Chemistry: analytical and immunological testing – Involving an insoluble carrier for immobilizing immunochemicals
Patent
1982-06-28
1985-04-30
Padgett, Benjamin R.
Chemistry: analytical and immunological testing
Involving an insoluble carrier for immobilizing immunochemicals
435 7, 435 68, 435810, 435811, 260112R, 424 85, 436531, 436548, G01N 3354
Patent
active
045145075
DESCRIPTION:
BRIEF SUMMARY
FIELD OF THE INVENTION
This invention relates to the field of immunometric assays. In particular it relates to an antibody excess immunometric assay for interferon involving the use of monoclonal antibody to interferon.
BACKGROUND OF THE INVENTION
The interferons are a group of related proteins present in the mammalian body. An interferon is a protein factor which exerts virus non-specific, antiviral activity at least in homologous cells through cellular metabolic processes involving synthesis of both RNA and protein. The interferons are classified into types on the basis of antigenic specificity, the designations being alpha, beta and gamma (these correspond to previous designations leucocyte, fibroblast and type II (immune) interferons respectively). In addition to its antiviral effect interferon has been implicated as a mediator or immune function of other cellular phenomena. Interferon research has been hampered by problems in its assay. The only widespread assays use tissue cultured cells and compare some parameter of viral growth (for example viral RNA synthesis or host cell death) in the presence and absence of interferon. These complex biological assays, though sensitive, are laborious and subject to inherent variability. In particular, components other than interferon present in the assay sample often influence viral growth. There is a need for a simple indirect interferon assay. Such an assay would find widespread application in at least three areas of interferon research:
(1) the monitoring of both laboratory and large-scale production and purification of interferon;
(2) the quantitation of interferon doses in research and clinical applications;
(3) the measurement of interferon in biological fluids.
The object of the present invention is to provide an antibody excess immuno assay for interferon which will fulfill this need.
The human body reacts to the presence of antigens by producing antibody molecules from its lymphocyte cells. Antibodies have the property of selectively binding to certain distinctive sites known as determinants on antibodies thereby rendering the antigen innocuous. The nature of the interaction between antigen and antibody is not fully understood but it is clear that antibodies have a physical affinity for specific determinants of antigenic material. A reaction between an antibody and a determinant on an antigen for which the antibody is specific results in an adduct, commonly referred to as an "immunocomplex". The formation of such species makes possible a wide variety of assays for antigenic material. Such assays are known generically as immuno assays.
Immunoassays fall broadly into two categories:
(1) Analyte excess; labelled antigen. (The term analyte is a term of art and in this context means "that to be analysed"). In this type of assay an antibody having specificity to the analyte is incubated with a solution containing the analyte and a known quantity of a labelled antigen. In this way a competitive equilibrium is set-up in which the unknown amount of analyte competes with the known amount of labelled antigen to form immunocomplexes with the antibody. A method of determining the number of immunocomplexes formed between labelled antigen and the antibody make it possible to deduce the amount of analyte. This type of assay has certain disadvantages in that the ultimate sensitivity of the assay is limited by the relative stability constants of the immunocomplexes formed.
(2) Antibody excess; labelled antibody. In this type of assay the analyte to be determined is incubated with an excess of labelled antibody molecules. The estimation of the amount of analyte is therefore linear and its maximum sensitivity is, in theory at least, one molecule of the analyte. A refinement of this method involves insolubilising an antibody to a solid substrate, in excess and allowing the analyte to form immunocomplexes therewith. Subsequently, an excess of labelled antibody to a second determinant on the analyte may be incubated with the solid substrate. This type of assay, commonly re
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Moskowitz M.
Padgett Benjamin R.
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