The cytopathic effect of staphylococcal enterotoxin B upon mammalian cell cultures is expressed fully only if the cultures are grown in human serum-supplemented media. Little cytotoxicity is seen if the cells are grown in horse serum-supplemented media and none is seen if the cells are grown in calf serum-supplemented media. The cytotoxicity of the toxin is reversible at any stage of intoxication. Cytotoxicity is rarely seen before 5 hr and is most often not evident before 8 hr. Cytoxicity is

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... t lethal. A number of reports have been published in recent years concerning the interaction of staphylococcal enterotoxin B with a variety of cell culture lines and strains. The authors of these reports have been in disagreement as to whether cell culture systems are sensitive or resistant to the cytotoxic properties of enterotoxin B. One of the earliest reports in which a highly purified toxin preparation was used, that of Guerin et al. (Abstr. Annu. Meeting Tissue Culture Ass., 12th, p. 37, 1961) showed that chick embryonic fibroblasts are sensitive to graded concentrations of enterotoxin. Milone (3), by using HEp-2, HeLa, and human heart cell cultures, showed that the cytotoxic response of these cells diminished as more highly purified toxin preparations were employed. Further, and more significantly, the cytotoxicity of none of the preparations could be neutralized with specific antitoxin. Grigorova and Danon (1), with a variety of primary and established cell cultures, showed a cytotoxic response of the cells to enterotoxin. However, no purification step of the toxin was reported, nor was there any report of the specificity of the cytotoxic effect of their culture filtrates. Hallander and Bengtsson (2), with a highly purified preparation of enterotoxin B, reported no cytotoxicity after 2 hr of exposure in the cell cultures they employed. In two earlier reports (4, 5), we have shown that a specific cytotoxic response to enterotoxin B by human embryonic intestine cell cultures can be achieved and, further, that this response may be altered if the cells are trypsinized before challenge. We have continued our examination of the factors influencing the interaction of enterotoxin B with our cell culture system and have recently tested our methodology in five additional cell culture systems. The purpose of this report is to demonstrate how it is possible to see sensitivity and resistance in one cell line, depending upon the conditions one uses to grow the cultures or challenge the cells. MATERIALS AND METHODS Cell cultures. Six heteroploid cell lines were used in this study: human embryonic intestine, HeLa, HEp-2, Chang liver, KB, and L-cells. In addition, one primary cell culture of human foreskin fibroblasts was employed. Toxin preparation. The toxin preparation used was the same as described previously (4) . The toxin preparation is between 96 and 99% pure. The usual tests for a and # hemolysins, apyrase, and dermonecrotic substances were negative. Cell didture maintenance and experimental procedures. The stock cultures were grown in Eagle's basal medium containing Earle's salt solution and were incubated in a 5% CO2 atmosphere. The serum supplement used initially was 10% human serum for the human embryonic intestine cell line or 10% fetal calf serum for the other cell cultures. All cell culture supplies were purchased from Grand Island Biological Co., Grand Island, N.Y. To adapt the cell cultures to grow in media supplemented with human serum or calf serum, two batches of Eagle's basal medium were prepared, one containing 10% fetal calf serum and one containing 10% human serum. Various mixtures of the two were made (9:1, 8:2, 7:3, etc.) as required and the cell cultures were either successively fed or planted with mixtures of the two. Cultures were retained at each successive 455 on May 7, 2020 by guest http://iai.asm.org/ Downloaded from