CA1303981C - Polymer-coated optical structures - Google Patents
Polymer-coated optical structuresInfo
- Publication number
- CA1303981C CA1303981C CA000542873A CA542873A CA1303981C CA 1303981 C CA1303981 C CA 1303981C CA 000542873 A CA000542873 A CA 000542873A CA 542873 A CA542873 A CA 542873A CA 1303981 C CA1303981 C CA 1303981C
- Authority
- CA
- Canada
- Prior art keywords
- polymer
- diffraction grating
- ligand
- binding partner
- specific binding
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Expired - Lifetime
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Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/543—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals
- G01N33/54366—Apparatus specially adapted for solid-phase testing
- G01N33/54373—Apparatus specially adapted for solid-phase testing involving physiochemical end-point determination, e.g. wave-guides, FETS, gratings
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/543—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals
- G01N33/544—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals the carrier being organic
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/543—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals
- G01N33/551—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals the carrier being inorganic
- G01N33/553—Metal or metal coated
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y10—TECHNICAL SUBJECTS COVERED BY FORMER USPC
- Y10S—TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y10S436/00—Chemistry: analytical and immunological testing
- Y10S436/805—Optical property
Abstract
Abstract Polymer-coated optical structures A method of treating the surface of an optical structure which comprises forming on said surface a thin layer of organic polymer using a solvent casting technique. Preferably the process includes the subsequent treatment of the polymer layer with a solution of a specific ligand.
Complex formation between the bound ligand and its specific binding partner present in a sample to be analysed alters the optical properties of the diffraction grating surface and the change can form the basis of an assay method.
Complex formation between the bound ligand and its specific binding partner present in a sample to be analysed alters the optical properties of the diffraction grating surface and the change can form the basis of an assay method.
Description
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12W Sl 820 Polymer-coat~d optical structures The present invention relates to methods o producing polymer-coated surfaces suitable ~or use as optical structures. In particular it relates to methods of producing surfaces suitable for use in sensors, for example in biosensors in which one of a pair of binding partners is applied to the surface of a polymer-coated optical structure to form a device for detecting the presence of the complementary component in a sample subsequently brought into contact with the surface.
The properties of simple optical structures have been known for many years and structures such as~ for example, diffraction gratings have widespread lS use, not only as tools for understanding and analysing the wave properties of electromagnetic radiation but also, more recently, as sensors for detecting chemical, biochemical or biological species in a sample.
US-3926564 and US-3979184 both describe the adsocption of immunologically reactive proteins onto rough metallic surfaces such that binding of a complementary component changes the optical properties of the surface in a manner detectable 2S by the unaided eye.
EP-0112721 describes the preparation, and use as biosensors, of diffraction gratinqs coated with a material (hereinafter called "specific binding partnern) capable of binding with the species to be assayed (hereinafter called "ligand") to form a complex. The optical properties of such diffraction gratings are altered as a result of complex formation between the specific binding partner and the ligand and this phenomenon can consequently form the basis , , .
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of an assay system.
A problem common not only to the use of coated optical structures as biosensors but also to the use of standard optical structures by experimental physicists, is the dif~iculty of controlling their surface properties. The supporting substrate of such optical structures is often of glass or plastics material but the surface of the structure may comprise a thin metal layer formed, for example, by vacuum deposition. Problems can arise with the use of metal-coated surfaces in corrosive environments which may des~roy the integrity of a metal layer.
Other inorganic layers e.g. silicon oxide have also been described as diffraction grating surfaces (see, for example, EP-0112721 and EP-0178083).
Certain of the optical properties of an optical structure, for example its reflection and/or transmission properties and any surface plasmon resonance effect exhibited, will depend on the composition, thickness and uniformity of such surface layers; the composition of the surface layer also governs its chemical properties. However, the range of chemical and physical properties of known inorganic layers is limited. A still further problem which may occur when using such optical structures as biosensors is that some biologically active molecules, e.g.
proteins, may be at least partial}y inactivated by direct contact with metallic and certain inorganic surfaces.
In contrast, an extremely wide range of chemical and physical properties can be achieved by using appropriate organic polymers and a large number of techniques are known for bonding thereto or adsorbing thereon chemical, biochemical and biological entities~ Unfortunately, previous attempts to prepare polymer-coated difÇraction gratings have resulted in the polymer coating failing to con~orm "- ~3~3~
adequately to the surface relief profile of the yrating thereby grossly distorting its inherent physical properties.
We have now found that by employing a particular technique known as "solvent casting" it is possible to prepare an optical structure having a uniformly distributed surface layer of organic polymer which conforms well to the surface relief profile of the optical structure.
Thus, in its broadest aspect, the invention provides a method of preparing an optical structure useful for the detection of a ligand in a spectrophotometric assay technique, said optical structure having a surface relief profile or being capable of supporting surface plasmon resonance, which method comprises applying to the entire surface of the optical s~ructure a unifoxm, thin layer of an organic polymer by means of a solvent casting technique and absorbing on or binding to the thin layer of organic polymer, either directly or indirectly, a specific binding partner for the ligand.
In general, the layer of polymer applied to the surface of the grating will have a thickness in the range of 5 nanometers ~o 100 nanometers, preferably 15 to 20 nanometers. The polymer may be any suitable material which can be cast in a thin layer using a solvent casting technique.
The use of cellulose derivatives such as, for example, cellulose nitrate~ as the polymer layer is particularly pre~erred but other polymers may also be used, for example, those mentioned hereinafter.
As mentioned previously the optical characteristics of an optical surface depend not only on its physical relief profile but also on the composition of the ~3~
- 3a -surface layer or layers. Thus, the process of the prQsent invention can be applied to produce novel coated optical structures which may prove to be useful tools for experimental physicists and the like. In addition, certain preferred surface layers such as, for example, those capable of supporting surface plasmon resonance e.g. silver may be susceptible to corrosion by, for example, aqueous environments;
the present invention provides a means of passivating such a layer such that its integrity is maintained . .~.
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under practical working conditions.
However, the present invention is of particular advantage where the optical structures are diffraction gratings intended to be used as biosensors in a manner analogous to previous proposals (see, for example, EP-0112721). ~irect contact between a biologically active material, such as, for example, an antibody or antigen, and a metallic surface may result in contamination or destruction of its biological activity. In this situation the polymer layer-conveniently acts as a barrier. Furthermore, the wide range of chemical properties of different polymer surfaces ~e.g. availability of ~ree groups for covalent bonding, hydrophilicity or hydrophobicity, surface charge and dielectric properties) provide versatility and scope for optimising the binding or adsorption of any particular binding partner, such as, for example, an antibody or an antigen.
It is important to maintain the native conformation and biological activity of the desired binding partner and to obtain satisfactory immobilisation onto the surface of the optical structure whilst minimising any non-specific binding or steric hindrance.
The method of the present invention hence has particular use in the preparation of biosensors and thus according to one embodiment of the present invention there is provided a method of preparing a device for the detection of a ligand which comprises applying to the surface o~ an optical structure a thin layer of an organic polymer by means of a solvent casting technique and adsorbing on or binding to the said organic polymer, either directly or indirectly, a specific binding partner for the ligand it is desired to detect.
Biosensors produced in accordance with the invention are especially useful ~or the detection of antibodies or antigens (in which case the speci~ic .
.:
binding partner will be an antigen or an antibody, monoclonal or polyclonal, respectively) but other ligands may be detected by the use of other specific binding partners, as discussed hereinafter. However, S the sensors produced in accordance with the invention are not limited to biosensors and may comprise chemical sensors such as, for example, gaseous chemical detectors wherein specific adsorption onto or absorption into the polymer layer may occur.
The optical properties of an optical structure coated with a specific binding partner are altered by complex formation between the specific binding partner and the complementary ligand and, in one embodiment of the invention, by comparing the optical characteristics o~ a standard (untreated) region with those of a treated test region of the surface it is possible to determine qualitatively whether a binding reaction has occurred in the test region.
In an alternative embodiment where, for example, the specific binding partner is an antibody, an antibody specific to the antigen to be tested for i5 adsorbed on or bound to at least one discrete test region on the surface of an optical structure and a protein which is not a specific binding partner for any ligand which may be present in the sample to be tested (denoted herein as a "non-specific protein") is adsorbed on or bound to at least one discrete reference region on said surface. ~he non-specific protein may be, for example, an inactivated antibody or an antibody raised against a ligand not present in the samples to be tested e.g. where the sa~ples to be tested are human sera, the non-specific protein may be an anti-mouse antibody. Any differences between the non-specific binding of e.g. proteins present in the test sample to either the specific antibody or the non-specific protein can be determined by comparing the optical properties o~ the test r~
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region with the reference region after exposure to a sample which does not contain the antigen to be tested for, so that an appropriate correction can, if necessary, be made. Comparison of the optical characteristics of the test and standard regions of a similar biosensor during or after exposure to a sample to be tested can then provide a measurement of complex formation between the antigen to be tested for and its specific antibody.
Each region may comprise a continuous layer of a specific binding partner or each binding partner may be present at discrete intervals within any given region to form a discontinuous layer.
The biosensors produced in accordance with the invention may, for example, be used in assays in ways analogous to those described in EP-0112721 and EP-0178083.
Thus according to a further feature of the present invention there is provided a method of detecting a ligand in a sample which comprises contacting said sample with a biosensor prepared by the process hereinbefore defined and determining whether, and if desired the extent to which and/or rate at which, an optical characteristic of the biosensor is altered by formation of a complex between the ligand and the specific binding partner.
The present invention further provides a biosensor eor detecting a ligand comprising an optical structure bearing a thin layer of an organic polymer, which has been applied to the surface of the optical structure by a solvent casting technique, the said organic polymer having adsorbed thereon or bound thereto, either directly or indirectly, a specific binding partner for the ligand it is desired to detect.
Solvent casting of polymers onto solid or liquid sur~aces is a well known technique, reviewed .
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by Mano and Durao, Journal of Chemical Education (1973), 50, 22~3. The article entitled "On the Technique of Making Thin Celluloid Films" ~y James Taylor, published in the Journal of Scienti~ic Instruments for 1925 to 1926, Volume 3, pages 400-40~, describes the formation on a water surEace oE celluloid Eilms having thicknesses of the order of 50 nm.
The solvent casting technique ~referably used in accordance with the invention involves the ormation of a thin polymer ~ilm on a liquid surface using a solvent, and subsequently causing relative movement between the surface of the optical structure on which the film is to be formed and the thin polymer film on the liquid surface such that they are brought into contact. The thickness of the film on the liquid sur~ace will to a large extent determine the thickness of the film on the surace of the optical structure, and the thickness of the film on the liquid surEace will itself be governed by the quantity and the nature of the polymer used, the surface area and nature of the casting liquid (the term "casting liquid" as used herein refers to the bulk liquid on the surface of which the polymer Eilm is cast). Thus, ~or example, one drop Erom a capillary tube o a mixture of 3 g cellulose nitrate in 100 ml of amyl acetate can be made to spread out on a sufeiciently large water surface to give a film of cellulose nitrate o only a few nanometers thickness.
