SARS-CoV细胞空斑试验

目的建立适用于生物安全三级实验室操作的SARS．CoV空斑试验方法。方法SARS－CoVl0倍系列稀释接种Vero－E6细胞，用0．6％琼脂糖覆盖中性红着色（琼脂糖．中性红法）或覆盖1％甲基纤维素，4％甲醛固定，结晶紫染色（甲基纤维素－结晶紫法），空斑计数。结果琼脂糖－中性红法病毒滴度为6．14Lg pfu／mL，甲基纤维素－结晶紫法病毒滴度为6．57Lg pfu／mL。结论二种空斑试验方法相比，病毒滴度无明显差异。甲基纤维素－结晶紫空斑试验法形成的空斑比琼脂糖－中性红法清晰、操作简单安全、结果可长期保存。

Plaque Assay in SARS-CoV Titration

Objective To optimize and assess the method of plaque assay of severe acute respiratory syndrome （SARS）associated eoronavirus （SARS-CoV） for pathogenesis studies and monitor the viral titers for quantifying the amounts of SARS-CoV. Method A series of 10-fold dilutions of viruses were added into Vero-E6 cells seeded plates. The plates were overlaid with DMEM containing 1% methyleellulose 4,000 CP. The Vero-E6 cells were fixed with 4% formaldehyde, and stained with 0.5 % crystal violet. Plaques were counted after the plates were air dried. Result The viral titer was 6.57 Lg pfu/mL. Clear plaques were formed in wells infected with SARS-CoV. Conclusion This procedure is useful for SARS-CoV titration and has less risk of SARS-CoV handling hazards in bio-safety laboratory. The plates can be kept for a longer time than other assays. This improved plaque formation technique can also be used and applied to measure the neutralizing antibody activities against SARS-CoV infection.