Detection of serum proteins in the electrophoretic patterns of total proteins of mycoplasma cells

SummaryThe contamination of mycoplasma cell preparations by serum proteins originating from culture medium was studied.A. laidlawiiandM. arthritidiscells were grown in the presence of [14C]-aminoacids, and the cells were washed with 0·9% NaCl by threefold centrifugation. Total proteins of the washed cells were analysed by SDS gel electrophoresis. Coomassie-stained electrophoretic patterns were compared with autoradiographs of the same gels. The stained electrophoretic pattern of washedA. laidlawiigrown without serum was identical with autoradiographs of the same cells grown without or with serum. That of washedA. laidlawiigrown with serum differed from the corresponding autoradiograph by the presence of extra protein bands I, II, III, and IV with molecular weights of over 160,000, 80,000–87,000, 55,000 and 25,000, respectively. The same extra bands were found in stained electrophoretic patterns of washed: (a)A. laidlawiicells grown without serum and mixed with serum in the stationary phase, (b)M. arthritidiscells, as compared with their autoradiographs, (c) serum precipitate. The bands III and IV may be due to the heavy and light chains of γ-globulin, the band II might belong to transferrin or to some component of complement. Acidification of serum to pH 5 brought about 100-fold rise of amount of serum precipitate, the number of bands in the electrophoretic pattern of the precipitate being also increased. Stained electrophoretic patterns of cells purified by twofold centrifugation in step sucrose density gradient (1·20–1·27 g./cm.3forA. laidlawii,and 1·15–1·25 forM. arthritidis) contained no extra bands and matched completely with their autoradiographs.It was concluded that contamination of washed mycoplasma cells by serum proteins is mainly due to co-precipitation of aggregated serum proteins together with cells during centrifugation rather than to adsorption of serum proteins on the cell surface.

Factors in the interpretation of the inhibitive and fixation serum reactions in pulmonary tuberculosis

Calmette and Massoll have reported that immunisation of a calf with "bacillen-emulsion" has produced, not complemetn-fixation bodies, but an inhibitive effect which they designate as an "inhibiteur." Previous to this they had reported upon a serum, obtained by intravenous inoculation, which had an extremely active agglutinating property, and under certain conditions was capable of precipitating various solutions of tuberculin. The reaction of fixation was not, however, obtained with either the serum or the serum precipitate. Independently I had observed a somewhat analogous serum reaction (inhibitive reaction) in certain cases of pulmonary tuberculosis, and I first reported this at the meeting of the Canadian Medical Association, Toronto, June, 1910, subsequently publishing more extended investigations along these lines. These reactions have become an additional aid in the clinical estimation of the type of case. Emery records an observation which he regards as similar in character to that of Clamette and Massol. The variations in technique at present, however, prevent one from drawing any very close comparisons.