As an alternative to solvent casting techniques involving a casting liquid, in certain circumstances it may be possible to cast the Eilm directly onto the surEace oE the optical structure itselE or onto another solid surEace Eollowed by transEer of the Eilm to the surEace of the optical structure.
The invention will now he described in more detail with reEerence to a oreEerred embodiment -- ~3~3g~
wherein the optical structure is a diffraction grating. ~owever, it is to be understood that other optical structures such as, for example, op~ical waveguides, optical fibres and metal-coated prisms in the correct configuration to exhibit surface plasmon resonance, are all included within the scope of the invention.
As previously mentioned, the supporting substrate of a diffraction grating may comprise glass or plastics material; polycarbonate is particularly preferred. The diffraction grating may be formed in the surface of the supporting substrate which may be covered with, for example, a metal film, such as silver, gold, copper, nickel or aluminium, formed by a process of vapour deposition, electroplating, sputtering, coating, painting, spraying or otherwiseO
Preferably the metal surface is capable of supporting surface plasmon resonance, and more particularly is of silver, and the metallic film is deposited by vacuum deposition.
Where the diffraction grating is to be used in an optical transmission mode, the metal film must be sufficiently thin, for example up to 50 nm for silver, so as not to unduly impede the passage of light therethrough. Where the diffraction grating is to be used in a reflective mode, then the metal layer can be thicker, for example up to S00 nm, preferably around 100 nm for silver, and is preferably made sufficiently dense to provide a good reflecting surface on the diffraction grating pattern in the surface of the supporting substrate e.g. of plastics or glass.
A particularly preferred method of applying a polymer film to the surface of a metallised diffraction grating comprises the steps of:-1. inserting a metallised polycarbonate diffraction grating into a bath containlng the casting , .: . ' :. ' , ': . ' ' -': ' , :' '~ ' ' ' ~3~
liquid, preferably de-ionised distilled water, 2. forming a thin film of the desired polymer over the surface of the casting liquid, and 3. either raising the diffraction grating through the casting liquid to engage the film of polymer on the surface of the liquid or allowing the castin~ liquid to drain out of the bath so that the polymer film on the surface of the liquid drops as the liquid level drops until it engages and spreads over the surface of the metallised diffraction grating, in either case the diffraction grating being located at a small angle to the horizontal to facilitate draining of the casting liquid from the surface of the diffraction grating.
Where the diffraction grating is intended to be used as a biosensor as described above the solution of the specific binding partner is preferably left in contact with the polymer for a controlled period of time and, where appropriate, under controlled conditions of temperature, pressure, humidity and the like. The process will normally have to he performed under "clean room" conditions and, particularly where the binding partner is a biologically active substance, all the usual precautions must be taken for handling such materials in order to avoid contam-ination and denaturation or inactivation in any way.
Any surplus solution is preferably removed from the polymer surface and may be recovered by suction or washing and recycled or simply washed away as waste.
The final coated di~fraction grating may be dried or otherwise treated and packaged, again generally under ~Iclean room" conditions.
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Techniques for bonding specific binding partners to solid phase supports are described in the literature~
The binding partner may be bound to the polymer either directly or indirectly. Indirect binding S may, for example, be effected by binding to the polymer a reagent Y which selectively interacts with a reagent Z provided on the specific binding partner~ In such cases, the reagent Z may for example be such as to render the specific binding partner antigenic to reagent Y which in that case will be an antibody raised to Z. Z may for example be fluorescein isothiocyanate, rhodamine isothiocyanate, 2,4-dinitrofluorobenzene, phenyl isothiocyanate or dansyl chloride. Reagents Y and Z may alternatively be a specific binding protein and the corresponding ligand such as for example avidin and biotin.
rt is a desirable but not an essential feature of the invention to provide covalent bonding of the binding partner to the polym~r.
The bonding of the specific binding partner, either directly or indicectly, to the polymer may be facilitated by activating the polymer layer, to provide free reactive groups for bonding. Thus, for example, cellulose nitrate can be activated by cyanogen bromide in a~ueous solution of p~eferred pH 10-11. Typically, the cellulose surface is left in contact with the cyanogen bromide solution for a controlled period of time, typically of the order of a few minutes, whilst pH is controlled within the limits indicated above. After this reaction the diffraction grating is removed from the cyanogen bromide solution and washed in cold distilled water and thereafter is optionally dried and stored pending the application theeeto of a binding paetner, such as, for example, an antibody or an antigenic peotein.
.
, .
'' ' The binding of a specific binding partner to an activated polymer surface may be conveniently effected by contacting a solution of the specific binding partner with the polymer surface such that, where the polymer has been activated by the provision of reactive groups as described above, the binding partner may attach to or associate with the reactive groups on the polymer surface. Typically droplets of the solution may be app~ied to the polymer surface and left for a controlled period of time (typically of the order of some hours) in a controlled environment (particularly a humidity-controlled environment to prevent drying) and in clean room conditions, so that a sufficient quantity of the binding partner becomes bound to the surface of the polymer.
However, the use of cyanogen bromide may not be appropriate if the material which is to be bonded to the polymer is other than a protein, such as, for example, an antigenic polysaccharide.
The invention has wide application and merely as one example may be used in the detection of the Group A Streptococcal bacteria or bacterial antigen in childhood illnesses. However, it is to be understood that the coated diffraction gratings and biosensor diagnostic testing devices produced by the invention are not in any way li~ited to such an application and can be used for any diagnostic test provided the appropriate binding partner has been applied to the surface of the diffraction grating.
The invention will be particularly described hereinafter with reference to an antibody or an antigen as the specific binding partner or li~and.
Howevec, the invention is not to be taken as being limited to methods of preparing biosensors suitable for use in antibody or antigen assays but includes wlthin its scope any sensors prepsred by the process ' ' , .
3~
l2 -of the invention which can be used to detect any chemical, biochemical or biological species in a sample. Examples of suitable binding partners which may be immobilised on an optical structure prepared by the process of the invention and of ligands which may be assayed by the rnethod of the invention are given in Table I helow.
Table I
Ligand Speci~ic Binding Partner . _ _ _ _ antigen specific antibody antibody antigen hormone hormone receptor hormone receptor hormone polynucleotide complementary polynucleotide strand strand avidin biotin biotin avidin protein A immunoglobulin immunoglobulin protein A
enzyme enzyme co~actor (substrate) or inhihitor enzyme cofactor enzyrne tsubstrate) or inhibi~or lectins specific carbohydrate specific carbohydrate lectins of lectins .
The method o~ the invention has very broad applicability but in particular may be used to provide biosensors suitable ~or assaying: hormones, including peptide hormones (e.q. thyroi~ stirnulatin~
. , ~. , 3~3~
hormone (TSH), luteinising hormone (L,H), human chorionic gonadotrophin (hC~,), follicle stimulating hormone (FSH), insulin and prolactin) and non-peptide hormones (e.g. steroid hormones such as cortisol, estradiol, progesterone and testosterone, or thyroid hormones such as thyroxine (T4) and triiodothyronine), ~roteins (e.g. carcinoembryonic antigen (CEA) and alphafetoprotein (AFP)), drugs (e.g. digoxin), sugars, toxins, vitamins, viruses, bacteria o~
microorganisms.
It will be understood that the term "antibody"
used herein includes within its scope a) any of the various classes or sub-classes of immunoglobulin, e.g. IgG, IgM, derived from any of the animals conventionally used, e.g. sheep, rabbits, goats or mice, b) monoclonal antibodies, c) intact molecules or l'fragments" of antibodies, monoclonal or polyclonal, the fragments being those which contain the binding reqion of the antibody, i.e. fragments devoid of the Fc portion (e.g., Fab, Fab', F(ab')~) or the so-called "half-molecule" fragments obtained by reductive cleavage of the disulphide bonds connecting the heavy chain components in the intact antibody.
The method of preparation of fragments of antibodies is well known in the art and will not be described herein.
The term "antigen" as used herein will be understood to include both permanently antigenic species (for example, proteins, bacteria, bacteria fragments, cells, cell fragments and viruses) and haDtens which may be rendered antigenic under suitab1e conditions.
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The ~ollowing examples describe the preparation of cellulose nitrate ~ilms and their subsequent transfer onto diffraction gratings and the binding or adsorptlon o~ biological materials onto the sur~ace of such polymer-coated gratings.
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- lS -Preparation o Cellulose Nitrate Films and Transfer onto Diffraction Grating All solutio,1s and casting liquids used for the preparation of polymer films and their transfer onto di~raction gratings should be filtered to remove contaminating oarticles and all operations should be carried out in a clean room.
In a preferred method, a casting tank is chosen such that the area of the water surface is larger (by a factor of S or more) than the area o the grating to be covered. This ensures that the diffraction grating is not covered by an edge of the cast polymer film. Where a very thin layer -of polymer is re~uired the surface area of the casting tank is preferably sufficiently larger than the area of the cast film to avoid any "edge effect" e.g. five times larger. However, for thicker films it may be oreferred to cast the polymer into a ring on the surface on the casting liquid, the area of which can be readily adjusted to control the thickness of the polymer layer.
The tank is cleaned, particularly to remove grease, surfactants or other contaminants, and a diraction grating support is placed at the bottom of the tank. This support may, for example, comprise a wice grid located at a small angle to the horizontal to allow water to drain between the cast ~iLm and the diffraction grating as the two are brought into contact.
The tank is~ilIed with distilled water.
It may be advantageous but not essential to adjust the pH or ionic strength of this water in order to promote good adhesion of~the polymer ~ilm to the dif~raction grating. The water level must be above the surface Oe the diffraction gratings which are to rest on the support.
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- l6 Di~fraction gratings are place~ on the support before or after addition of water, grating side up. The diffraction gratings should be clean and free of grease, surEactants or other contaminants before enterinq the castin~ tank.
Once the water surface has settled a drop of cellulose nitrate solution, Eor example cellulose nitrate in amyi acetate, is dropped onto the centre of the water surface. Provided the water surEace is Eree of surfactants, the cellulose nitrate solution spreads to a well defined circular area and the liquid film dries within a few seconds. The rate of drying of the film may be controlled by covering the castinq tank with a lid provided with a s~all hole in the centre through which the cellulose solution is applied. With a uniform film thickness, reflected light interference colours may be observed as the film dries successively thinner.
If the water surface is not clean, the thickness of the cellulose film formed is not uniform and it spreads out into a non-circular patch with 'fingers' extending outwards. The non-uniform film thickness is apparent from the random distribution of reflected interference colours.
A polymer film thickness of about 16 nanometers is produced by addition of lO microlitres of a 9% cellulose nitrate solution (9 g cellulose nitrate in 100 ml amyl acetate).
Once the film has dried, water is drained out through a tap at the bottom of the tank, such that the dried polymer film is lowered onto the diffraction grating surface. The difEraction grating and its support are gently removed Erom the tank and left to dry Eor several minutes.
~e have Eound that the a~hesion of cellulose nitrate to silver-coated polycarbonate diE~raction qratin~s is enllanced ~v castin~ the ~olymer Eilms :
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onto distilled water whose p~ has been ad~usted to 9 by the addition of, for example, NaOH.
