Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving virus or bacteriophage
Reexamination Certificate
1994-06-17
2003-11-25
Stucker, Jeffrey (Department: 1648)
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving virus or bacteriophage
C435S007100, C435S007900, C435S007920, C435S007940, C435S007950, C435S971000, C435S973000, C435S974000, C435S967000, C436S825000, C436S536000, C436S541000, C436S544000, C436S545000, C436S546000, C436S548000, C436S063000, C436S811000, C436S809000, C436S810000, C423S055000, C423S056000, C423S058000, C423S060000
Reexamination Certificate
active
06653066
ABSTRACT:
FIELD OF THE INVENTION
The present invention relates to devices and methods for the detection or quantification of polyvalent substances such as immunoglobulin, and more particularly to devices and methods for performing inverse sandwich assays for the detection or quantification of polyvalent substances such as immunoglobulin.
BACKGROUND OF THE INVENTION
There are a wide variety of naturally occurring and synthetically produced polyvalent substances of scientific and commercial interest. Prototypical of such substances are immunoglobulin or antibody, which are proteins usually existing as glycoproteins in vivo. The protein moiety of an immunoglobulin comprises a basic unit of two light polypeptide chains and two heavy polypeptide chains, for a total of four polypeptide chains. An immunoglobulin molecule can have more than one of these basic units or can incorporate polypeptide chains in addition to those found as part of a basic unit.
One function of an immunoglobulin molecule is to bind to a substance at a site on the substance called an “epitope”. The “epitope binding site” on an immunoglobulin molecule is comprised of a segment of one light polypeptide chain and a segment of one heavy polypeptide chain such that the basic unit of four polypeptide chains defines two epitope binding sites.
Naturally occurring immunoglobulins are divided into classes and subclasses on the basis of amino acid sequence differences in the heavy polypeptide chain. In humans, there are five classes, IgD, IgE, IgG each having two epitope binding sites, IgA having two or four epitope binding sites, and IgM having ten epitope binding sites.
Antibodies are of particular scientific and commercial interest among the types of polyvalent substances because of their role in the mammalian immune response. They are markers of current and past infection, and important in host reactions against foreign substances such as viruses, bacteria, tissue grafts, toxins and drugs. Further, the inappropriate production of antibodies against host tissues contributes to many diseases including myasthenia gravis, hemolytic anemia, scleroderma, Graves' disease and rheumatoid arthritis.
Because of their importance, a variety of methods have been developed for the detection or quantification of antibody, including an antiviral neutralization bioassay and a radioimmunoassay. In most antibody detection or quantification methods, sample containing analyte antibody is applied to the surface of a device having an suitable immobilized antigen on the surface. The analyte antibody is allowed to bind to the immobilized antigen creating immobilized antigen/analyte antibody complexes. A ligand is then applied to the immobilized antigen/analyte antibody complex-coated surface and binds to the analyte antibody of the immobilized antigen/analyte antibody complex at a site other than one of the analyte antibody molecules' epitope binding sites. The ligand contains a chromogen, enzyme or radioactive label. Detection of the chromogen, enzyme or radioactive label remaining on the surface is a measure of the presence or quantity of the analyte antibody.
More recently, there has been described a sandwich-type of enzyme immunoassay for the detection or quantification of antibody, dubbed an “inverse sandwich” immunoassay. This assay utilizes the presence of the two epitope binding sites on an immunoglobulin molecule which allows each immunoglobulin molecule to simultaneously bind to two separate antigen moieties having a substantially-identical epitope.
In the inverse sandwich immunoassays heretofore described, an antigen is immobilized to the surface of plastic beads or a microtitre plate. Disadvantageously, the known techniques for utilizing an inverse sandwich immunoassay for the detection or quantification of immunoglobulin requires specialized equipment not readily available to a consumer or present in a physician's office. A further disadvantage of these prior techniques of performing an inverse sandwich immunoassay using beads or microtitre plates is that tight control is required over several of the parameters involved in performing the assay. Also disadvantageously, using beads or microtitre plates to perform an inverse sandwich immunoassay necessitates one or two washing steps.
SUMMARY OF THE INVENTION
According to one aspect of the present invention, there is provided a device for detecting the presence or amount of analyte antibody in a sample. The device includes a first zone that includes a first matrix and a first substance that includes an epitope that is capable of binding to the analyte antibody. The first substance has a label. The device further includes a second zone spatially separate from the first zone and in fluid communication with the first zone. The second zone includes a second matrix and a second substance that includes an epitope substantially identical to the epitope on the first substance. The second substance is immobilized on the second matrix and the first substance is capable of moving from the first zone to the second zone after application of the sample to the first zone.
According to another aspect of the present invention, the device can be used to detect antibody to Human Immunodeficiency Virus (HIV). In one preferred embodiment, the device can detect the presence or amount of antibody to HIV-1 glycoprotein 41. In another preferred embodiment, the device can detect the presence or amount of antibody to HIV-2 glycoprotein 36.
According to another aspect of the present invention, there is provided a method for detecting the presence or amount of analyte antibody to Human Immunodeficiency Virus in a sample. First, a device according to one of the foregoing aspects of the present invention is provided. Next, sample is applied to the first zone, the antibody in the sample comprising first and second epitope binding sites. After binding the first epitope binding site on the analyte antibody to the first epitope on the first substance, thereby forming an analyte antibody/first substance complex, the complex is allowed to migrate from the first zone to the second zone. Next, the second epitope binding site on the analyte antibody of the first substance/analyte antibody is allowed to bind to the second epitope on the immobilized second substance in the second zone, thereby forming immobilized first substance/analyte antibody/second substance. Finally, the label upon the first substance of the immobilized first substance/analyte antibody/second substance is detected. In a preferred embodiment, the label includes a chromogen and the detecting step includes visually identifying a color change on the second zone.
In another aspect of the invention, there is provided a device for detecting the presence or amount of analyte antibody to each of two antigens, antigen-1 and antigen-2, in a sample. In a preferred embodiment, antigen-1 is Human Immunodeficiency Virus 1 and antigen-2 is Human Immunodeficiency Virus 2. The device includes a first zone that includes a first matrix, an antigen-1 first substance and an antigen-2 first substance. The antigen-1 first substance includes an epitope from antigen-1, and the antigen-2 first substance includes an epitope from antigen-2. Each of the first substances has a label. The device further includes a second zone spatially separate from the first zone and in fluid communication with the first zone. The second zone includes a second matrix, an antigen-1 second substance and an antigen-2 second substance. The antigen-1 second substance includes an epitope from antigen-1 that is substantially identical to the epitope from antigen-1 on the antigen-1 first substance. The antigen-2 second substance includes an epitope from antigen-2 that is substantially identical to the epitope from antigen-2 on the antigen-2 first substance an epitope. Each of the second substances is immobilized on the second matrix. Each of the first substances is capable of moving from the first zone to the second zone after application of the sample to the first zone. In one particularly preferred em
Knobbe Martens Olson & Bear LLP
Stucker Jeffrey
Trinity Biotech
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