Enzyme-linked immunosorbent assay for the determination of blood coagulation factor XIII A-subunit in plasma and in cell lysates.
A new one-step ELISA was developed for the determination of the concentration of blood coagulation factor XIII subunit A (FXIII-A) in plasma and in cell lysates. Monoclonal antibodies directed against different epitopes on FXIII-A were used for the assay. The capture antibody was biotinylated on its carbohydrate moiety and the detection antibody was labelled with horseradish peroxidase. The antigen-antibody reaction was carried out in the well of a streptavidin-coated microplate. Complex formation with FXIII subunit B (FXIII-B) and association to fibrinogen did not influence the accessibility of the antibodies to FXIII-A. The method could be performed within 2 h and demonstrated good reproducibility, recovery and sensitivity. Plasma samples could be assayed after storage at -20 degrees C for at least 6 months. However, in the case of platelet lysates freezing and rethawing resulted in a significant loss of FXIII-A. FXIII-A concentrations measured in the plasma samples of healthy individuals and patients correlated well with the concentrations of complexed plasma FXIII (A2B2) and with the results of FXIII activity measurements. A reference range of 46-82 fg/platelet was established for platelet FXIII-A.
Katona E
E
E
Department of Clinical Biochemistry and Molecular Pathology, Medical and Health Science Centre, University of Debrecen, P.O. Box 40, H-4012, Debrecen, Hungary.
Ajzner
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E
Tóth
K
K
Kárpáti
L
L
Muszbek
L
L
D000911:Antibodies, Monoclonal; D001792:Blood Platelets / Q000737:chemistry; D002458:Cell Fractionation; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D000939:Epitopes; D005176:Factor XIII / Q000032:analysis* / Q000276:immunology; D006801:Humans
D000911:Antibodies, Monoclonal; D000939:Epitopes; C064015:factor XIII subunit A; D005176:Factor XIII; C054380:factor XIIIb