The use of any other solvent castable polymers is Possible with the correct choice ~, solvents and casting surfaces. Thus, for example, cellulose acetate films may be cast from amyl acetate, polystyrene from xylene, ~olyisobutylene ~rom petroleum ether and polyvinyl chloride - acetate copolymer ~ilms can be cast From cyclohexanone onto water surfaces.
In some circumstances enforced spreading of the solution on the casting liquid, using eonventional teehniques, may be neeessary.
Film thickness may be eontrolled by eare~ul seleetion of:
- polymer solution eoneentration - volume o~ material aoplied to the casting liquid sur~aee - type of solvent.
Amyl acetate ~or example is amphiphilic and spreads rapidly over a clean water surface. Other solvents may not spread as rapidly, with coneomitant increases in film thickness.
The technique is not limited to casting on a water sur~ace and other liquids may be used for the casting surface provided the solvent used is largely immiseible with, and the polymer-eontaining solution is o~ lower density than, the casting liquid. Thus, for example, ethyl eellulose ean be east onto mercury, diisocyanates together with polyols can be co-cast onto mercury and gelatin can be cast from water onto carbon tetrachloride.
It is also possible to produce thin polymer layers using the solvent casting technique to cast the polymer onto a solid surface. Thus, for example, polyethylene terephthalate can be cast from trifluoro-acetic acid, polycarbonate from dichloromethane, ~olyvinyl formal from dioxan, polysulfone Ero~
.
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chloroform, cellulo~se acetate ~rom acetone and polypropylene can be cast ~rom xylene, in each case on to an appropriate substrate. It ;s to be understood that where the polymer is cast onto a cast.lg liquid the casting liquid must be compati~le with the material of the di~raction grating immersed therein and where the polymer is cast onto a solid sur~ace the solvent used to bring the polymer into solution prior to casting must he compati~le ~ith the material of the solid sur~ace.
Bindinq of Bioloqical Materials to the Polymer Surface Three different methods of couplinq biological lS molecules to a cellulose nitrate polymer surface on a diffraction grating, have been considered:
a) covalent bonding b) non-covalent adsorption c) covalent bonding of the biological molecule to a pol~mer which is entrapped within the cellulose nitrate.
a) Covalent ~onding Established ~rocedures exist ~or covalently bonding biological materials, such as proteins, to cellulosic material. Published procedures include the use of chemical reagents which modify the polymer unit to form reactive groups which covalently bind to typical protein groups such as, ~or example, free amino groups.
Commonly used chemical reagents include:
- cyanogen bromicle - tosyl chloride - ~ titanium complexes - carbodiimide - cyanurlc chloride `
' .
~r . 13~3~8~
-- - oxirane - periodate.
b) Non-Covalent Adsorption Biological molecules, for example, sheep serum proteins, eibonucleases and immunoglobulins, have been found to hind very efficiently to cellulose nitrate. ~imple addition o~ an aqueous protein solution at room temperature or Lower has resulted in bound protein which could not easily be removed by washing with water, buffer solutions or detergents.
For example, incubation of a cellulose nitrate-coated grating for 2 hours at room temperature in lO mgml l bovine ~-globulin, 50 mM NaHCO3, pH 9, resulted in extensive coverage of the diffraction grating with the protein. Various treatments, such as, eor example, with mild detergent (5~ Tween*
20), buffer (50 mM NaHCO3), n heptane, and vH cycling between pH 2 and 10.6 did not affect the binding of the protein. Stronger detergent treatment (e.g.
2% SDS) has been found to damage the cellulose nitrate film.
c) Reactive PolYmer Entrapment This procedure separates the two important functions of the diffraction grating layer - adhesion and conformation to the grating surface - reactivity towards protein or other bioloqical material or binding partner and meets each requirement with a separate polymer.
A monomeric material is allowe~ to difeuse into the polymer aefixed to the dif~raction qratinq.
The monomer is then polymerised to give a network of the second polymer entangled in the first, a~Eixed * TRADE MARK
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polymer. The second polymer is chosen to have chemical groups that are reactive towards protein or other biological molecules.
Thus, ~or example, dif~raction gratings coated with cellulose nitrate according to the ~rocedure described herein are incubated with 1~ aqueous glutaraldehyde monomer solution pH 5.5 for 30 minutes at room temperature. The p~ is then adjusted to 10, and the diffraction gratings left for ~0 minutes to allow polymerisation of the glutaraldehyde.
Polymerisation may be monitored by optical absorbance measurements at 235 nm. Such techniques have been previously described, see for example, US-4001583.
Free reactive carbonyl groups in the polyglutaraldehyde are thus exposed at the surface of the polymer layer.
Treatment of these diffraction gratings with protein solutions, e.g. 5 mgml 1 bovine ~-globulin or 5 mgml 1 ribonuclease, at 4C overnight in 50 mM
NaHCO3 pH 9, results in strong binding between the protein and the polyglutaraldehyde, believed to be the result of covalent bonding between the polyglutaraldehyde carbonyl groups and the amino groups of the proteins.
~, . . .
12W Sl 820 Polymer-coat~d optical structures The present invention relates to methods o producing polymer-coated surfaces suitable ~or use as optical structures. In particular it relates to methods of producing surfaces suitable for use in sensors, for example in biosensors in which one of a pair of binding partners is applied to the surface of a polymer-coated optical structure to form a device for detecting the presence of the complementary component in a sample subsequently brought into contact with the surface.
The properties of simple optical structures have been known for many years and structures such as~ for example, diffraction gratings have widespread lS use, not only as tools for understanding and analysing the wave properties of electromagnetic radiation but also, more recently, as sensors for detecting chemical, biochemical or biological species in a sample.
US-3926564 and US-3979184 both describe the adsocption of immunologically reactive proteins onto rough metallic surfaces such that binding of a complementary component changes the optical properties of the surface in a manner detectable 2S by the unaided eye.
EP-0112721 describes the preparation, and use as biosensors, of diffraction gratinqs coated with a material (hereinafter called "specific binding partnern) capable of binding with the species to be assayed (hereinafter called "ligand") to form a complex. The optical properties of such diffraction gratings are altered as a result of complex formation between the specific binding partner and the ligand and this phenomenon can consequently form the basis , , .
- ~3~39~
of an assay system.
A problem common not only to the use of coated optical structures as biosensors but also to the use of standard optical structures by experimental physicists, is the dif~iculty of controlling their surface properties. The supporting substrate of such optical structures is often of glass or plastics material but the surface of the structure may comprise a thin metal layer formed, for example, by vacuum deposition. Problems can arise with the use of metal-coated surfaces in corrosive environments which may des~roy the integrity of a metal layer.
Other inorganic layers e.g. silicon oxide have also been described as diffraction grating surfaces (see, for example, EP-0112721 and EP-0178083).
Certain of the optical properties of an optical structure, for example its reflection and/or transmission properties and any surface plasmon resonance effect exhibited, will depend on the composition, thickness and uniformity of such surface layers; the composition of the surface layer also governs its chemical properties. However, the range of chemical and physical properties of known inorganic layers is limited. A still further problem which may occur when using such optical structures as biosensors is that some biologically active molecules, e.g.
proteins, may be at least partial}y inactivated by direct contact with metallic and certain inorganic surfaces.
In contrast, an extremely wide range of chemical and physical properties can be achieved by using appropriate organic polymers and a large number of techniques are known for bonding thereto or adsorbing thereon chemical, biochemical and biological entities~ Unfortunately, previous attempts to prepare polymer-coated difÇraction gratings have resulted in the polymer coating failing to con~orm "- ~3~3~
adequately to the surface relief profile of the yrating thereby grossly distorting its inherent physical properties.
We have now found that by employing a particular technique known as "solvent casting" it is possible to prepare an optical structure having a uniformly distributed surface layer of organic polymer which conforms well to the surface relief profile of the optical structure.
Thus, in its broadest aspect, the invention provides a method of preparing an optical structure useful for the detection of a ligand in a spectrophotometric assay technique, said optical structure having a surface relief profile or being capable of supporting surface plasmon resonance, which method comprises applying to the entire surface of the optical s~ructure a unifoxm, thin layer of an organic polymer by means of a solvent casting technique and absorbing on or binding to the thin layer of organic polymer, either directly or indirectly, a specific binding partner for the ligand.
In general, the layer of polymer applied to the surface of the grating will have a thickness in the range of 5 nanometers ~o 100 nanometers, preferably 15 to 20 nanometers. The polymer may be any suitable material which can be cast in a thin layer using a solvent casting technique.
The use of cellulose derivatives such as, for example, cellulose nitrate~ as the polymer layer is particularly pre~erred but other polymers may also be used, for example, those mentioned hereinafter.
As mentioned previously the optical characteristics of an optical surface depend not only on its physical relief profile but also on the composition of the ~3~
- 3a -surface layer or layers. Thus, the process of the prQsent invention can be applied to produce novel coated optical structures which may prove to be useful tools for experimental physicists and the like. In addition, certain preferred surface layers such as, for example, those capable of supporting surface plasmon resonance e.g. silver may be susceptible to corrosion by, for example, aqueous environments;
the present invention provides a means of passivating such a layer such that its integrity is maintained . .~.
~,....
~ d,i~
~3~3~
under practical working conditions.
However, the present invention is of particular advantage where the optical structures are diffraction gratings intended to be used as biosensors in a manner analogous to previous proposals (see, for example, EP-0112721). ~irect contact between a biologically active material, such as, for example, an antibody or antigen, and a metallic surface may result in contamination or destruction of its biological activity. In this situation the polymer layer-conveniently acts as a barrier. Furthermore, the wide range of chemical properties of different polymer surfaces ~e.g. availability of ~ree groups for covalent bonding, hydrophilicity or hydrophobicity, surface charge and dielectric properties) provide versatility and scope for optimising the binding or adsorption of any particular binding partner, such as, for example, an antibody or an antigen.
It is important to maintain the native conformation and biological activity of the desired binding partner and to obtain satisfactory immobilisation onto the surface of the optical structure whilst minimising any non-specific binding or steric hindrance.
The method of the present invention hence has particular use in the preparation of biosensors and thus according to one embodiment of the present invention there is provided a method of preparing a device for the detection of a ligand which comprises applying to the surface o~ an optical structure a thin layer of an organic polymer by means of a solvent casting technique and adsorbing on or binding to the said organic polymer, either directly or indirectly, a specific binding partner for the ligand it is desired to detect.
Biosensors produced in accordance with the invention are especially useful ~or the detection of antibodies or antigens (in which case the speci~ic .
.:
binding partner will be an antigen or an antibody, monoclonal or polyclonal, respectively) but other ligands may be detected by the use of other specific binding partners, as discussed hereinafter. However, S the sensors produced in accordance with the invention are not limited to biosensors and may comprise chemical sensors such as, for example, gaseous chemical detectors wherein specific adsorption onto or absorption into the polymer layer may occur.
The optical properties of an optical structure coated with a specific binding partner are altered by complex formation between the specific binding partner and the complementary ligand and, in one embodiment of the invention, by comparing the optical characteristics o~ a standard (untreated) region with those of a treated test region of the surface it is possible to determine qualitatively whether a binding reaction has occurred in the test region.
