Chemistry: analytical and immunological testing – Involving an insoluble carrier for immobilizing immunochemicals – Carrier is organic
Patent
1995-01-17
1997-06-17
Spiegel, Carol A.
Chemistry: analytical and immunological testing
Involving an insoluble carrier for immobilizing immunochemicals
Carrier is organic
435 71, 435 792, 435 793, 435 794, 435 795, 435 96, 435962, 435968, 435971, 435975, 435500, 435538, 435539, 435540, 435543, 435808, 435817, 435822, 435824, G01N 3353
Patent
active
056396708
DESCRIPTION:
BRIEF SUMMARY
The present invention relates to a method for the determination of the amount of a ligand in a sample of a biological fluid, in which the biological fluid containing the ligand to be determined is incubated in the presence of a dissolved ligand derivative, which can be converted into an insoluble form, in the liquid phase with less than the stoichiometric amount of a labelled specific binder which binds both the ligand to be determined and the ligand derivative so that the ligand to be determined and the ligand derivative compete for the labelled specific binder, and in which the ligand derivative is converted into an insoluble form and, after separation of the insoluble components of the assay system from those components of the assay system remaining in the liquid phase, the amount of the ligand to be determined in the biological fluid is calculated from the amount of the labelled specific binder bound to the ligand derivative, and a kit for carrying out such a method.
The present invention relates in particular to a method of the stated type for the determination of the free portion of a monovalent ligand, i.e. of a ligand which represents a monovalent antigen or a hapten, which is to say that it has only one binding site for the immunological binding by a specific binding partner, namely a thyroid hormone, which is present in biological fluids partly in a form bound to its physiological binding proteins and partly in unbound, i.e. free, form, and the object of the process is the quantitative determination of this free proportion.
The method according to the invention is one of the immunological assay methods in which labelled antibodies (labelled specific binders) are employed and which are also referred to as immunometric assay methods. It belongs to the version of such methods which is referred to as a competitive method. In contrast to typical sandwich methods, in which the total amount of the ligand to be determined is bound in a sandwich, a proportion of the labelled binding partner is bound to the ligand to be determined and a further proportion to a competing derivative of this ligand. Further statements on various types of immunometric assay methods are to be found in the introduction of EP-B-89806 in which, exactly as in EP-B-103605, the immunometric version of a method for the determination of free proportions of ligands is described, which method is described as the so-called analogue tracer assay method in EP-B-26103.
In immunometric methods of the type according to the invention for the determination of the free proportions of ligands, in particular monovalent ligands such as the thyroid hormones, the ligand to be determined and the ligand derivative taken in a known amount are allowed to compete for a minor amount of the labelled antibodies according to the above description of the fundamental method on which the present invention is based. The labelled antibody is bound essentially completely to the free ligand and the ligand derivative, and its distribution over the two binding partners reflects the concentration to be determined of the ligand in the sample. In order to obtain information about the concentration to be determined for the ligand and the distribution of the labelled antibody over the ligand to be measured and the ligand derivative, the reaction product of ligand derivative and labelled antibody is preferably immobilised for separation from the remaining assay solution, which, according to EP-P-103605, is effected by converting the ligand derivative into an insoluble form before or after incubation with the labelled antibody. In practice, the procedure generally adopted to date is to initially take the ligand derivative from the outset in a form immobilised by binding to the solid substrate.
Embodiments of a method of this type are described in EP-A-303284 and EP-A-324540.
In a version of the method of the stated type, which version is described in EP-A-149631, an exogenous binder for the ligand derivative is used in addition to the conventional components of the assay meth
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patent: H0001018 (1992-01-01), Hantke
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patent: 4839299 (1989-06-01), Charlton et al.
patent: 5278080 (1994-01-01), Midgley et al.
patent: 5304498 (1994-04-01), Ekins
Bergmann Andreas
Struck Joachim
B.R.A.H.M.S. Diagnostica GmbH
Spiegel Carol A.
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