Evaluation of the Affinity Measurement of Anti-HIV-1 P17 Monoclonal Antibody by BIAcore

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Abstract
The methods of measuring the affinity constants of anti-HIV-1 p17 monoclonal antibodies (MAbs) using the double antibody methods in the liquid phase and the biomolecular interaction analysis by BIAcore system (Pharmacia Biosensor AB, Uppsala, Sweden) were compared. MAbs, HyHIV1-6, recognizing residues 12-29 (P12-29) of pl7 and the naive protein, pl7, were used. The kinetic association constants (KAs) obtained using the double antibody method were 2.40 X 107 - 1.40 X 108M-1 for P12-29, and 4.80 x 106 - 1.80 x 107M-1 for pl7. In the BIAcore system where P12-29 or pl7 was used as immobilized antigens onto the sensorchip, the KAs were 1.57 x 109 - 4.81 x 109M-1 for P12-29, and 1.52 x 109 - 1.21 X l010M-1 for pl7. On the other hand, when MAbs were immobilized onto the sensorchip and P12-29 or rpl7 was used as analyte, the KAs for P12-29 and pl7 were in the region 3 X l08 - 3 X 109, 1 X 108 - 3 X 109M-1, respectively. These data show that the KAs were higher than those obtained using the double antibody method, however, no significant difference could be observed. Moreover, the KAs obtained for pl7 using MAbs as ligand were similar for BIAcore and the double antibody method except for HyHIV2. Therefore, the BIAcore system can be used for the affinity measurement instead of the double antibody method.

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