In an alternative embodiment where, for example, the specific binding partner is an antibody, an antibody specific to the antigen to be tested for i5 adsorbed on or bound to at least one discrete test region on the surface of an optical structure and a protein which is not a specific binding partner for any ligand which may be present in the sample to be tested (denoted herein as a "non-specific protein") is adsorbed on or bound to at least one discrete reference region on said surface. ~he non-specific protein may be, for example, an inactivated antibody or an antibody raised against a ligand not present in the samples to be tested e.g. where the sa~ples to be tested are human sera, the non-specific protein may be an anti-mouse antibody. Any differences between the non-specific binding of e.g. proteins present in the test sample to either the specific antibody or the non-specific protein can be determined by comparing the optical properties o~ the test r~
~3~3~18~
region with the reference region after exposure to a sample which does not contain the antigen to be tested for, so that an appropriate correction can, if necessary, be made. Comparison of the optical characteristics of the test and standard regions of a similar biosensor during or after exposure to a sample to be tested can then provide a measurement of complex formation between the antigen to be tested for and its specific antibody.
Each region may comprise a continuous layer of a specific binding partner or each binding partner may be present at discrete intervals within any given region to form a discontinuous layer.
The biosensors produced in accordance with the invention may, for example, be used in assays in ways analogous to those described in EP-0112721 and EP-0178083.
Thus according to a further feature of the present invention there is provided a method of detecting a ligand in a sample which comprises contacting said sample with a biosensor prepared by the process hereinbefore defined and determining whether, and if desired the extent to which and/or rate at which, an optical characteristic of the biosensor is altered by formation of a complex between the ligand and the specific binding partner.
The present invention further provides a biosensor eor detecting a ligand comprising an optical structure bearing a thin layer of an organic polymer, which has been applied to the surface of the optical structure by a solvent casting technique, the said organic polymer having adsorbed thereon or bound thereto, either directly or indirectly, a specific binding partner for the ligand it is desired to detect.
Solvent casting of polymers onto solid or liquid sur~aces is a well known technique, reviewed .
~3~39~3~
by Mano and Durao, Journal of Chemical Education (1973), 50, 22~3. The article entitled "On the Technique of Making Thin Celluloid Films" ~y James Taylor, published in the Journal of Scienti~ic Instruments for 1925 to 1926, Volume 3, pages 400-40~, describes the formation on a water surEace oE celluloid Eilms having thicknesses of the order of 50 nm.
The solvent casting technique ~referably used in accordance with the invention involves the ormation of a thin polymer ~ilm on a liquid surface using a solvent, and subsequently causing relative movement between the surface of the optical structure on which the film is to be formed and the thin polymer film on the liquid surface such that they are brought into contact. The thickness of the film on the liquid sur~ace will to a large extent determine the thickness of the film on the surace of the optical structure, and the thickness of the film on the liquid surEace will itself be governed by the quantity and the nature of the polymer used, the surface area and nature of the casting liquid (the term "casting liquid" as used herein refers to the bulk liquid on the surface of which the polymer Eilm is cast). Thus, ~or example, one drop Erom a capillary tube o a mixture of 3 g cellulose nitrate in 100 ml of amyl acetate can be made to spread out on a sufeiciently large water surface to give a film of cellulose nitrate o only a few nanometers thickness.
As an alternative to solvent casting techniques involving a casting liquid, in certain circumstances it may be possible to cast the Eilm directly onto the surEace oE the optical structure itselE or onto another solid surEace Eollowed by transEer of the Eilm to the surEace of the optical structure.
The invention will now he described in more detail with reEerence to a oreEerred embodiment -- ~3~3g~
wherein the optical structure is a diffraction grating. ~owever, it is to be understood that other optical structures such as, for example, op~ical waveguides, optical fibres and metal-coated prisms in the correct configuration to exhibit surface plasmon resonance, are all included within the scope of the invention.
As previously mentioned, the supporting substrate of a diffraction grating may comprise glass or plastics material; polycarbonate is particularly preferred. The diffraction grating may be formed in the surface of the supporting substrate which may be covered with, for example, a metal film, such as silver, gold, copper, nickel or aluminium, formed by a process of vapour deposition, electroplating, sputtering, coating, painting, spraying or otherwiseO
Preferably the metal surface is capable of supporting surface plasmon resonance, and more particularly is of silver, and the metallic film is deposited by vacuum deposition.
Where the diffraction grating is to be used in an optical transmission mode, the metal film must be sufficiently thin, for example up to 50 nm for silver, so as not to unduly impede the passage of light therethrough. Where the diffraction grating is to be used in a reflective mode, then the metal layer can be thicker, for example up to S00 nm, preferably around 100 nm for silver, and is preferably made sufficiently dense to provide a good reflecting surface on the diffraction grating pattern in the surface of the supporting substrate e.g. of plastics or glass.
A particularly preferred method of applying a polymer film to the surface of a metallised diffraction grating comprises the steps of:-1. inserting a metallised polycarbonate diffraction grating into a bath containlng the casting , .: . ' :. ' , ': . ' ' -': ' , :' '~ ' ' ' ~3~
liquid, preferably de-ionised distilled water, 2. forming a thin film of the desired polymer over the surface of the casting liquid, and 3. either raising the diffraction grating through the casting liquid to engage the film of polymer on the surface of the liquid or allowing the castin~ liquid to drain out of the bath so that the polymer film on the surface of the liquid drops as the liquid level drops until it engages and spreads over the surface of the metallised diffraction grating, in either case the diffraction grating being located at a small angle to the horizontal to facilitate draining of the casting liquid from the surface of the diffraction grating.
Where the diffraction grating is intended to be used as a biosensor as described above the solution of the specific binding partner is preferably left in contact with the polymer for a controlled period of time and, where appropriate, under controlled conditions of temperature, pressure, humidity and the like. The process will normally have to he performed under "clean room" conditions and, particularly where the binding partner is a biologically active substance, all the usual precautions must be taken for handling such materials in order to avoid contam-ination and denaturation or inactivation in any way.
Any surplus solution is preferably removed from the polymer surface and may be recovered by suction or washing and recycled or simply washed away as waste.
The final coated di~fraction grating may be dried or otherwise treated and packaged, again generally under ~Iclean room" conditions.
, . : .
. . .
- '' , '.
~3~39~
Techniques for bonding specific binding partners to solid phase supports are described in the literature~
The binding partner may be bound to the polymer either directly or indirectly. Indirect binding S may, for example, be effected by binding to the polymer a reagent Y which selectively interacts with a reagent Z provided on the specific binding partner~ In such cases, the reagent Z may for example be such as to render the specific binding partner antigenic to reagent Y which in that case will be an antibody raised to Z. Z may for example be fluorescein isothiocyanate, rhodamine isothiocyanate, 2,4-dinitrofluorobenzene, phenyl isothiocyanate or dansyl chloride. Reagents Y and Z may alternatively be a specific binding protein and the corresponding ligand such as for example avidin and biotin.
rt is a desirable but not an essential feature of the invention to provide covalent bonding of the binding partner to the polym~r.
The bonding of the specific binding partner, either directly or indicectly, to the polymer may be facilitated by activating the polymer layer, to provide free reactive groups for bonding. Thus, for example, cellulose nitrate can be activated by cyanogen bromide in a~ueous solution of p~eferred pH 10-11. Typically, the cellulose surface is left in contact with the cyanogen bromide solution for a controlled period of time, typically of the order of a few minutes, whilst pH is controlled within the limits indicated above. After this reaction the diffraction grating is removed from the cyanogen bromide solution and washed in cold distilled water and thereafter is optionally dried and stored pending the application theeeto of a binding paetner, such as, for example, an antibody or an antigenic peotein.
.
, .
'' ' The binding of a specific binding partner to an activated polymer surface may be conveniently effected by contacting a solution of the specific binding partner with the polymer surface such that, where the polymer has been activated by the provision of reactive groups as described above, the binding partner may attach to or associate with the reactive groups on the polymer surface. Typically droplets of the solution may be app~ied to the polymer surface and left for a controlled period of time (typically of the order of some hours) in a controlled environment (particularly a humidity-controlled environment to prevent drying) and in clean room conditions, so that a sufficient quantity of the binding partner becomes bound to the surface of the polymer.
However, the use of cyanogen bromide may not be appropriate if the material which is to be bonded to the polymer is other than a protein, such as, for example, an antigenic polysaccharide.
The invention has wide application and merely as one example may be used in the detection of the Group A Streptococcal bacteria or bacterial antigen in childhood illnesses. However, it is to be understood that the coated diffraction gratings and biosensor diagnostic testing devices produced by the invention are not in any way li~ited to such an application and can be used for any diagnostic test provided the appropriate binding partner has been applied to the surface of the diffraction grating.
The invention will be particularly described hereinafter with reference to an antibody or an antigen as the specific binding partner or li~and.
Howevec, the invention is not to be taken as being limited to methods of preparing biosensors suitable for use in antibody or antigen assays but includes wlthin its scope any sensors prepsred by the process ' ' , .
3~
l2 -of the invention which can be used to detect any chemical, biochemical or biological species in a sample. Examples of suitable binding partners which may be immobilised on an optical structure prepared by the process of the invention and of ligands which may be assayed by the rnethod of the invention are given in Table I helow.
Table I
Ligand Speci~ic Binding Partner . _ _ _ _ antigen specific antibody antibody antigen hormone hormone receptor hormone receptor hormone polynucleotide complementary polynucleotide strand strand avidin biotin biotin avidin protein A immunoglobulin immunoglobulin protein A
enzyme enzyme co~actor (substrate) or inhihitor enzyme cofactor enzyrne tsubstrate) or inhibi~or lectins specific carbohydrate specific carbohydrate lectins of lectins .
The method o~ the invention has very broad applicability but in particular may be used to provide biosensors suitable ~or assaying: hormones, including peptide hormones (e.q. thyroi~ stirnulatin~
. , ~. , 3~3~
hormone (TSH), luteinising hormone (L,H), human chorionic gonadotrophin (hC~,), follicle stimulating hormone (FSH), insulin and prolactin) and non-peptide hormones (e.g. steroid hormones such as cortisol, estradiol, progesterone and testosterone, or thyroid hormones such as thyroxine (T4) and triiodothyronine), ~roteins (e.g. carcinoembryonic antigen (CEA) and alphafetoprotein (AFP)), drugs (e.g. digoxin), sugars, toxins, vitamins, viruses, bacteria o~
microorganisms.
It will be understood that the term "antibody"
used herein includes within its scope a) any of the various classes or sub-classes of immunoglobulin, e.g. IgG, IgM, derived from any of the animals conventionally used, e.g. sheep, rabbits, goats or mice, b) monoclonal antibodies, c) intact molecules or l'fragments" of antibodies, monoclonal or polyclonal, the fragments being those which contain the binding reqion of the antibody, i.e. fragments devoid of the Fc portion (e.g., Fab, Fab', F(ab')~) or the so-called "half-molecule" fragments obtained by reductive cleavage of the disulphide bonds connecting the heavy chain components in the intact antibody.
The method of preparation of fragments of antibodies is well known in the art and will not be described herein.
