Fluorescent excitation transfer immunoassay. A general method for determination of antigens.
J Biol Chem
; 251(14): 4172-8, 1976 Jul 25.
Article
en En
| MEDLINE
| ID: mdl-945272
ABSTRACT
A general immunochemical method for the assay of haptens and proteins has been devised and applied to morphine, a morphine-albumin conjugate, and human immunoglobulin G. A fluorescein-labeled antigen and a quencher-labeled antibody are employed. By use of fluorescein and rhodamine as the fluorescer and quencher, respectively, dipole-dipole-coupled excitation energy transfer can occur within the antigen-antibody complex. The resulting quenching of fluorescence can be inhibited by competitive binding with unlabeled antigen, Alternatively, separate antibody samples can be labeled with fluorescein and rhodamine, respectively. Unlabeled antigen causes aggregation of the separately labeled components with resultant quenching. Using the latter method, experiments suggest that up to about 20 anti-morphine antibody binding sites will associate with morphine-albumin conjugates. When an excess of the conjugate is present the antibodies appear to assemble in clumps on the protein surface. Mathematical analysis of the quenching of fluorescein-labeled morphine by rhodamine-labeled anti-morphine gives an approximate fit to the quenching data, but the calculations are very dependent on the assumptions used.
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Colección:
01-internacional
Banco de datos:
MEDLINE
Asunto principal:
Inmunoensayo
/
Antígenos
Límite:
Animals
/
Humans
Idioma:
En
Revista:
J Biol Chem
Año:
1976
Tipo del documento:
Article