Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving antigen-antibody binding – specific binding protein...
Reexamination Certificate
1999-09-29
2001-09-25
Stucker, Jeffrey (Department: 1648)
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving antigen-antibody binding, specific binding protein...
C436S526000, C436S528000, C436S534000, C436S536000, C436S538000, C436S806000, C422S236000
Reexamination Certificate
active
06294342
ABSTRACT:
BACKGROUND OF THE INVENTION
1. Field of the Invention
This invention relates to a method for determining the presence or amount of analyte in a test sample using magnetically responsive materials. More particularly, the invention relates to the use of magnetically responsive materials to change the properties of components in binding assays.
2. Discussion of the Art
Diagnostic assays have become an indispensable means for detecting analytes in test samples by using the mutual reaction between the analyte and a specific binding member for the analyte, such as the immunoreaction between an antigen and an antibody that binds to that antigen. Typically, detectable tags or labels attached to antibodies, which in turn bind to the analyte of interest, are employed in such diagnostic assays, wherein the detection of the resultant labeled antibody-analyte complex, or detection of the labeled antibody that does not bind to the analyte to form a complex, is used to indicate the presence or amount of analyte in the test sample.
Two commonly used diagnostic assay techniques employing specific binding members are the radioimmunoassay (RIA) and the enzyme immunoassay (EIA), both of which employ a labeled specific binding member. The RIA uses a radioactive isotope as the detectable tag or label attached to a specific binding member. Because the radioactive isotope can be detected in very small amounts, it can be used to detect or quantify small amounts of analyte. However, substantial disadvantages associated with the RIA include the special facilities and extreme caution that are required in handling radioactive materials, the high costs of such reagents, and their unique disposal requirements.
The EIA uses an enzyme as the detectable tag or label attached to a specific binding member, wherein the enzymatic activity of the enzyme is used to detect the immunoreaction. While the EIA does not have some of the same disadvantages of the RIA, EIA techniques typically require the addition of substrate materials to elicit the detectable enzyme reaction. In addition, enzyme substrates are often unstable and have to be prepared just prior to use or be stored under refrigeration. Moreover, enzyme labels may be difficult to purify and conjugate to binding members, and may be unstable during storage at room temperature or even under refrigerated conditions. Enzyme immunoassays are also unsatisfactory in that the methods typically require complex incubations, multiple liquid additions, and multiple wash steps.
More recently, assay techniques using metallic sol particles as visual labels have been developed. In these techniques, a metal (e.g., gold, silver, platinum), a metal compound, or a nonmetallic substance coated with a metal or a metal compound, is used to form an aqueous dispersion of particles. Generally, the specific binding member to be labeled is adsorbed onto the metallic sol particles, and the particles are captured or aggregated in the presence of analyte. Although the metallic sol particles have the advantage of producing a signal that is visually detectable as well as measurable by an instrument, they are difficult to measure quantitatively. The metallic sol particles also have a limited color intensity, and consequently, limited sensitivity in some assays. In addition, the surfaces of inorganic metallic sol particles, such as gold, may not readily accept the covalent attachment of specific binding members. Thus, during use in a binding assay, care must be taken so that the adsorbed specific binding members are not removed from the inorganic particles through the combination of displacement by other proteins or surface active agents and the shear forces that accompany washing steps used to remove non-specifically bound material. Metallic sol particles can be difficult to coat without inducing aggregation; they may aggregate upon storage or they may aggregate upon the addition of buffers or salts. Furthermore, such particulate labels are difficult to concentrate and can be difficult to disperse.
Other materials for labels include chemiluminescent and fluorescent substances. However, these substances can be unstable, and fluorescent materials may undergo quenching. Non-metallic particles, such as dyed or colored latex particles and selenium particles, have also been used as visual labels.
Self-performing immunoassay devices have proven to be of great benefit in the field of diagnostics. A self-performing immunoassay device is a kit containing immunoreagents to which a biological sample can first be added by the patient or laboratory technician, then the diagnostic assay performed without the need for complex laboratory instruments. Commercially available self-performing immunoassay devices, such as the strip assay device having the trademark “TESTPACK PLUS”, distributed by Abbott Laboratories, enable immunoassays to be performed quickly and reliably.
Typically, self-performing immunoassay devices involve chromatographic test strips. For example, U.S. Pat. No. 4,960,691 discloses a test strip for analysis of an analyte in a sample by means of a sequential series of reactions. The test strip comprises a length of chromatographic material having capillarity and the capacity for chromatographic solvent transport of non-immobilized reagents and reactive components of a sample by means of a selected chromatographic solvent. The test strip includes (1) a first end at which chromatographic solvent transport begins, (2) a second end at which chromatographic solvent transport ends, and (3) a plurality of zones positioned between the first and second ends. These zones include (1) a first zone impregnated with a first reagent which is mobile in the solvent and capable of a specific binding reaction with the analyte, (2) a second zone for receiving the sample, and (3) a third zone, downstream of the second zone, impregnated with a second reagent that is immobilized against solvent transport and is capable of a specific binding reaction with the analyte so as to immobilize the analyte in the third zone. The test strip is designed so that the first reagent can be detected at the third zone as a measure of the analyte.
A common feature of chromatographic test strips involves the flow of a fluid or a mixture of a fluid and particles through a porous matrix. The test strip typically includes a reaction zone where binding reactions can occur. For proper binding reactions to occur in chromatographic test strips, the fluid or mixture must flow substantially uniformly through the reaction zone.
A problem with assay devices of this type is the inherent variability in the material from which the porous matrix is formed. This variability (for example, in porosity) directly affects the flow of fluid through the matrix and may adversely affect the precision of the assay device. Furthermore, the matrix will often non-specifically bind the particles or reagents at sites at the intended reaction zone or elsewhere, thereby necessitating the use of elaborate passivating procedures after the immobilized reagent has been applied. Consequently, there is a desire to develop a rapid, simple, self-performing assay device that does not require a fluid to flow through a porous matrix.
Another problem with self-performing immunoassay devices is the necessity of immobilizing a specific binding reagent on the test strip so that reagents involved in the assay can be captured at the reaction zone. The process of immobilizing the specific binding reagents on the test strip can be difficult to control, leading to lot-to-lot variations in the binding capacity of the reaction zone. Furthermore, the immobilized binding reagents can be unstable, causing the binding capacity of the reaction zone to change after shipping or storage. Because the immobilized specific binding reagent is specific for the assay of a particular analyte, test strips must be dedicated to a particular assay. An additional problem with self-performing immunoassay devices is lot-to-lot variation resulting from manufacturing processes, especially variation of the activit
Elstrom Tuan A.
Howard Lawrence V.
Rohr Thomas E.
Shain Eric B.
Abbott Laboratories
Stucker Jeffrey
Weinstein David L.
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