Chemistry: analytical and immunological testing – Thyroid hormone tests
Patent
1998-07-08
2000-03-14
Chin, Christopher L.
Chemistry: analytical and immunological testing
Thyroid hormone tests
436501, 436503, 436504, 436517, 436518, 436523, 436524, 436528, 436533, 436537, 436541, 436542, 436543, 436544, 436545, 436546, 435 4, 435 6, 435 71, 435 79, 435 792, 435 794, 435 795, 435 76, 435 78, G01N 33566, G01N 33557, C12Q 168, A61K 39395
Patent
active
060371854
DESCRIPTION:
BRIEF SUMMARY
This invention relates to a method for the non-competitive determination of analytes.
BACKGROUND OF THE INVENTION
The publications and other materials used herein to illuminate the background of the invention, and in particular, cases to provide additional details with respect to the practice, are incorporated by reference.
The application of biospecific binding partners for the determination of analytes from complex samples has gained widespread use in in vitro diagnostics. At present, most of such determinations use antibodies--either polyclonal or monoclonal--as the biospecific binding partner. The determinations that use antibodies are often called immunoassays. Immunoassays are often divided into non-competitive and competitive ones, where non-competitive assays involve the use of an excess of reagents and two biospecific binding partners binding to the same analyte (this type of assay is commonly called the sandwich assay). Competitive assays on the other hand rely on the measurement of the ratio between the free and bound labelled marker, with the ratio being modified by the amount of the analyte in the sample.
In usual non-competitive immunoassays, a sample containing the antigen to be determined is incubated with an excess of a capture antibody immobilized to a solid support. A labelled antibody, specific for another epitope on the same antigen, is added in excess. A sandwich comprising "catching antibody--antigen--labelled antibody" is thus formed. After completion of the incubation, the unbound labelled antibody is removed and the signal from the label in the sandwich is measured. The signal is thus directly proportional to the antigen concentration in the sample.
The above non-competitive method cannot, however, be easily applied to the determination of small molecular weight analytes. The small molecular weight analyte is too small to simultaneously bind to two different antibodies. The immunoassay of these analytes is therefore normally performed by a competitive assay. In a typical implementation of competitive assay, the sample containing the analyte to be determined as well as a labelled derivative of the analyte are added to an immobilized antibody specific for said analyte. The unlabelled and labelled analytes compete for the binding sites on the antibody. After completion of the incubation, unbound analytes are removed and the signal from the bound labelled analyte is detected. Increased concentrations of the analyte in the sample will thus result in a decreased signal. When the signal strength is plotted as a function of increasing analyte concentration, a sigmoidal, descending curve is obtained. The sensitivity of this competitive assay is not as high as that of non-competitive assays due to the fact that the signal is at its highest value at the zero dose. As Ekins et al. [1,2] have pointed out, sensitivity can be defined as the smallest dose distinguishable from the zero dose. A usual criterion for this distinction is that the signal of the dose differs by more that two standard deviations (SD) from the signal of the zero dose. In a competitive assay one has to detect a small difference between two high signals, whereas in a non-competitive assay one has to detect a small difference between two low signals. As the SD of the low signal tends to be less than that of the high signal, the non-competitive assay is able to detect smaller differences in signal than the competitive assay. Assuming that the slopes of the dose-response curves of both assays have the same absolute value in the low dose range, then non-competitive assay is more sensitive, because its ability to detect smaller differences in signal is in direct proportion to its ability to detect smaller differences in the dose, too.
We have recently discovered a new non-competitive method for the determination of small molecular weight analytes such as haptens. Contrary to the usually employed competitive assay described above, the new method gives a linear, ascending curve when signal strength is plotted as a function of in
REFERENCES:
patent: 5476770 (1995-12-01), Pradelles
patent: 5516635 (1996-05-01), Ekins et al.
patent: 5573920 (1996-11-01), Randle
patent: 5573921 (1996-11-01), Behnke et al.
patent: 5710009 (1998-01-01), Fitzpatrick et al.
patent: 5817525 (1995-05-01), DeAlwis
patent: 5863740 (1995-09-01), Kientsch-Engel et al.
Pradelles et al., "Immunometric Assay of Low Molecular Weight Haptens Containing Primary Amino Groups," 66 Anal. Chem. 16-22 (1994).
Patent Abstracts of Japan, vol. 4, No. 142, p. 30 JP 55-90858, (1980) (Abstract).
Chin Christopher L.
Do Pensee T.
Orion-yhtyma Oy
Wallac Oy
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