The term "antigen" as used herein will be understood to include both permanently antigenic species (for example, proteins, bacteria, bacteria fragments, cells, cell fragments and viruses) and haDtens which may be rendered antigenic under suitab1e conditions.
~3~3~
The ~ollowing examples describe the preparation of cellulose nitrate ~ilms and their subsequent transfer onto diffraction gratings and the binding or adsorptlon o~ biological materials onto the sur~ace of such polymer-coated gratings.
' 13~?3~8~L
- lS -Preparation o Cellulose Nitrate Films and Transfer onto Diffraction Grating All solutio,1s and casting liquids used for the preparation of polymer films and their transfer onto di~raction gratings should be filtered to remove contaminating oarticles and all operations should be carried out in a clean room.
In a preferred method, a casting tank is chosen such that the area of the water surface is larger (by a factor of S or more) than the area o the grating to be covered. This ensures that the diffraction grating is not covered by an edge of the cast polymer film. Where a very thin layer -of polymer is re~uired the surface area of the casting tank is preferably sufficiently larger than the area of the cast film to avoid any "edge effect" e.g. five times larger. However, for thicker films it may be oreferred to cast the polymer into a ring on the surface on the casting liquid, the area of which can be readily adjusted to control the thickness of the polymer layer.
The tank is cleaned, particularly to remove grease, surfactants or other contaminants, and a diraction grating support is placed at the bottom of the tank. This support may, for example, comprise a wice grid located at a small angle to the horizontal to allow water to drain between the cast ~iLm and the diffraction grating as the two are brought into contact.
The tank is~ilIed with distilled water.
It may be advantageous but not essential to adjust the pH or ionic strength of this water in order to promote good adhesion of~the polymer ~ilm to the dif~raction grating. The water level must be above the surface Oe the diffraction gratings which are to rest on the support.
.~ --- , :
- l6 Di~fraction gratings are place~ on the support before or after addition of water, grating side up. The diffraction gratings should be clean and free of grease, surEactants or other contaminants before enterinq the castin~ tank.
Once the water surface has settled a drop of cellulose nitrate solution, Eor example cellulose nitrate in amyi acetate, is dropped onto the centre of the water surface. Provided the water surEace is Eree of surfactants, the cellulose nitrate solution spreads to a well defined circular area and the liquid film dries within a few seconds. The rate of drying of the film may be controlled by covering the castinq tank with a lid provided with a s~all hole in the centre through which the cellulose solution is applied. With a uniform film thickness, reflected light interference colours may be observed as the film dries successively thinner.
If the water surface is not clean, the thickness of the cellulose film formed is not uniform and it spreads out into a non-circular patch with 'fingers' extending outwards. The non-uniform film thickness is apparent from the random distribution of reflected interference colours.
A polymer film thickness of about 16 nanometers is produced by addition of lO microlitres of a 9% cellulose nitrate solution (9 g cellulose nitrate in 100 ml amyl acetate).
Once the film has dried, water is drained out through a tap at the bottom of the tank, such that the dried polymer film is lowered onto the diffraction grating surface. The difEraction grating and its support are gently removed Erom the tank and left to dry Eor several minutes.
~e have Eound that the a~hesion of cellulose nitrate to silver-coated polycarbonate diE~raction qratin~s is enllanced ~v castin~ the ~olymer Eilms :
.
~3~39~
onto distilled water whose p~ has been ad~usted to 9 by the addition of, for example, NaOH.
The use of any other solvent castable polymers is Possible with the correct choice ~, solvents and casting surfaces. Thus, for example, cellulose acetate films may be cast from amyl acetate, polystyrene from xylene, ~olyisobutylene ~rom petroleum ether and polyvinyl chloride - acetate copolymer ~ilms can be cast From cyclohexanone onto water surfaces.
In some circumstances enforced spreading of the solution on the casting liquid, using eonventional teehniques, may be neeessary.
Film thickness may be eontrolled by eare~ul seleetion of:
- polymer solution eoneentration - volume o~ material aoplied to the casting liquid sur~aee - type of solvent.
Amyl acetate ~or example is amphiphilic and spreads rapidly over a clean water surface. Other solvents may not spread as rapidly, with coneomitant increases in film thickness.
The technique is not limited to casting on a water sur~ace and other liquids may be used for the casting surface provided the solvent used is largely immiseible with, and the polymer-eontaining solution is o~ lower density than, the casting liquid. Thus, for example, ethyl eellulose ean be east onto mercury, diisocyanates together with polyols can be co-cast onto mercury and gelatin can be cast from water onto carbon tetrachloride.
It is also possible to produce thin polymer layers using the solvent casting technique to cast the polymer onto a solid surface. Thus, for example, polyethylene terephthalate can be cast from trifluoro-acetic acid, polycarbonate from dichloromethane, ~olyvinyl formal from dioxan, polysulfone Ero~
.
~3~3~
chloroform, cellulo~se acetate ~rom acetone and polypropylene can be cast ~rom xylene, in each case on to an appropriate substrate. It ;s to be understood that where the polymer is cast onto a cast.lg liquid the casting liquid must be compati~le with the material of the di~raction grating immersed therein and where the polymer is cast onto a solid sur~ace the solvent used to bring the polymer into solution prior to casting must he compati~le ~ith the material of the solid sur~ace.
Bindinq of Bioloqical Materials to the Polymer Surface Three different methods of couplinq biological lS molecules to a cellulose nitrate polymer surface on a diffraction grating, have been considered:
a) covalent bonding b) non-covalent adsorption c) covalent bonding of the biological molecule to a pol~mer which is entrapped within the cellulose nitrate.
a) Covalent ~onding Established ~rocedures exist ~or covalently bonding biological materials, such as proteins, to cellulosic material. Published procedures include the use of chemical reagents which modify the polymer unit to form reactive groups which covalently bind to typical protein groups such as, ~or example, free amino groups.
Commonly used chemical reagents include:
- cyanogen bromicle - tosyl chloride - ~ titanium complexes - carbodiimide - cyanurlc chloride `
' .
~r . 13~3~8~
-- - oxirane - periodate.
b) Non-Covalent Adsorption Biological molecules, for example, sheep serum proteins, eibonucleases and immunoglobulins, have been found to hind very efficiently to cellulose nitrate. ~imple addition o~ an aqueous protein solution at room temperature or Lower has resulted in bound protein which could not easily be removed by washing with water, buffer solutions or detergents.
For example, incubation of a cellulose nitrate-coated grating for 2 hours at room temperature in lO mgml l bovine ~-globulin, 50 mM NaHCO3, pH 9, resulted in extensive coverage of the diffraction grating with the protein. Various treatments, such as, eor example, with mild detergent (5~ Tween*
20), buffer (50 mM NaHCO3), n heptane, and vH cycling between pH 2 and 10.6 did not affect the binding of the protein. Stronger detergent treatment (e.g.
2% SDS) has been found to damage the cellulose nitrate film.
c) Reactive PolYmer Entrapment This procedure separates the two important functions of the diffraction grating layer - adhesion and conformation to the grating surface - reactivity towards protein or other bioloqical material or binding partner and meets each requirement with a separate polymer.
A monomeric material is allowe~ to difeuse into the polymer aefixed to the dif~raction qratinq.
The monomer is then polymerised to give a network of the second polymer entangled in the first, a~Eixed * TRADE MARK
~ ",~
.~
~3~3~3~
polymer. The second polymer is chosen to have chemical groups that are reactive towards protein or other biological molecules.
Thus, ~or example, dif~raction gratings coated with cellulose nitrate according to the ~rocedure described herein are incubated with 1~ aqueous glutaraldehyde monomer solution pH 5.5 for 30 minutes at room temperature. The p~ is then adjusted to 10, and the diffraction gratings left for ~0 minutes to allow polymerisation of the glutaraldehyde.
Polymerisation may be monitored by optical absorbance measurements at 235 nm. Such techniques have been previously described, see for example, US-4001583.
Free reactive carbonyl groups in the polyglutaraldehyde are thus exposed at the surface of the polymer layer.
Treatment of these diffraction gratings with protein solutions, e.g. 5 mgml 1 bovine ~-globulin or 5 mgml 1 ribonuclease, at 4C overnight in 50 mM
NaHCO3 pH 9, results in strong binding between the protein and the polyglutaraldehyde, believed to be the result of covalent bonding between the polyglutaraldehyde carbonyl groups and the amino groups of the proteins.
~, . . .
Claims (20)
1. A method of applying a uniform, thin polymer film to the surface of a metallised diffraction grating which comprises the steps of:-i) inserting the metallised diffraction grating into a bath containing a casting liquid, ii) forming a thin film of the desired polymer over the surface of the casting liquid, and iii) either raising the diffraction grating through the casting liquid to engage the film of polymer on the surface of the liquid or allowing the casting liquid to drain out of the bath so that the polymer film on the surface of the liquid drops as the liquid level drops until it engages and spreads over the surface of the diffraction grating, in either case the diffraction grating being located at a small angle to the horizontal to facilitate draining of the casting liquid from the surface of the diffraction grating.
2. A method of preparing an optical structure useful for the detection of a ligand in a spectrophotometric assay technique, said optical structure having a surface relief profile or being capable of supporting surface plasmon resonance, which method comprises applying to the entire surface of the optical structure a uniform, thin layer of an organic polymer by means of a solvent casting technique and adsorbing on or binding to the thin layer of organic polymer, either directly or indirectly, a specific binding partner for the ligand.
3. The method of claim 2 wherein the optical structure is a metallized diffraction grating.
4. The method of claim 3 wherein the thin layer of organic polymer is 5 to 100 nm thick.
5. The method of claim 4 wherein the metallized diffraction grating is adapted to support surface plasmon resonance of a ligand/specific binding partner complex formed thereon.
6. The method of claim 5 wherein the thin layer of organic polymer is activated prior to contact with the specific binding partner to facilitate binding thereof.
7. The method of claim 6 wherein the specific binding partner is applied to at least one discrete region of the thin layer of organic polymer to form at least one test region and a non-binding protein is applied to at least one other discrete region of the thin layer of organic polymer to form at least one reference region.
8. The method of claim 7 wherein the metallized diffraction grating has a surface comprising a thin film of silver less than 500 nm thick.
9. The method of claim 8 wherein the specific binding partner is an antigen or antibody.
10. The method of claim 9 wherein the organic polymer is cellulose nitrate.
11. An optical biosensor useful for the detection of a ligand in a spectrophotometric assay technique which comprises an optical structure having a surface relief profile coated over its entire surface with a uniform, thin layer of an organic polymer of 5 to 100 nm thick to which is bound or adsorbed, directly or indirectly, a specific binding partner for the ligand.
12. The optical biosensor of claim 11 wherein the optical structure is a metallized diffraction grating.
13. The optical biosensor of claim 12 wherein the metallized diffraction grating has a surface comprising a thin film of silver less than 500 nm thick.
14. The method of claim 1 wherein the thin polymer film is 5 to 100 nm thick.
15. The method of claim 14 wherein the casting solution is water and the polymer is cellulose nitrate.
16. The method of claim 15 wherein the thin film of polymer is formed by dropping a solution of cellulose nitrate in amyl acetate onto the water surface.
