Studies on the measurement of protein S in plasma.
The anticoagulant factor Protein S circulates in two major forms, free and bound to C4b binding protein (C4BP). We report a monoclonal antibody-based enzyme-linked immunosorbent assay of human Protein S that is sensitive (detection limit 30 ng) and free of notable competition from other vitamin K-dependent proteins. We demonstrate assay sensitivity to Protein S-C4BP complex, as well as to free Protein S by (a) results of immunoaffinity chromatography involving the assay antibody; (b) parallelism between the standard curve and a plasma dilution curve, including dilutions greater than 100-fold; (c) lack of effect of added C4BP on Protein S values; and (d) reactivity in the assay by high-molecular-mass plasma Protein S in gel permeation studies. These latter studies also indicate that uncomplexed Protein S has an apparent molecular mass of approximately 150,000 Da, attributable to either polymerization or conformational considerations. Inter- and intra-assay CVs were 9.8% and 5.1%, respectively, and 90% of Protein S added to plasma was recovered. Reference ranges (mean and SD) for total Protein S in middle-aged normal subjects were 16.6 (1.9) mg/L for men (n = 54) and 16.0 (3.0) mg/L for women (n = 38). In the polyethylene glycol precipitation method, the reference values for free Protein S were 5.4 (1.6) mg/L for men and 4.4 (1.1) mg/L for women.
Bovill
E G
EG
Department of Pathology, University of Vermont, Burlington 05405.
Landesman
M M
MM
Busch
S A
SA
Fregeau
G R
GR
Mann
K G
KG
Tracy
R P
RP
D000368:Aged; D015153:Blotting, Western; D002352:Carrier Proteins / Q000378:metabolism; D015935:Complement C4b / Q000378:metabolism; D005260:Female; D006023:Glycoproteins / Q000097:blood*; D006801:Humans; D007118:Immunoassay / Q000379:methods; D016167:Integrin alphaXbeta2; D008297:Male; D008875:Middle Aged; D011498:Protein Precursors / Q000097:blood*; D017293:Protein S
D002352:Carrier Proteins; D006023:Glycoproteins; D016167:Integrin alphaXbeta2; D011498:Protein Precursors; D017293:Protein S; C050930:protein S precursor; D015935:Complement C4b