JPH07508350A - Verification devices formed from materials with different porosity - Google Patents

Abstract

(57) [Summary] This bulletin contains application data before electronic filing, so abstract data is not recorded.

Description

[Detailed description of the invention] This invention is a device composed of materials with different porosities. Specifically, regarding testing equipment that can be used specifically when performing "flow-through" testing. Ru. More specifically, the present invention relates to assay devices formed from materials with different porosity. Regarding the location.

Immunoassays can be performed using a single device, which includes certain absorbent materials, That is, it contains absorption bands, and these are the antigens of one analyte (analyte). Antibodies or Directing the flow of fluid containing the analyte through membranes that support other types of binding agents.

U.S. Patent No. 4,366,241 issued to Tom et al. Flow-through assay devices and assays using such devices are disclosed. This inspection The fixing device has an immunoadsorbent band, which immobilizes the members of the immunocouple. Liquid absorption band is immune placed adjacent to an adsorption zone that pulls the liquid sample through the immunoadsorption zone. wear. Absorption bands control the rate at which liquid samples are attracted through immunoadsorption bands can do.

U.S. Patent No. 4,632,901 issued to Valkers et al. Discloses an apparatus and method for performing a bacterial infection assay, in which antibodies such as monoclonal antibodies are Or coupled to a filter. Absorbent material absorbs liquid through a membrane or filter. It is located below the membrane or filter that directs the flow of the sample. liquid sump The analyte in the sample binds to the antibody on the membrane or filter. Labeled anti-analyte The body is then added. A wash step then removes unbound labeled antibody. wash Following the cleaning step, the presence of labeled antibodies on the membrane or filter is assayed for the analyte. It shows its existence.

According to one aspect of the invention, an assay device is provided that includes a first absorbent material and a second absorbent material. be done. The first absorbent material has at least one binding agent disposed surface for the analyte. have. The second absorbent material directs the flow of the sample to be assayed to at least one of the analytes. Direct to the binding agent. The second absorbent material is adjacent to the second absorbent material and has more pores than the first absorbent material. rate is low.

In a preferred embodiment, the second absorbent material surrounds the first absorbent material (e.g., The absorbent material is in the form of a cylinder and the second absorbent material has at least one bond for the analyte. Surround or encircle the cylinder to the surface of the cylinder supporting the agent. one When the sample is applied to the top of the assay device, the absorbent material with less porosity will direct the flow of the sample to the first absorbent material or cylinder, where the sample is It flows through the at least one binder and is absorbed by the first absorbent material.

In one embodiment, the first absorbent material is about 0.01 cc/g to about 0.400 O1 cc/g. cc/g, preferably from about 0.010 cc/g to about 0.280 cc/g. has a pore volume of . The first absorbent material also has a diameter from about 0.005μ to about 1254μ, as desired. Preferably, the pore diameter is between about 0.03μ and about 3μ.

In a preferred embodiment, the first absorbent material is a porous ceramic material.

[Second] The surface of the absorbent material is coated with humidifiers to speed up or slow down the desired absorption time. Alternatively, it is pretreated with a polymer. The surface of the first absorbent material further reduces non-specific binding. treated with surfactants and/or proteins to reduce The surface is coated with latex particles or immunoprecipitate-like particles that act as a sample carrier. can be processed with.

In one embodiment, the second absorbent material is non-porous and has a weight of about 1.8 g/cc to about 5 g/cc. ．． having a density of 0 g/cc, preferably about 2.2 g/cc to about 4.4 g/cc . Desirably, the second absorbent material is a non-porous ceramic material. Same as for the first absorbent material Similarly, the surface of the second absorbent material can be coated with humidifier to speed up or slow down the absorption time. or pretreated with polymers and/or surfactants and/or tannins. Processed with protein.

The first and second absorbent materials are of the same type of material (e.g., porous plastic, fibrous material). materials, ceramics, etc.), where such materials have a high porosity primary absorption to form a porosity gradient from the material to the low porosity or non-porous secondary absorbent material. It is processed. The porosity gradient can be determined, for example, by dry-pressing ceramic powder in a mold. It is formed by The lower die of the mold has a moving mold center. This central part is lifted during the dry powder filling stage to reduce the amount of dry powder above it. Make it less. The central part is pulled down during the compaction stage, which makes the finished part more porous. The sexual midsection is provided.

In one embodiment, the absorbent material is a non-fibrous porous plastic material; It is "wettable" and can provide capillary flow of the plastic.

Examples of non-fibrous plastic absorbent materials used include polyolefins, polyester Examples include Stell, porous polyvinyl chloride, and Boristine. porous plastic The stick is impregnated with a suitable material, such as cellulose material, and the material has the desired pores. Fill the pores of the plus deck to provide a porous material with a size of . one In embodiments, the porous plastic has a thickness of about 5μ to 500μ, preferably about 10μ. It has a diameter from u to 40μ.