17. The method of claim 16 additionally comprising adsorbing on or binding to the thin polymer film either directly or indirectly, a specific binding partner for a ligand which is desired to be assayed.
18. In a method of detecting a ligand in a sample utilizing a spectrophotometric assay technique which comprises contacting the sample with an optical biosensor and measuring the change in optical characteristics of the optical biosensor caused by formation of a ligand/specific binding partner complex thereon, the improvement wherein the optical biosensor comprises an optical structure having a surface relief profile coated over its entire surface with a uniform, thin layer of an organic polymer of 5 to 100 nm thick to which is bound or adsorbed, directly or indirectly, the specific binding partner for the ligand.
19. The method of claim 18 wherein the optical structure is a metallized diffraction grating.
20. The method of claim 19 wherein the metallized diffraction grating has a surface comprising a thin film of silver less than 500 nm thick.
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| GB868618133A GB8618133D0 (en) | 1986-07-24 | 1986-07-24 | Biosensors |
| GB8618133 | 1986-07-24 |
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| Publication Number | Publication Date |
|---|---|
| CA1303981C true CA1303981C (en) | 1992-06-23 |
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| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CA000542873A Expired - Lifetime CA1303981C (en) | 1986-07-24 | 1987-07-23 | Polymer-coated optical structures |
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|---|---|
| US (1) | US4992385A (en) |
| EP (1) | EP0254575B1 (en) |
| JP (2) | JP2528134B2 (en) |
| AT (1) | ATE86744T1 (en) |
| CA (1) | CA1303981C (en) |
| DE (1) | DE3784576T2 (en) |
| ES (1) | ES2038183T3 (en) |
| GB (1) | GB8618133D0 (en) |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| GB8705650D0 (en) * | 1987-03-10 | 1987-04-15 | Pa Consulting Services | Assay technique |
| GB8705649D0 (en) * | 1987-03-10 | 1987-04-15 | Pa Consulting Services | Assay sensor |
| GB8804669D0 (en) * | 1988-02-27 | 1988-03-30 | Medical Res Council | Immobilisation of haptens |
| GB2217447A (en) * | 1988-04-21 | 1989-10-25 | Atomic Energy Authority Uk | Bonding to metal surfaces |
| SE462454B (en) * | 1988-11-10 | 1990-06-25 | Pharmacia Ab | METHOD FOR USE IN BIOSENSORS |
| SE8804074D0 (en) * | 1988-11-10 | 1988-11-10 | Pharmacia Ab | SENSOR UNIT AND ITS USE IN BIOSENSOR SYSTEM |
| AU6354190A (en) * | 1989-08-21 | 1991-04-03 | Board Of Regents Of The University Of Washington, The | Multiple-probe diagnostic sensor |
| JPH03122553A (en) * | 1989-10-04 | 1991-05-24 | Olympus Optical Co Ltd | Photosensor |
| US5252743A (en) * | 1989-11-13 | 1993-10-12 | Affymax Technologies N.V. | Spatially-addressable immobilization of anti-ligands on surfaces |
| JPH05504195A (en) * | 1990-02-22 | 1993-07-01 | ザ・ロイヤル・インステイチユーシヨン・フオー・ジ・アドバンスメント・オブ・ラーニング （マクギル・ユニバーシテイ） | Solid phase interference immunoassay system |
| JPH04211202A (en) * | 1990-03-19 | 1992-08-03 | Canon Inc | Reflection type diffraction grating and device by use of same deffraction grating |
| DE4024476C1 (en) * | 1990-08-02 | 1992-02-27 | Boehringer Mannheim Gmbh, 6800 Mannheim, De | |
| US5763191A (en) * | 1990-12-12 | 1998-06-09 | Boehringer Mannheim Gmbh | Universal binding film |
| DE4039677A1 (en) * | 1990-12-12 | 1992-06-17 | Boehringer Mannheim Gmbh | UNIVERSAL BINDING FILM |
| JPH05506314A (en) * | 1991-02-11 | 1993-09-16 | バイオスター・メディカル・プロダクツ・インコーポレイテッド | Method of manufacturing test surfaces for optical detection of biological binding |
| EP0643833A1 (en) * | 1991-06-04 | 1995-03-22 | FISONS plc | Analytical device |
| GB9111912D0 (en) * | 1991-06-04 | 1991-07-24 | Fisons Plc | Analytical methods |
| JPH07500413A (en) * | 1991-07-22 | 1995-01-12 | クロエン、インストルメンツ、リミテッド | Substrates for surface-enhancing analytical processing and substrates prepared for specific purposes |
| US5837552A (en) * | 1991-07-22 | 1998-11-17 | Medifor, Ltd. | Surface-enhanced analytical procedures and substrates |
| CA2139242A1 (en) * | 1992-07-17 | 1994-02-03 | Richard Calvin Ebersole | Analyte detection by means of an analyte-responsive polymer |
| US5395754A (en) * | 1992-07-31 | 1995-03-07 | Hybritech Incorporated | Membrane-based immunoassay method |
| EP0598340B1 (en) * | 1992-11-17 | 2012-06-13 | O.S.P. Inc. | Use of a copolymer film on a substrate for detecting chemical substances |
| GB2273772A (en) * | 1992-12-16 | 1994-06-29 | Granta Lab Ltd | Detection of macromolecules utilising light diffraction |
| US5395587A (en) * | 1993-07-06 | 1995-03-07 | Smithkline Beecham Corporation | Surface plasmon resonance detector having collector for eluted ligate |
| US5955153A (en) * | 1993-10-01 | 1999-09-21 | Johnson & Johnson Clinical Diagnostics, Inc. | Production of carriers for surface plasmon resonance |
| ATE208653T1 (en) * | 1994-03-04 | 2001-11-15 | Usf Filtration & Separations | LARGE-PORED MEMBRANE MADE OF SYNTHETIC POLYMERS |
| GB9406142D0 (en) | 1994-03-28 | 1994-05-18 | British Tech Group | A sensor |
| US5854863A (en) * | 1996-03-15 | 1998-12-29 | Erb; Judith | Surface treatment and light injection method and apparatus |
| US5852229A (en) * | 1996-05-29 | 1998-12-22 | Kimberly-Clark Worldwide, Inc. | Piezoelectric resonator chemical sensing device |
| US6020047A (en) * | 1996-09-04 | 2000-02-01 | Kimberly-Clark Worldwide, Inc. | Polymer films having a printed self-assembling monolayer |
| US6048623A (en) * | 1996-12-18 | 2000-04-11 | Kimberly-Clark Worldwide, Inc. | Method of contact printing on gold coated films |
| US5922550A (en) * | 1996-12-18 | 1999-07-13 | Kimberly-Clark Worldwide, Inc. | Biosensing devices which produce diffraction images |
| US5958704A (en) * | 1997-03-12 | 1999-09-28 | Ddx, Inc. | Sensing system for specific substance and molecule detection |
| US6180288B1 (en) | 1997-03-21 | 2001-01-30 | Kimberly-Clark Worldwide, Inc. | Gel sensors and method of use thereof |
| US5955378A (en) * | 1997-08-20 | 1999-09-21 | Challener; William A. | Near normal incidence optical assaying method and system having wavelength and angle sensitivity |
| US5925878A (en) * | 1997-08-20 | 1999-07-20 | Imation Corp. | Diffraction anomaly sensor having grating coated with protective dielectric layer |
| US5994150A (en) * | 1997-11-19 | 1999-11-30 | Imation Corp. | Optical assaying method and system having rotatable sensor disk with multiple sensing regions |
| JP3437170B2 (en) * | 1997-12-12 | 2003-08-18 | ピーイー コーポレイション （エヌワイ） | Optical resonance analysis system |
| US6060256A (en) | 1997-12-16 | 2000-05-09 | Kimberly-Clark Worldwide, Inc. | Optical diffraction biosensor |
| US5986762A (en) * | 1998-06-15 | 1999-11-16 | Imation Corp. | Optical sensor having optimized surface profile |
| GB9820919D0 (en) * | 1998-09-26 | 1998-11-18 | Secr Defence | Diagnostic method |
| US6320991B1 (en) | 1998-10-16 | 2001-11-20 | Imation Corp. | Optical sensor having dielectric film stack |
| US6221579B1 (en) * | 1998-12-11 | 2001-04-24 | Kimberly-Clark Worldwide, Inc. | Patterned binding of functionalized microspheres for optical diffraction-based biosensors |
| US6579673B2 (en) * | 1998-12-17 | 2003-06-17 | Kimberly-Clark Worldwide, Inc. | Patterned deposition of antibody binding protein for optical diffraction-based biosensors |
| US6771376B2 (en) * | 1999-07-05 | 2004-08-03 | Novartis Ag | Sensor platform, apparatus incorporating the platform, and process using the platform |
| EP1192448B1 (en) | 1999-07-05 | 2006-09-27 | Novartis AG | Process of using a sensor platform |
| US7167615B1 (en) | 1999-11-05 | 2007-01-23 | Board Of Regents, The University Of Texas System | Resonant waveguide-grating filters and sensors and methods for making and using same |
| US8111401B2 (en) * | 1999-11-05 | 2012-02-07 | Robert Magnusson | Guided-mode resonance sensors employing angular, spectral, modal, and polarization diversity for high-precision sensing in compact formats |
| US6399295B1 (en) | 1999-12-17 | 2002-06-04 | Kimberly-Clark Worldwide, Inc. | Use of wicking agent to eliminate wash steps for optical diffraction-based biosensors |
| EP1266207B1 (en) * | 2000-03-22 | 2010-10-27 | Axela Inc. | Method and apparatus for multiple-analyte assay |
| US7264973B2 (en) * | 2000-10-30 | 2007-09-04 | Sru Biosystems, Inc. | Label-free methods for performing assays using a colorimetric resonant optical biosensor |
| US20030092075A1 (en) * | 2000-10-30 | 2003-05-15 | Sru Biosystems, Llc | Aldehyde chemical surface activation processes and test methods for colorimetric resonant sensors |
| US7306827B2 (en) * | 2000-10-30 | 2007-12-11 | Sru Biosystems, Inc. | Method and machine for replicating holographic gratings on a substrate |
| US6951715B2 (en) * | 2000-10-30 | 2005-10-04 | Sru Biosystems, Inc. | Optical detection of label-free biomolecular interactions using microreplicated plastic sensor elements |
| US7142296B2 (en) * | 2000-10-30 | 2006-11-28 | Sru Biosystems, Inc. | Method and apparatus for detecting biomolecular interactions |
| US20030113766A1 (en) * | 2000-10-30 | 2003-06-19 | Sru Biosystems, Llc | Amine activated colorimetric resonant biosensor |
| US7153702B2 (en) * | 2000-10-30 | 2006-12-26 | Sru Biosystems, Inc. | Label-free methods for performing assays using a colorimetric resonant reflectance optical biosensor |
