Amplification method for solid phase immunoassays

Abstract

The present invention is directed to immunoassays for detecting one or more target analytes in a fluid sample wherein the detection reaction occurs on a solid support and involves an amplification system. In particular, the invention is directed to making and using a test device having at least one site for detecting the presence of at least one target analyte, wherein a conjugate comprising dextran-polystreptavidin is immobilized at the test site(s) as a capture reagent for a complex containing the target analyte.

Description

CROSS REFERENCE TO RELATED APPLICATION [0001] The present application claims priority from U.S. Provisional Patent Application Ser. No. 60 / 604,378 filed on Aug. 25, 2004, which is expressly incorporated by reference herein.FIELD OF THE INVENTION [0002] The present invention is directed to immunoassays for detecting one or more target analytes in a fluid sample wherein the detection reaction occurs on a solid support. BACKGROUND OF THE INVENTION [0003] Immunoassays have become methods of choice for rapid in vitro diagnostics due in part to the ease of use, speed of reaction, and relatively low cost. A common immunoassay format comprises a “capture” reagent immobilized on a solid substrate wherein the capture reagent is capable of binding the target analyte, or a complex containing the target analyte, or a molecule that competes with the target analyte for binding sites on the capture reagent. By localizing the capture reagent in a restricted zone or area, the result of the immunoassa...

AI Technical Summary

Benefits of technology

[0010] Streptavidin moieties within the immobilized dextran-polystreptavidin conjugate bind specifically to biotin; thus a biotinylated component, such as a biotinylated binding partner to the analyte, is used. For example, biotinylated antibodies specific for a target analyte can be utilized, or if the target analyte is itself an antibody, then a biotinylated antigen can be used. Likewise, hormones, receptors, and the like can serve as target analytes, and appropriate ligands that specifically bind to the hormones, receptors, or other target analyte are conjugated to biotin. In one embodiment, biotinylated conjugates are dried or lyophilized in a separate area from other dried components, such as dried labeled conjugates, because separating the biotinylated conjugate in a separate site from the site containing a labeled conjugate appears to further improve the sensitivity of the device using dextran-polystreptavidin as a capture component. In one embodiment, the biotinylated conjugate, such as a biotinylated antibody or antibody fragment capable of specifically binding to a target analyte, is placed upstream of the labeled component, which is, in turn, upstream from the test site(s).

Problems solved by technology

Such restrictions on processing present a challenge when the biologically or physiologically meaningful concentration of a target analyte is low.

Addition of such reagents leads to further dilution of the concentration of target analytes, compounding the problem of detecting low levels of target analytes that may be present in a mucus specimen, throat swab, semen, and the like.

However, conjugation of streptavidin to latex particles adds to the labor and cost of manufacturing the test device and entrapping particles within the pores of the carrier material is not equivalent to chemically or physically binding, directly or indirectly, the capture reagent to the carrier material.

Moreover, entrapment of latex particles within a carrier material requires porosity sufficient to contain particles, but is not desirable for use in connection with samples containing cellular contaminants and / or debris, such as sputum samples, semen, vagina and cervical secretions, whole blood, and the like.

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