The cellulosic material does not adversely affect the absorbency and wetting performance of the plastic. messenger Examples of cellulosic materials used are codon linters and starches such as arrowroot starch. It is.

Porous plastics can also be hydrophobic porous plastics, which are The addition of humectants such as surfactants can impart hydrophilicity or wettability. Sometimes it happens. Surfactants (e.g. porous plastic binders are cellulose applied to all or part of a porous plastic material (after being impregnated with material) I can do it. Plus Deck, which is hydrophobic, is part of the surface of porous plastic. is contacted with a surfactant to impart hydrophilic, or "wetting" properties to the surface. Certification The binder used in is applied to the "wettable" portion of the porous material. If limited When a small amount of detergent is applied, the specimen and labeled antibody will flow into the absorbent material. -rate will be slower, thus affecting the sensitivity of the assay.

The surfactant may be applied to the entire surface or a portion of the surface of the binder, thus covering the entire surface. Alternatively, a portion of the surface may be rendered hydrophilic.

In another embodiment, the absorbent material is formed from compressed fibrous material.

Compressed fibers are hydrophilic, or wettable, providing capillary flow to the fiber disc You can. The fibers are compressed to provide the desired porosity gradient. Supports the binder Absorbent surfaces with , preferably a smooth surface. Examples of textile materials used are not necessarily limited to However, glass fiber, polyester, polyamide, cellulose fiber, etc. , the fibrous material is placed into a compression die and then a perforator with an inverted cone-shaped cavity , and a porosity gradient is formed. The binder is applied directly to the surface of the absorbent material or solid particles such as latex particles, or diatomaceous earth, porous It is supported by glass, resin particles, or the like.

The surface of the absorbent material is coated with one material, e.g. or coated with any blocking material known in the art.

In a preferred embodiment, the absorbent material is made of porous ceramic material. ceramic Absorbent material is defined as a non-fibrous inorganic porous matrix. Ceramic material is , so as to provide a porosity gradient from a high porosity first absorbent material to a low porosity second absorbent material. It is formed.

This device is used in conjunction with a wide variety of assays. Such a test is not necessarily including but not limited to competitive assays, sandwich assays, enzyme-linked immunosorbents test (ELISA), agglutination test, indirect test, and their equivalents known in the art. Includes other tests.

In competitive assays, the analyte and tracer are analyte and tracer specific. competes with other binding agents. Tracers can be detected! bound to a marker or analyte or a suitable analogue thereof. Tracer and analyte are on the binder the amount of tracer that competes for a limited number of binding sites and is bound to the binding agent. is inversely proportional to the amount of analyte in the sample. amount of tracer, and similarly tested The amount can be determined by the amount of label present. in the tracer part Some labels or markers may be 1 e.g. iodine, cobalt, or tritium released. Radioactive isotopes, enzymes, fluorescent dyes, absorption dyes, luminescent substances, spin labels, biotin, Detection markers known to those skilled in the art such as colored particles or other labeling substances It is. A preferred label is a colored particle such as colloidal gold.

When a sandwich assay is used, the analyte-specific binding agent contains the analyte. have come into contact with a sample that contains or is suspected of containing the analyte. It will be done. Analyte present within the sample binds to the binding agent. The sample passes through the binder. After flowing into the absorbent material, the analyte/binder complex is contacted with the tracer. tray The sir is a ligand specific to the analyte being assayed. For example, tray The sensor can be an antibody that is elicited in response to the analyte being assayed. Distribution The ligand is preferably labeled as a detection marker as described above and is also present in the sample. The amount of 1M analyte present is determined by the amount of label present on the surface of the absorbent material.

Indirect Sandwich Assay 1 In this, the analyte bound to the supported binder is The analyte bound to the analyte is contacted with the analyte binding agent. child The tracer used in this assay binds to the analyte bound to the supported binding agent. f! 1111 ligand.

In the ELISA assay, the tracer or ligand is labeled with an enzyme and The amount of analyte present in the sample is determined by the amount of bound enzyme label present. It will be done. The ELISA test is performed similarly to a sandwich or competitive test.

In agglutination assays, particles that sensitize with antigens or antibodies specific to the analyte are used. The binding agent is contacted with a sample suspected of containing the analyte. Subject The presence of a body is evidenced by the agglomeration of solid particles.

The binding agent used in the assay depends on the analyte being assayed, e.g. if the analyte is antigenic or or habten (partial antigen), the binding agent is specific to the antibody or analyte. It is a naturally occurring substance that is unique. When the analyte is an antibody, the binding agent is an antibody, An antigen, or naturally occurring substance that is specific to the analyte.

