Isolation and purification of viable Ureaplasma urealyticum cells free from medium components

Abstract

A procedure was devised to produce Ureaplasma uredyticum preparations free of adsorbed components of the growth medium, which contains high concentrations of serum. The ureaplasmas were cultivated in a medium containing PPLO-serum fraction as a replacement for horse serum. High titres of ureaplasmas (> lo7 c.f.u. ml-l) were obtained. Harvested cells were then purified by Urograh density gradient centrifugation. By use of 3H-labelled ureaplasma cells and 251-labelled medium components, a distinct band of viable cells devoid of serum constituents was demonstrated. The absence of medium components was verified by immunoblotting cells from this band with antiserum to medium components. Medium components that had been present before the purification procedure were undetectable in the purified cell fraction obtained. The viability of the purified ureaplasma cells represented an 85 % recovery rate and their antigenicity, examined with anti-serotype specific antiserum, remained intact. This easy and reproducible procedure can be used to prepare purified ureaplasmas for investigation of ureaplasmal antigens and their expression and/or role in disease.