EPITOPE LOCATION IN THE CRYPTOCOCCUS-NEOFORMANS CAPSULE IS A DETERMINANT OF ANTIBODY EFFICACY

Monoclonal antibodies (mAbs) to the polysaccharide capsule of Cryptoco ccus neoformans can prolong survival in mice. However, the properties of antibodies that mediate protection are not fully understood. The Ig M mAbs 12A1 and 13F1 originated from the same B cell and differ only b y somatic mutations in their variable regions; yet mAb 12A1 protects a gainst serotype D infection, while mAb 13F1 does not. Phage peptide di splay libraries were used to analyze the fine specificity of these two mAbs. The selection of distinct peptide motifs from identical librari es confirmed that mAbs 12A1 and 13F1 bound to two distinct epitopes. I mmunofluorescence and immunoelectron microscopy studies revealed diffe rences in antibody localization within the capsule of serotype D strai n; mAb 12A1 bound to the outer rim of the capsule resulting in an annu lar pattern, whereas mAb 13F1 bound throughout the capsule and had a p unctate appearance. The difference in the binding pattern of mAb 12A1 and 13F1 was not observed on serotype A organisms, where both mAbs bou nd to the capsule with an annular fluorescence pattern. The fluorescen ce pattern of binding correlated with protective efficacy; mAb 13F1 pr olonged survival of mice infected with the J11 serotype A strain (annu lar fluorescence), but not serotype D strains (punctate pattern). Annu lar binding, but not punctate binding, was associated with increased o psonic efficacy for phagocytosis of C. neoformans by J774.16 macrophag e-like cells. The correlation between capsular binding pattern, opsoni c activity, and ability to prolong survival suggests that the efficacy of anticryptococcal antibodies is dependent upon where they bind in t he polysaccharide capsule.