A sensitive immunochemical assay for measuring the concentration of the activated protein C-protein C inhibitor complex in plasma: use of a catcher antibody specific for the complexed/cleaved form of the inhibitor.
Activated protein C (APC) is a serine proteinase that regulates blood coagulation. In plasma it is inhibited mainly by the protein C inhibitor (PCI). The plasma concentrations of APC-PCI complex is increased in hypercoagulative states such as deep venous thrombosis. Formation of the APC-PCI complex induces a drastic conformational change in PCI that exposes new epitopes (neoepitopes) on the molecule. We have devised a simple immunofluorometric sandwich assay for measurements of the concentrations of APC-PCI complex, employing as the catcher, a monoclonal antibody that has a high affinity (K(D) = 4 x 10(-11) M) for a complexation-specific neoepitope that is expressed on PCI. A monoclonal antibody against protein C is employed as the tracer. The method gives a linear dose-response curve (0.06-50 microg/l), has a low detection limit (0.06 microg/l) and no crossreactivity with native PCI at physiologic plasma concentrations. We have now determined the concentration of the APC-PCI complex in healthy individuals.
Strandberg
K
K
Department of Clinical Chemistry, University Hospital, Malmö, Lund University, Sweden.
Kjellberg
M
M
Knebel
R
R
Lilja
H
H
Stenflo
J
J
D002138:Calibration; D003429:Cross Reactions; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods / Q000592:standards; D000939:Epitopes / Q000276:immunology / Q000378:metabolism; D015200:Fluoroimmunoassay / Q000379:methods / Q000592:standards; D006801:Humans; D011485:Protein Binding / Q000276:immunology; D011486:Protein C / Q000032:analysis* / Q000276:immunology / Q000378:metabolism; D018046:Protein C Inhibitor / Q000097:blood* / Q000276:immunology / Q000378:metabolism; D011487:Protein Conformation; D012015:Reference Standards; D012680:Sensitivity and Specificity
D000939:Epitopes; D011486:Protein C; D018046:Protein C Inhibitor