目的  验证4%甲醛溶液使Sabin株Ⅰ、Ⅱ、Ⅲ型脊髓灰质炎（脊灰）病毒达到预期灭活效果所需要的时间。方法  用4%甲醛溶液对15批Sabin株Ⅰ、Ⅱ、Ⅲ型脊灰病毒在(36.5±1) ℃ 环境进行灭活，同时以15批未加入4%甲醛溶液的病毒纯化液作为对照组，按照企业注册标准采用微量细胞病变法测定病毒滴度。为了消除4%甲醛溶液对Hep-2细胞的影响，取4%甲醛溶液接种Hep-2细胞作为实验组，同时设置Hep-2细胞为对照组，每天观察对比两组细胞形态。对同一个Hep-2细胞悬液的总细胞数和活细胞数采用传统手工计数和全自动细胞计数仪计数，并比较分析结果。结果  实验组随着灭活时间的延长病毒滴度呈下降趋势，在灭活到第4天起，3个型别样品均检测不到病毒滴度；对照组病毒滴度在每毫升9.1～10.6 lg半数细胞培养物感染量范围内，符合企业注册标准的要求。4%甲醛溶液对Hep-2细胞生长有抑制作用，在培养观察到第3天时细胞全部失去活力死亡；对照组细胞形态良好呈多边形长成致密单层，活细胞组t值为18.098，总细胞组t值为4.005，P值均＜0.05，差异有统计学意义。结论  脊灰病毒在第4天能被彻底灭活，在进行此实验时应先消除甲醛对细胞培养的影响，避免结果出现假病变现象，确保结果的准确性；应用全自动细胞计数能提高计数结果的精确度。

 Objective  To verify the time required for 4%formaldehyde solution to achieve the expected inactivation effect on Sabin strain Ⅰ, Ⅱ and Ⅲ poliovirus. Methods fifteen batches of Sabin poliovirus type I, type Ⅱ and type Ⅲ were inactivated with 4%formaldehyde solution at (36.5 ± 1.0) ℃, and 15 batches of purified virus without 4%formaldehyde solution were used as control group. In order to eliminate the effect of 4%formaldehyde solution on Hep-2 cells, Hep-2 cells were inoculated with 4%formaldehyde solution as the experimental group, and Hep-2 cells were set as the control group. At the same time, SPSS 23 independent sample t test was used to compare and analyze the results of traditional manual counting and automatic cell counting. Results  In the experimental group, the virus titer showed a downward trend with the prolongation of inactivation time, and no virus titer was detected in the three types of samples since the 4th day of inactivation. In the control group, the virus titer was in the range of 9.1-10.6 lg 50% cell culture infectious dose / ml. All met the requirements of enterprise registration standard. The growth of Hep-2 cells was inhibited by 4%formaldehyde solution. On the third day of culture, all the cells in experimental group contracted, lost their vitality and died. In the control group, the cells were in good shape and grew into a dense monolayer. The t value of living cells group was 18.098, and that of total cells group was 4.005, all P < 0.05. Conclusions  Poliovirus can be completely inactivated on the fourth day. In this experiment, the effect of formaldehyde on cell culture should be eliminated to avoid false pathological changes and ensure the accuracy of the results. The application of automatic cell counting can improve the accuracy of the counting results.