US EPA

5214994 

Journal Article 

Purification of Borrelia burgdorferi outer surface protein A (OspA) and analysis of antibody binding domains 

Jiang, W; Gorevic, PD; Dattwyler, RJ; Dunn, JJ; Luft, BJ 

1994 

Yes 

Clinical and Diagnostic Laboratory Immunology
ISSN: 1071-412X
EISSN: 1098-6588 

1 

4 

406-412 

English 

8556477 

WOS:A1994PT56500008 

The major outer surface protein, OspA, of Borrelia burgdorferi is a lipoprotein which is a particular interest because of its potential as a vaccine candidate. However, serotypic and genetic analysis of OspA from both European and North American strains have demonstrated antigenic and structural heterogeneities. We purified OspA to homogeneity by exploiting its resistance to trypsin digestion. By treating spirochetes with trypsin and then using Triton X-114 extraction and ion-exchange chromatography, we obtained a yield of 2 mg of pure OspA protein per liter of culture. INtrinsic labeling with [14C]palmitic acid confirmed that OspA was lipidated, and partial digestion established lipidation at the amino-terminal end of the molecule. The reactivity of five anti-OspA murine monoclonal antibodies to nine different isolates of B. burgdorferi was ascertained by Western blot (immunoblot) analysis. Purified OspA was fragmented by enzymatic or chemical cleavage, and the monoclonal antibodies were able to define four distinct immunogenic domains. Further resolution of the epitope specificity to determine humoral and cellular immune responses to OspA has implications for vaccine development and for the utility of this protein as a reagent in diagnostic testing for Lyme borreliosis.