The assays described above can be used to determine a wide variety of analytes. child Examples of analytes assayed in accordance with the invention include, but are not limited to, pharmaceuticals. products, hormones, polymers, antibodies, microorganisms, toxins, polypeptides, proteins, polysaccharides etc., nucleic acids, etc. The selection of a deceased subject is considered to be within the scope of those skilled in the art. It will be done.

The surface of the first absorbent material is small (some of the analyte (antigen, habten, antibody) Such an assay can be performed by placing one binding agent. The binder is , speckled or dispersed, sprayed or pre-coated onto the film to create symbolic forms. is placed on the surface of the first absorbent material in the form of a lint.

The binder is applied (either by passive or covalent methods of attachment) to the surface of the first absorbent material. A direct device (can be used or supported on solid particles that is placed on the surface of the first absorbent material) You can. The surfaces of the first and second absorbent materials then contain the analyte. Contacted with possible samples. The binder supported on the surface of the first absorbent material is Specific to the body. The analyte present binds to the binding agent on the surface of the first absorbent material. will combine to form an analyte binding agent complex. Apply the sample to the first and second aspiration. Upon application to the absorbent material, the sample flow is directed to and placed in the first absorbent material. the binder. This flow of sample is caused by the low pores of the second absorbent material. This is due to porosity, and the second absorbent material is more porous than the first absorbent material. creating a "furnelling" effect that flows towards the binder sample, thus This will provide optimal binding of the analyte. The surfaces of the first and second absorbent materials are as follows. contact with the tracer at the same time as, or subsequent to, contact with the sample. It will be done. A flow of tracer is also directed to the first absorbent material. tracer is a ligand The labeled form and the ligand employed depends on the assay format. competitive test , the tracer ligand is the ligand bound by the analyte binding agent. be. In a sandwich assay, the tracer ligand is bound (directly detected) by the analyte. sample) or bound by a binding agent for the analyte (indirect assay). If the Doinch test is employed, the tracer is preferably It is a labeled form of the ligand that is specific. In this assay, the analyte and tracer is bound to a supporting binder (tracer is bound directly to a supporting binder in competitive assays). , bound to the supporting binder through the analyte in the sandwich assay) and therefore Stays on the surface of the harvested material. Capillary movement through the first absorbent material allows the sample and tray to This occurs while the surfer passes through the supporting binder and the unbound portion of the flow to the absorbent material. The combination of occurs. The flow rate of the sample through the assay device is determined by the It depends on the porosity of the device obtained and the treatment, if any.

Generally, a 0.25 ml sample is sampled for a period of about 2 seconds to about 60 minutes, preferably about from about 6 seconds to about 45 minutes, and more preferably from about 10 seconds to about 20 seconds. 1clI+ flows through the absorbent material.

Supporting a binder after one or more steps and prior to a subsequent step The surface can be cleaned. For the Sand Inch test, following sample addition, Prior to and/or following the addition of tracers. Prior to its deployment, the surface can be cleaned. Washing with standard aqueous buffer , e.g. phosphate buffer and Tris buffer (tris(hydroxymethyl)amino) Contains nomethane). Washing should also be done using detergents and chemicals to minimize non-specific binding. Contains otrobism agents.

The method for determining the analyte in the sample being assayed may be used with a ligand or tracer. Depends on the type of marker used. If the dye is used as a label or marker If so, it appears on the surface of the first absorbent material and what cleaning steps it has gone through. It remains on its surface even afterward. In the case of enzymatic labeling, a suitable usually precipitating substrate provides the color. used to provide. Radioactivity, fluorescence, luminescence, enzymes remaining on the surface of the first absorbent material , chromogens, spin labels, biotin, gold particles, or other The presence of type markers can be determined by any conventional technique.

The test can be a qualitative or quantitative test, and the term "determined" as used here means Refers to the qualitative and/or quantitative determination of an analyte.

According to another aspect of the invention, a test piece comprising a first absorbent material and a second absorbent material is provided. equipment is provided. The first absorbent material has at least one binding agent for the analyte. Has a surface. The second absorbent material is adjacent to the first absorbent material. The first absorbent material is located next to the second absorbent material. has a lower porosity than the first one, so that the flow of liquid applied to the surface of the device is It is guided from the absorbent material to the second absorbent material. In a preferred embodiment, the second absorbent material is Surrounding the absorbent (for example, the first absorbent material is in the shape of a cylinder, and the second absorbent material is in the form of a cylinder as described above) The street surrounds or surrounds this column. In this aspect of the invention, A small amount of sample that is suspected to contain the analyte is placed on the top of the first absorbent material. The surface includes at least one binding agent for the analyte. The sample is contact with at least one binding agent for a sufficient time to form an analyte binding agent complex; Continue. During this verification process, someone must use one or more of the verification devices. When a person wants to clean through the above cleaning steps, he uses a large amount of cleaning agent. No. Because the first absorbent material has a lower porosity than the second absorbent material, the first absorbent material has a lower porosity than the second absorbent material. This is directed towards a second absorbent material which absorbs a greater amount of cleaning agent.

