Activity of antiphospholipid antibody ELISA cofactor in different animal sera

A plasma protein cofactor, beta 2-glycoprotein I (beta 2GPI), also known as apolipoprotein H, is necessary to detect certain antiphospholipid antibodies (aPA) to negatively charged phospholipids (PL) in the ELISA. Inasmuch as sera are diluted 1:100 before testing, the concentration of native beta 2GPI may be insufficient to provide an optimal aPA ELISA signal. Therefore, many laboratories add adult bovine serum (ABS) to the diluent buffer to provide a consistent level of cofactor for optimal aPA binding. To determine if other animal sera can provide the cofactor, cat, chicken, dog, horse, goat, guinea pig, mouse, pig, rat, and sheep were tested as diluent supplements in the aPA ELISA. To measure cofactor activity in these animal sera, ELISA for aPA to anionic phospholipids were performed. Two aPA positive patient plasmas were selected for study; one with cofactor-dependent and one with cofactor-independent aPA. Only four of the animal sera tested (bovine, pig, sheep, and cat) supported the cofactor-dependent aPA in ELISA. The cofactor-independent aPA was positive in the presence of each animal serum except bovine and rat. In order to determine whether these animal sera contain a beta 2GPI-like molecule, Western blot analyses were performed. By using a polyclonal antiserum produced to human beta 2GPI, specific beta 2GPI-like cross-reactivity was observed with all animal sera except the chicken. In summary, cofactor activity in animal sera varied significantly; however, bovine and pig sera appear to allow optimal binding of cofactor dependent aPA.