An Agarose Plate Method for Detecting Alloantisera to Coagulant Factor IX and Factor IX Antigen

Abstract

An agarose plate method for detecting human alloantibodies (inhibitors) to coagulant factor IX has been developed. The assay is based on the ability of such antisera to inhibit the coagulation of a mixture of haemophilia B plasma, normal plasma and platelet subsitute in an agarose matrix. The agarose plate method was also adopted to measure levels of FIX antigen (IX:Ag) in plasma. Using this technique, 12 of 17 obligate carriers of haemophilia B demonstrated reduced levels of IX:Ag. Three of the five carriers with normal IX:Ag levels were members of kindred in which affected individuals had normal or near normal levels of IX:Ag.

Antibody Technics and Blood Coagulation

“…Methods were developed for the quantitative as say of Factor VIII-related antigen in various condi tions. Immunoradiometric assays [285,288,289,[291][292][293][294][296][297][298][299], radioimmunoassays [286], electroim muno assays [290], radial immunodiffusion [300], and a staphylococcal coagglutination assay [301] were developed. These assays have been used to quantitate Factor VIII-related antigen for the dif ferentiation of hemophilia A and vW disease [286], to identify variants of the vW disease [288,290,291], to study clotting abnormalities [289], to determine antigen levels in adult and fetal blood [292][293][294], to investigate porcine Factor VIII-related antigen [296], to study Factor VIII concentrates [297], to assess alloantibodies [289,299], and to quantitate Factor VIII-related antigen [301].…”

“…Recently, an agarose plate method for detecting human alloantibodies to procoagulant Factor IX has been developed. With this technic, levels of Factor IX in plasma and hemo philia B carrier states can also be determined [300]. The same group of investigators developed a radio immunoassay for the detection of Factor IX anti bodies using radiolabeled Factor IX and formalinfixed heat killed Staph A as a solid phase adsorbent for immune complexes [301].…”

“…Radioimmunoassays [298,300,308] were devel oped and used to detect carriers [298], to measure alloantibodies [301] and to quantitate abnormal Factor IX in patients on warfarin [308]. Other tech nics were the immunoradiometric assay [299] and the agarose plate method [300]. Factor X is the inactive precursor of Factor Xa and is a vitamin K-dependent glycoprotein contain ing γ-carboxyglutamic acid residues.…”