| US7371562B2 (en) * | 2000-10-30 | 2008-05-13 | Sru Biosystems, Inc. | Guided mode resonant filter biosensor using a linear grating surface structure |
| US7094595B2 (en) * | 2000-10-30 | 2006-08-22 | Sru Biosystems, Inc. | Label-free high-throughput optical technique for detecting biomolecular interactions |
| US7175980B2 (en) * | 2000-10-30 | 2007-02-13 | Sru Biosystems, Inc. | Method of making a plastic colorimetric resonant biosensor device with liquid handling capabilities |
| US7202076B2 (en) | 2000-10-30 | 2007-04-10 | Sru Biosystems, Inc. | Label-free high-throughput optical technique for detecting biomolecular interactions |
| US7217574B2 (en) * | 2000-10-30 | 2007-05-15 | Sru Biosystems, Inc. | Method and apparatus for biosensor spectral shift detection |
| US7101660B2 (en) * | 2000-10-30 | 2006-09-05 | Sru Biosystems, Inc. | Method for producing a colorimetric resonant reflection biosensor on rigid surfaces |
| US7023544B2 (en) | 2000-10-30 | 2006-04-04 | Sru Biosystems, Inc. | Method and instrument for detecting biomolecular interactions |
| US7615339B2 (en) * | 2000-10-30 | 2009-11-10 | Sru Biosystems, Inc. | Method for producing a colorimetric resonant reflection biosensor on rigid surfaces |
| US7070987B2 (en) * | 2000-10-30 | 2006-07-04 | Sru Biosystems, Inc. | Guided mode resonant filter biosensor using a linear grating surface structure |
| US7575939B2 (en) * | 2000-10-30 | 2009-08-18 | Sru Biosystems, Inc. | Optical detection of label-free biomolecular interactions using microreplicated plastic sensor elements |
| US7300803B2 (en) * | 2000-10-30 | 2007-11-27 | Sru Biosystems, Inc. | Label-free methods for performing assays using a colorimetric resonant reflectance optical biosensor |
| US7875434B2 (en) * | 2000-10-30 | 2011-01-25 | Sru Biosystems, Inc. | Label-free methods for performing assays using a colorimetric resonant reflectance optical biosensor |
| US6576430B1 (en) * | 2000-11-20 | 2003-06-10 | Becton, Dickinson And Company | Detection of ligands by refractive surface methods |
| DE10064146A1 (en) * | 2000-12-22 | 2002-07-04 | Andreas Hofmann | Biosensor and method for its production |
| JP4554205B2 (en) * | 2001-09-13 | 2010-09-29 | アクセラ インコーポレーテッド | Method and apparatus for calibration based on diffraction of light |
| US7098041B2 (en) | 2001-12-11 | 2006-08-29 | Kimberly-Clark Worldwide, Inc. | Methods to view and analyze the results from diffraction-based diagnostics |
| US7102752B2 (en) | 2001-12-11 | 2006-09-05 | Kimberly-Clark Worldwide, Inc. | Systems to view and analyze the results from diffraction-based diagnostics |
| US7384598B2 (en) * | 2001-12-21 | 2008-06-10 | Kimberly-Clark Worldwide, Inc. | Diagnostic device |
| US7244393B2 (en) * | 2001-12-21 | 2007-07-17 | Kimberly-Clark Worldwide, Inc. | Diagnostic device and system |
| US20030119203A1 (en) * | 2001-12-24 | 2003-06-26 | Kimberly-Clark Worldwide, Inc. | Lateral flow assay devices and methods for conducting assays |
| US6837171B1 (en) | 2002-04-29 | 2005-01-04 | Palmer/Snyder Furniture Company | Lightweight table with unitized table top |
| US8367013B2 (en) * | 2001-12-24 | 2013-02-05 | Kimberly-Clark Worldwide, Inc. | Reading device, method, and system for conducting lateral flow assays |
| US7485453B2 (en) * | 2002-05-03 | 2009-02-03 | Kimberly-Clark Worldwide, Inc. | Diffraction-based diagnostic devices |
| US7223368B2 (en) * | 2002-05-03 | 2007-05-29 | Kimberly-Clark Worldwide, Inc. | Diffraction-based diagnostic devices |
| US7214530B2 (en) * | 2002-05-03 | 2007-05-08 | Kimberly-Clark Worldwide, Inc. | Biomolecule diagnostic devices and method for producing biomolecule diagnostic devices |
| US7771922B2 (en) * | 2002-05-03 | 2010-08-10 | Kimberly-Clark Worldwide, Inc. | Biomolecule diagnostic device |
| US7223534B2 (en) * | 2002-05-03 | 2007-05-29 | Kimberly-Clark Worldwide, Inc. | Diffraction-based diagnostic devices |
| US7118855B2 (en) * | 2002-05-03 | 2006-10-10 | Kimberly-Clark Worldwide, Inc. | Diffraction-based diagnostic devices |
| DE10222979B4 (en) * | 2002-05-23 | 2006-06-08 | Sartorius Ag | Method for amplifying a signal |
| US7091049B2 (en) * | 2002-06-26 | 2006-08-15 | Kimberly-Clark Worldwide, Inc. | Enhanced diffraction-based biosensor devices |
| US20040126875A1 (en) * | 2002-09-12 | 2004-07-01 | Putnam Martin A. | Assay stick |
| US7126755B2 (en) * | 2002-09-12 | 2006-10-24 | Moon John A | Method and apparatus for labeling using diffraction grating-based encoded optical identification elements |
| EP1535241A1 (en) * | 2002-08-20 | 2005-06-01 | Cyvera Corporation | Diffraction grating-based optical identification element |
| US7872804B2 (en) * | 2002-08-20 | 2011-01-18 | Illumina, Inc. | Encoded particle having a grating with variations in the refractive index |
| US7900836B2 (en) * | 2002-08-20 | 2011-03-08 | Illumina, Inc. | Optical reader system for substrates having an optically readable code |
| US7164533B2 (en) * | 2003-01-22 | 2007-01-16 | Cyvera Corporation | Hybrid random bead/chip based microarray |
| US7190522B2 (en) * | 2002-09-12 | 2007-03-13 | Cyvera Corporation | Chemical synthesis using diffraction grating-based encoded optical elements |
| US7901630B2 (en) * | 2002-08-20 | 2011-03-08 | Illumina, Inc. | Diffraction grating-based encoded microparticle assay stick |
| US7441703B2 (en) * | 2002-08-20 | 2008-10-28 | Illumina, Inc. | Optical reader for diffraction grating-based encoded optical identification elements |
| US7508608B2 (en) * | 2004-11-17 | 2009-03-24 | Illumina, Inc. | Lithographically fabricated holographic optical identification element |
| EP1535242A1 (en) * | 2002-08-20 | 2005-06-01 | Cyvera Corporation | Diffraction grating-based encoded micro-particles for multiplexed experiments |
| US20050227252A1 (en) * | 2002-08-20 | 2005-10-13 | Moon John A | Diffraction grating-based encoded articles for multiplexed experiments |
| US7923260B2 (en) | 2002-08-20 | 2011-04-12 | Illumina, Inc. | Method of reading encoded particles |
| US7285424B2 (en) * | 2002-08-27 | 2007-10-23 | Kimberly-Clark Worldwide, Inc. | Membrane-based assay devices |
| US7432105B2 (en) * | 2002-08-27 | 2008-10-07 | Kimberly-Clark Worldwide, Inc. | Self-calibration system for a magnetic binding assay |
| US7314763B2 (en) * | 2002-08-27 | 2008-01-01 | Kimberly-Clark Worldwide, Inc. | Fluidics-based assay devices |
| US20040043508A1 (en) * | 2002-09-03 | 2004-03-04 | Frutos Anthony G. | Polymer-coated substrates for binding biological molecules |
| US7927822B2 (en) * | 2002-09-09 | 2011-04-19 | Sru Biosystems, Inc. | Methods for screening cells and antibodies |
| US7429492B2 (en) * | 2002-09-09 | 2008-09-30 | Sru Biosystems, Inc. | Multiwell plates with integrated biosensors and membranes |
| US20100255603A9 (en) | 2002-09-12 | 2010-10-07 | Putnam Martin A | Method and apparatus for aligning microbeads in order to interrogate the same |
| US7092160B2 (en) | 2002-09-12 | 2006-08-15 | Illumina, Inc. | Method of manufacturing of diffraction grating-based optical identification element |
| WO2004024328A1 (en) * | 2002-09-12 | 2004-03-25 | Cyvera Corporation | Method and apparatus for aligning elongated microbeads in order to interrogate the same |
| WO2004025563A1 (en) * | 2002-09-12 | 2004-03-25 | Cyvera Corporation | Diffraction grating-based encoded micro-particles for multiplexed experiments |
| US7169550B2 (en) * | 2002-09-26 | 2007-01-30 | Kimberly-Clark Worldwide, Inc. | Diffraction-based diagnostic devices |
| US7781172B2 (en) * | 2003-11-21 | 2010-08-24 | Kimberly-Clark Worldwide, Inc. | Method for extending the dynamic detection range of assay devices |
| US20040106190A1 (en) * | 2002-12-03 | 2004-06-03 | Kimberly-Clark Worldwide, Inc. | Flow-through assay devices |
| US7309614B1 (en) | 2002-12-04 | 2007-12-18 | Sru Biosystems, Inc. | Self-referencing biodetection method and patterned bioassays |
| US20040121334A1 (en) * | 2002-12-19 | 2004-06-24 | Kimberly-Clark Worldwide, Inc. | Self-calibrated flow-through assay devices |
| US7247500B2 (en) * | 2002-12-19 | 2007-07-24 | Kimberly-Clark Worldwide, Inc. | Reduction of the hook effect in membrane-based assay devices |
| US7851209B2 (en) * | 2003-04-03 | 2010-12-14 | Kimberly-Clark Worldwide, Inc. | Reduction of the hook effect in assay devices |
| US20040197819A1 (en) * | 2003-04-03 | 2004-10-07 | Kimberly-Clark Worldwide, Inc. | Assay devices that utilize hollow particles |
| US7070406B2 (en) * | 2003-04-29 | 2006-07-04 | Hewlett-Packard Development Company, L.P. | Apparatus for embossing a flexible substrate with a pattern carried by an optically transparent compliant media |
| US7497992B2 (en) * | 2003-05-08 | 2009-03-03 | Sru Biosystems, Inc. | Detection of biochemical interactions on a biosensor using tunable filters and tunable lasers |
| US20060057729A1 (en) * | 2003-09-12 | 2006-03-16 | Illumina, Inc. | Diffraction grating-based encoded element having a substance disposed thereon |
| US8298780B2 (en) * | 2003-09-22 | 2012-10-30 | X-Body, Inc. | Methods of detection of changes in cells |
| AU2004290375A1 (en) * | 2003-11-06 | 2005-05-26 | Sru Biosystems, Inc. | High-density amine-functionalized surface |
| US7943395B2 (en) * | 2003-11-21 | 2011-05-17 | Kimberly-Clark Worldwide, Inc. | Extension of the dynamic detection range of assay devices |
| US20050112703A1 (en) * | 2003-11-21 | 2005-05-26 | Kimberly-Clark Worldwide, Inc. | Membrane-based lateral flow assay devices that utilize phosphorescent detection |
