Production, characterization, and application of monoclonal antibodies which distinguish three glucosyltransferases...

Thirty-three murine monoclonal antibodies (MAbs) against the three glucosyltransferases (GTFs) (GTF-I, -SI, and -S) from Streptococcus mutans were obtained by the fusion of murine myeloma cells (P3X63-Ag8-U1) with spleen cells of BALB/c mice immunized with pure GTF-S or partially purified GTF-I from serotype c S. mutans PS14. The immunoreactivities of these MAbs were tested by enzyme-linked immunosorbent assay and Western blotting (immunoblotting) with various GTF preparations. GTF-I and GTF-SI were expressed from two Streptococcus milleri or Escherichia coli transformants harboring gtfB or gtfC, respectively. All of the five MAbs raised against the GTF-S from PS14 reacted only with the homologous enzymes. Of these, 8 MAbs reacted only with the gtfB gene product (GTF-I), 4 MAbs reacted only with the gtfC gene product (GTF-SI), and the remaining 16 MAbs reacted with both gene products. The existence of GTF-SI in the purified GTF-I from PS14 was demonstrated by Western blot analysis using the representative monospecific MAbs. Further, the relative levels of the three GTFs in the extracellular and cellular fractions of S. mutans clinical isolates were examined by immunoblot analysis. The findings indicated that the relative level of GTF-SI, unlike that of GTF-I or GTF-S, differed markedly among isolates although the three GTFs were synthesized extracellularly by all the strains.