A simple method to quantify staphylococcal protein A in the presence of human or animal IgG in various samples.
J Immunol Methods
; 235(1-2): 61-9, 2000 Feb 21.
Article
em En
| MEDLINE
| ID: mdl-10675758
ABSTRACT
Immunoassays designed to measure low concentrations of staphylococcal protein A (SPA) that have been leached into antibody preparations intended for therapeutic use are subject to differing degrees of interference. Methods established to quantify SPA in murine antibody preparations are not accurate in the presence of human or humanized IgG. We report the development of an enzyme-linked immunosorbent assay (ELISA) for SPA with a detection limit of 7 pg/ml and the optimization of a method that permits complete dissociation of SPA-immunoglobulin-complexes. This assay is a modification of our heat-mediated dissociation (HD-SD) treatment with sodium dodecyl sulfate (SDS) and diethylenetriaminepentacetic acid (DTPA) for total immune-complex dissociation, in which the heat treatment has been prolonged and the diluent is characterized by increased protein content and buffering capacity. The diluent developed contains SDS, DTPA and bovine serum albumin dissolved in a 0.1 M phosphate buffer (pH 7.2). To validate the efficiency of this novel method, a series of samples have been assayed, including samples reconstituted in vitro, samples of purified antibodies, and plasma from patients. The described method has been shown to be generally efficient in quantitating all native and recombinant SPA in samples containing up to 50 mg/ml of human IgG. These data demonstrate the utility of this technique in determining SPA contamination of recombinant immunoglobulin therapeutic products.
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Base de dados:
MEDLINE
Assunto principal:
Proteína Estafilocócica A
/
Ensaio de Imunoadsorção Enzimática
Tipo de estudo:
Diagnostic_studies
Limite:
Humans
Idioma:
En
Ano de publicação:
2000
Tipo de documento:
Article