| US7713748B2 (en) * | 2003-11-21 | 2010-05-11 | Kimberly-Clark Worldwide, Inc. | Method of reducing the sensitivity of assay devices |
| US20050136550A1 (en) * | 2003-12-19 | 2005-06-23 | Kimberly-Clark Worldwide, Inc. | Flow control of electrochemical-based assay devices |
| US7943089B2 (en) * | 2003-12-19 | 2011-05-17 | Kimberly-Clark Worldwide, Inc. | Laminated assay devices |
| US7433123B2 (en) * | 2004-02-19 | 2008-10-07 | Illumina, Inc. | Optical identification element having non-waveguide photosensitive substrate with diffraction grating therein |
| US7796266B2 (en) * | 2004-04-30 | 2010-09-14 | Kimberly-Clark Worldwide, Inc. | Optical detection system using electromagnetic radiation to detect presence or quantity of analyte |
| US7815854B2 (en) * | 2004-04-30 | 2010-10-19 | Kimberly-Clark Worldwide, Inc. | Electroluminescent illumination source for optical detection systems |
| US20050244953A1 (en) * | 2004-04-30 | 2005-11-03 | Kimberly-Clark Worldwide, Inc. | Techniques for controlling the optical properties of assay devices |
| US7521226B2 (en) * | 2004-06-30 | 2009-04-21 | Kimberly-Clark Worldwide, Inc. | One-step enzymatic and amine detection technique |
| WO2006020363A2 (en) * | 2004-07-21 | 2006-02-23 | Illumina, Inc. | Method and apparatus for drug product tracking using encoded optical identification elements |
| WO2006055736A1 (en) | 2004-11-16 | 2006-05-26 | Illumina, Inc. | And methods and apparatus for reading coded microbeads |
| WO2006055735A2 (en) | 2004-11-16 | 2006-05-26 | Illumina, Inc | Scanner having spatial light modulator |
| US7604173B2 (en) * | 2004-11-16 | 2009-10-20 | Illumina, Inc. | Holographically encoded elements for microarray and other tagging labeling applications, and method and apparatus for making and reading the same |
| US20060110594A1 (en) * | 2004-11-24 | 2006-05-25 | Frutos Anthony G | Polymer-coated substrates for binding biomolecules and methods of making and using thereof |
| US20070121113A1 (en) * | 2004-12-22 | 2007-05-31 | Cohen David S | Transmission-based optical detection systems |
| US20060141527A1 (en) * | 2004-12-29 | 2006-06-29 | Caracci Stephen J | Method for creating a reference region and a sample region on a biosensor and the resulting biosensor |
| US7604984B2 (en) * | 2004-12-29 | 2009-10-20 | Corning Incorporated | Spatially scanned optical reader system and method for using same |
| US7629173B2 (en) * | 2004-12-29 | 2009-12-08 | Corning Incorporated | Optical reader system and method for monitoring and correcting lateral and angular misalignments of label independent biosensors |
| US7623624B2 (en) * | 2005-11-22 | 2009-11-24 | Illumina, Inc. | Method and apparatus for labeling using optical identification elements characterized by X-ray diffraction |
| US7781203B2 (en) * | 2005-12-29 | 2010-08-24 | Corning Incorporated | Supports for assaying analytes and methods of making and using thereof |
| US7830575B2 (en) * | 2006-04-10 | 2010-11-09 | Illumina, Inc. | Optical scanner with improved scan time |
| US20070264155A1 (en) * | 2006-05-09 | 2007-11-15 | Brady Michael D | Aerosol jet deposition method and system for creating a reference region/sample region on a biosensor |
| AU2007313830A1 (en) * | 2006-10-31 | 2008-05-08 | Sru Biosystems, Inc. | Method for blocking non-specific protein binding on a functionalized surface |
| CN101743465A (en) * | 2007-04-19 | 2010-06-16 | Sru生物系统公司 | Method for employing a biosensor to detect small molecules that bind directly to immobilized targets |
| WO2009009718A1 (en) * | 2007-07-11 | 2009-01-15 | Sru Biosystems, Inc. | Methods of identifying modulators of ion channels |
| US9134307B2 (en) * | 2007-07-11 | 2015-09-15 | X-Body, Inc. | Method for determining ion channel modulating properties of a test reagent |
| US7923241B2 (en) * | 2007-10-10 | 2011-04-12 | Corning Incorporated | Cell culture article and methods thereof |
| US20100021954A1 (en) * | 2008-07-23 | 2010-01-28 | Sophie Deshayes | High capacity nanoparticulate immobilization surface |
| WO2009058214A1 (en) * | 2007-10-30 | 2009-05-07 | Corning Incorporated | High capacity nanoparticulate immobilization surface |
| US8257936B2 (en) | 2008-04-09 | 2012-09-04 | X-Body Inc. | High resolution label free analysis of cellular properties |
| EP2304500A1 (en) * | 2008-06-04 | 2011-04-06 | SRU Biosystems, Inc. | Detection of promiscuous small submicrometer aggregates |
| US20100273185A1 (en) * | 2009-04-27 | 2010-10-28 | Sru Biosystems, Inc. | Detection of Biased Agonist Activation |
| AU2010248784A1 (en) * | 2009-05-15 | 2011-12-01 | Sru Biosystems, Inc | Detection of changes in cell populations and mixed cell populations |
| US8593630B2 (en) * | 2009-10-07 | 2013-11-26 | The Board Of Trustees Of The University Of Illinois | Discrete frequency spectroscopy and instrumentation |
| WO2011120042A1 (en) * | 2010-03-26 | 2011-09-29 | Sru Biosystems, Inc. | Use of induced pluripotent cells and other cells for screening compound libraries |
Family Cites Families (22)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| NL67090C (en) * | 1947-06-27 | |||
| DE1108461B (en) * | 1954-10-14 | 1961-06-08 | Nat Res Dev | Method for copying optical diffraction grating replicas |
| US3839067A (en) * | 1972-09-25 | 1974-10-01 | Bell Telephone Labor Inc | Method of making a thin film wave guide employing stress induced birefringence by desposition in a polymer from solution |
| US3926564A (en) * | 1974-02-25 | 1975-12-16 | Gen Electric | Substrate for immunological tests and method of fabrication thereof |
| US3979184A (en) * | 1975-05-27 | 1976-09-07 | General Electric Company | Diagnostic device for visually detecting presence of biological particles |
| US4080476A (en) * | 1976-11-15 | 1978-03-21 | Datascope Corporation | Anti-fog coated optical substrates |
| US4357142A (en) * | 1980-07-18 | 1982-11-02 | Akzona Incorporated | Glass support coated with synthetic polymer for bioprocess |
| US4363634A (en) * | 1980-07-18 | 1982-12-14 | Akzona Incorporated | Glass support coated with synthetic polymer for bioprocess |
| JPS5767860A (en) * | 1980-10-15 | 1982-04-24 | Fuji Photo Film Co Ltd | Material for multilayer analysis |
| JPS5821213A (en) * | 1981-07-31 | 1983-02-08 | Canon Inc | Optical coupler |
| DE3135196A1 (en) * | 1981-09-05 | 1983-03-17 | Merck Patent Gmbh, 6100 Darmstadt | METHOD, MEANS AND DEVICE FOR DETERMINING BIOLOGICAL COMPONENTS |
| DE3343636A1 (en) * | 1982-12-07 | 1984-06-07 | AVL AG, 8201 Schaffhausen | Sensor element for optically measuring fluorescence, and method of producing it |
| CA1237645A (en) * | 1982-12-21 | 1988-06-07 | John H. Fisher | Assay technique |
| US4537861A (en) * | 1983-02-03 | 1985-08-27 | Elings Virgil B | Apparatus and method for homogeneous immunoassay |
| DE3346795A1 (en) * | 1983-12-23 | 1985-07-04 | E. Prof. Dr. 3550 Marburg Geyer | Enzyme immunoassay and corresponding diagnostic aid |
| JPS60173842A (en) * | 1984-02-20 | 1985-09-07 | Canon Inc | Forming method of pattern |
| GB8413262D0 (en) * | 1984-05-24 | 1984-06-27 | Perkins P G | Lens coating material |
| US4647544A (en) * | 1984-06-25 | 1987-03-03 | Nicoli David F | Immunoassay using optical interference detection |
| EP0190123A1 (en) * | 1984-07-20 | 1986-08-13 | GAUNTT, Charles, O. | Immunoregulatory and anti-viral compound |
| GB8423204D0 (en) * | 1984-09-14 | 1984-10-17 | Comtech Res Unit | Assay technique and equipment |
| GB8509492D0 (en) * | 1985-04-12 | 1985-05-15 | Plessey Co Plc | Optical assay |
| DE3680999D1 (en) * | 1985-05-29 | 1991-09-26 | Artificial Sensing Instr Asi A | OPTICAL SENSOR FOR SELECTIVE DETECTION OF SUBSTANCES AND DETECTION OF REFRIGERATION CHANGES IN MEASURING SUBSTANCES. |
-
1986
- 1986-07-24 GB GB868618133A patent/GB8618133D0/en active Pending
-
1987
- 1987-07-14 IL IL83180A patent/IL83180A/en not_active IP Right Cessation
- 1987-07-22 US US07/076,519 patent/US4992385A/en not_active Expired - Lifetime
- 1987-07-23 AT AT87306535T patent/ATE86744T1/en not_active IP Right Cessation
- 1987-07-23 ES ES198787306535T patent/ES2038183T3/en not_active Expired - Lifetime
- 1987-07-23 DE DE8787306535T patent/DE3784576T2/en not_active Expired - Lifetime
- 1987-07-23 EP EP87306535A patent/EP0254575B1/en not_active Expired - Lifetime
- 1987-07-23 CA CA000542873A patent/CA1303981C/en not_active Expired - Lifetime
- 1987-07-24 JP JP62183772A patent/JP2528134B2/en not_active Expired - Lifetime
-
1993
- 1993-03-16 GR GR930400595T patent/GR3007383T3/el unknown
-
1995
- 1995-06-23 JP JP7157861A patent/JP2592588B2/en not_active Expired - Lifetime
Also Published As
| Publication number | Publication date |
|---|---|
| JPS63100355A (en) | 1988-05-02 |
| EP0254575B1 (en) | 1993-03-10 |
| AU594176B2 (en) | 1990-03-01 |
| JP2592588B2 (en) | 1997-03-19 |
| ATE86744T1 (en) | 1993-03-15 |
| US4992385A (en) | 1991-02-12 |
| JP2528134B2 (en) | 1996-08-28 |
| DE3784576T2 (en) | 1993-09-23 |
| EP0254575A2 (en) | 1988-01-27 |
| IL83180A (en) | 1992-09-06 |
| EP0254575A3 (en) | 1990-05-30 |
| GB8618133D0 (en) | 1986-09-03 |
| GR3007383T3 (en) | 1993-07-30 |
| AU7600787A (en) | 1988-01-28 |
| JPH0868749A (en) | 1996-03-12 |
| IL83180A0 (en) | 1987-12-31 |
| DE3784576D1 (en) | 1993-04-15 |
| ES2038183T3 (en) | 1993-07-16 |
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Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| MKEX | Expiry |