Flow cytometric assay for cytotoxic activity of crude Clostridium perfringens enterotoxin using non-adherent cell FM3A.
The flow cytometric assay method was tested for the cytotoxic activity of Clostridium perfringens enterotoxin (CPE) in culture using mouse mammary carcinoma cell line FM3A stained with propidium iodide (PI). From the results obtained, FM3A cells proved to be susceptible to CPE. A reproducible dose-response curve with FM3A was obtained between crude CPE at 13.9-109 ng/ml and between purified CPE at 40-400 ng/ml, respectively. These findings indicate that non-adherent FM3A is preferable to determine the cytotoxic activity of CPE because it can be used without detachment procedures with trypsinin compared with adherent African monkey kidney cell line (Vero cells). Furthermore, the flow cytometry with non-adherent cell FM3A stained with PI only proved to be a useful method to determine the biological activity of CPE in culture isolates.
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Laboratory of Veterinary Public Health, Department of Veterinary Medicine, College of Agriculture, Osaka Prefecture University, Sakai, Japan.
Sugii
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Kusunoki
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Uemura
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D000818:Animals; D002522:Chlorocebus aethiops; D003016:Clostridium perfringens / Q000472:pathogenicity*; D004306:Dose-Response Relationship, Immunologic; D004768:Enterotoxins / Q000633:toxicity*; D005434:Flow Cytometry / Q000379:methods*; D011419:Propidium; D014407:Tumor Cells, Cultured; D014709:Vero Cells
D004768:Enterotoxins; C035598:enterotoxin, Clostridium; D011419:Propidium