In one embodiment, the first absorbent material is about 1.8 g/cc to about 5.0 g/cc; Non-porous, preferably with a density of about 2.2 g/cc to about 4.4 g/cc It is a sexual material. In one embodiment, the first absorbent material is a non-porous ceramic.

In one embodiment, the second absorbent material is about 0.01 cc/g to about 0.400 cc/g. /g, preferably from about 0.010 cc/g to about 0.280 cc/g. and from about 0.005μ to about 125μ, preferably from about 0.030μ to about It has a diameter of up to 5μ. In one embodiment, the second absorbent material is a porous ceramic material. It is.

The first and second absorbent materials are formed from materials as described above. but formed a porosity gradient is formed from the high porosity second absorbent material to the low porosity first absorbent material; This has a surface with at least one binding agent for the analyte.

The surfaces of the first and second absorbent materials are arranged to speed up or slow down the absorption time. Pretreated with humectants or polymers.

The first and second absorbent materials contain a surfactant and a surfactant to further reduce non-specific binding. Or processed with proteins.

The assays employed when using this assay device include those described above. This outfit When performing assays using is placed on at least a portion of the surface of the first absorbent material. Contains the subject A small amount of sample, which is thought to be It remains on the surface of the first absorbent material for a sufficient time to form a binder complex. sample After application and before addition of tracer and/or post-development reagents, The harvested material is cleaned with a fairly large amount of cleaning agent. Cleaning agent is applied to this equipment Therefore, when cleaning the sample from the first absorbent material, the cleaning agent becomes less porous. from the first absorbent material to the more porous second absorbent material, which allows the sample to be and the cleaning agent is absorbed into the second absorbent material. This test method must be adopted. This applies to any additional washing steps.

However, it should be noted that the scope of the invention is not limited to the specific embodiments described above. must be understood. This invention may be practiced otherwise than as specifically described. Yes, and within the scope of the following claims.

Claims (14)

[Claims] 1. an assay device comprising a sample receiving surface, the sample receiving surface comprising: a sample receiving surface; the surface includes a first surface portion and a second surface portion; the first surface has at least one binding agent for one analyte; one surface portion is formed from a first absorbent material, and the second surface portion is formed from a liquid sample to be assayed. directing a flow of pull to the at least one binding agent for the analyte; a portion is adjacent to the first surface portion, the second surface portion is formed from a second absorbent material, and The second absorbent material has a porosity lower than that of the first absorbent material. Verification device. 2. The first absorbent material has pores of about 0.001 cc/g to about 0.400 cc/g. 2. Device according to claim 1, characterized in that it has a volume. 3. The first absorbent material has a pore volume of about 0.010 cc/g to about 0.280 cc/g. 3. A device according to claim 2, characterized in that it has: 4. The second absorbent material has a density of about 1.8 g/cc to about 5.0 g/cc. 2. A device according to claim 1, characterized in that: 5. The second absorbent material has a density of about 2.2 g/cc to about 4.4 g/cc. 5. A device according to claim 4, characterized in that: 6. Claim 1, wherein the first absorbent material is a porous ceramic material. The device according to item 1. 7. Claim 1, characterized in that the second absorbent material is an unpolished ceramic material. The device according to item 1. 8. an assay device comprising a sample receiving surface, the sample receiving surface comprising: a sample receiving surface; the surface includes a first surface portion and a second surface portion; the first surface has at least one binding agent for one analyte; one surface portion is formed from a first absorbent material, and the second surface portion is adjacent to the first surface portion. , the second surface portion is formed from a second absorbent material, wherein the first absorbent material is formed from the second absorbent material. the first surface has a porosity less than that of the absorbent material, and wherein the first surface has a porosity less than that of the absorbent material; characterized in that the flow of liquid applied to the second surface can be directed to the second surface. Verification device. 9. The first absorbent material has a density of about 1.8 g/cc to about 5.0 g/cc. 9. A device according to claim 8, characterized in that: 10. The first absorbent material has a density of about 2.2 g/cc to about 4.4 g/cc. 10. A device according to claim 9, characterized in that: 11. The second absorbent material is about 0.001 cc/g to about 0.400 cc/g. 9. Device according to claim 8, characterized in that it has a pore volume. 12. The second absorbent material is about 0.010 cc/g to about 0.280 cc/g. 12. Device according to claim 11, characterized in that it has a pore volume. 13. Claims characterized in that the first absorbent material is a non-porous ceramic material. Apparatus according to clause 8. 14. Claims characterized in that the second absorbent material is a porous ceramic material. Apparatus according to clause 8.