The question of free bacillary growth in the plasma surrounding lepromatous explants during tissue cultivation.

An accumulated turbidity and the presence of many bacilli in the plasma surrounding lepromatous tissue culture explants has been emphasized by Timofejewsky (1) and by Suwo and Kin (2) as indicating the multiplication of free bacilli in the plasma itself. Since this inference seems unwarranted by the observations made during the preceding studies with tissue cultures, it is desired to explain why different conclusions were reached. The opacity of the plasma sw"rounding the explants is not explained by the numbers of bacilli present. In preceding communications (3, 4) , reference has been made to difficulties due to the deposition of calcium phosphates in the explants and adjacent plasma when 10 or 15 per cent of chicken embryo juice was added to human serum media. A detailed analysis of this problem is to be presented elsewhere (5). Briefly, the fresh embryo juice is saturated with respect to calcium and phosphates, and itS high phosphatase activity continues to liberate inorganic phosphates from organic phosphorus, thus producing supersaturation. The embryo juice alone, or mixtures with serum and the diluting fluid, when stored or incubated in plain glass tubes for a few days deposit _calcium phosphates which are accompanied by complex materials absorbed from the medium. In the presence of plasma, autolyzing tissue explants, and especially of actively metabolizing cultures, similar deposits accumulate most strikingly in and around the centers of the tissue cultures. Carrel's earlier notes on tissue cultivation (6) warn against the opacity which characterizes rabbit plasma in tissue c;ultivation. Rabbit serum, due to its higher calcium content, is more prone than human serum to cause calcification of plasma in the presence of embryo juice (5). Timofejewsky described his best cultures as having been prepared in rabbit and human plasma and having been renewed with undiluted human embryo juice. Suwo and Kin employed chicken plasma and renewed the liquid phase with chicken embryo juice. From what has been said, it appears correct to infer that the cloudy plasma described by Timofejewsky was due to the