ABSTRACT
Hepatitis B surface antigen (HBsAg) specific monoclonal antibodies (mAbs) are potentially valuable therapeutic and diagnostic tool. We have previously established and characterized a panel of mAbs derived from immunized BALB/c mice with a yeast-derived recombinant HB vaccine subgentoype A2 and HBsAg subtype adw2. This study was conducted to evaluate the reactivity pattern of this anti-HBs mAbs panel with various genotypes and subgenotypes of HBV using the first WHO HBV genotype reference panel containing 15 serum samples representing the subgenotypes A1, A2, B1, B2, C2, D1-D3, E, F2, and H. Ten out of 21 anti-HBs mAbs were able to strongly recognize all gentopye/subtypes of HBsAg provided in the WHO reference panel. However, 10 out of 21 anti-HBs mAbs showed a moderate to profound loss of reactivity with HBV genotypes/HBsAg subtypes D2/ayw3, E/ayw4, F2/adw4, and H/adw4. Two mAbs from the second group displayed a profoundly reduced reactivity with only 1 out of 3 C2/adr genotype/subtype samples. The amino acid alignment of these 3 samples showed that this particular sample contains amino acid substitution at residue 127, which is located inside "a" determinant. This amino acid substitution, which profoundly affected the reactivity of anti-HBs antibodies, has been previously reported only in D/ayw3, E/ayw4, F/adw4, and H. Interestingly, the amino acid alignment of the samples in this WHO panel showed that P127T substitution can also be found in C2/adr. Comparing amino acids sequences inside the antigenic loop (AGL) showed that D2/ayw3 contains a T118A/P127T double substitution, E/ayw4 contains P127L/T140S, F2/adw4 contains P127L/T140S/ F158L, and H/adw4 contains P127L substitution. Therefore, amino acid variability at positions 118, 127, 140, and 158 was found to cause significant loss of reactivity with anti-HBs mAbs. Since HBsAg variability in different genotypes of HBV can profoundly affect the reactivity of anti-HBs mAbs, analytical sensitivity for HBsAg assays should be considered based on the circulating and common HBV variants in the relevant countries.
