Use of progesterone-3(O-carboxymethyl oxime)-horseradish peroxidase in a sensitive microtitre-plate EIA and its application to a visual membrane EIA of progesterone

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Title
Use of progesterone-3(O-carboxymethyl oxime)-horseradish peroxidase in a sensitive microtitre-plate EIA and its application to a visual membrane EIA of progesterone
Author(s)
Do Young Yoon; Eun Young Song; D H Kwon; Myung Ja Choi; Si Myung Byun; In Seong Choe; Tai Wha Chung
Bibliographic Citation
Journal of Immunoassay and Immunochemistry, vol. 16, no. 2, pp. 137-153
Publication Year
1995
Abstract
A simple method of visual membrane enzyme-immunoassay (EIA) for the detection of progesterone is described. When two types of progesterone- horseradish peroxidase (HRP) tracers were challenged for binding, in the presence of progesterone, to the monoclonal anti-progesterone antibody, 15A, coated on the microtitre plate, the HRP conjugated at the C-3 position (A- ring) of progesterone competed more effectively with progesterone to the binding site of the monoclonal antibody (mAb) than HRP conjugated at the C- 11 position of the C-ring. By using this combination of mAb, 15A, and progesterone-3(O-carboxymethyloxime)-HRP (P-3CMO-HRP), we developed a visual membrane EIA system in which free progesterone in the sample could be quantified by the degree of color development. In this system, free progesterone competed with P-3CMO-HRP for binding sites of mAb immobilized on the nitrocellulose membrane. The stable grey color was formed on the surface of membrane for progesterone-negative and no color for progesterone-positive sample using 3,3'-diaminobenzidine (DAB) with Co2+ as an insoluble substrate solution. To examine whether tetramethylbenzidine (TMB) can substitute for DAB in membrane EIA, an experiment was conducted where TMB was used as an insoluble substrate.
Keyword
progesteronevisual membrane immunoassay
ISSN
0197-1522
Publisher
Taylor & Francis
Type
Article
Appears in Collections:
1. Journal Articles > Journal Articles
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