Characterization of monoclonal antibodies to trichocyst antigens in Paramecium

Abstract

Ten mouse monoclonal antibodies (mAbs) were raised against trichocyst contaminants present in crude or enriched lysosome fractions of Paramecium multimicronucleatum. Using an indirect immunofluorescence assay (IFA) and immunogold labelling on frozen thin sections, epitopes were located on the outer edge, cortex and the core of the trichocyst body, as well as the sheath covering the tip. Except for the two on the tip, epitopes were reactive after SDS-PAGE under non-reducing conditions. Four mAbs (131C1E8, A1-3, A16-2, D7) were directed to a trio of bands of 37, 34 and 29 (x 10(3] Mr from the beaded or meshlike trichocyst body sheath. A fifth mAb (135B9E7), directed to epitopes on the cortex inside the beaded body sheath, reacted strongly with the 37 and 34 bands, but weakly with the 29 × 10(3) Mr band. The last three mAbs (270D5, 22D7F2, D8) were reactive with one or more of three families of antigens found on the trichocyst core. mAb 270D5 reacted mainly with the 34 and 29 (x 10(3] Mr bands of the family containing the above trio, while mAb 22C7F2 reacted consistently with the 47 × 10(3) band of the higher Mr family but variably with both the trio of bands and the 17 × 10(3) band of the lower Mr family. mAb D8, which was directed to epitopes on the trichocyst core and small vesicles in the endoplasm, reacted only with the 29 × 10(3) Mr band. The mAbs were cross-reactive with the trichocysts of P. primaurelia, P. tetraurelia, P. caudatum and P. calkinsi with some small variation in blotting patterns.(ABSTRACT TRUNCATED AT 250 WORDS)