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目的验证和评价医用胶原修复膜制备工艺中过氧化氢溶液、低p H孵放和γ射线辐照灭活/去除病毒的可行性。方法以水疱性口炎病毒(vesicular stomatitis virus,VSV)、伪狂犬病病毒(pseudorabies virus,PRV)、呼肠弧病毒Ⅲ(respiratory enteric orphan virusⅢ,Reo 3)、猪细小病毒(porcine parvovirus,PPV)作为指示病毒,采用非洲绿猴肾细胞(Vero)和猪肾细胞(PK-15)进行病毒增殖和感染性评价,Karber法测定病毒滴度;分别对牛腱片样品进行3%过氧化氢、低p H孵放处理(pH 3.4±0.3),检测病毒灭活/去除效果;将医用胶原修复膜中间品进行~(60)Coγ射线辐照(剂量为25 k Gy),检测病毒灭活/去除效果。结果经3%过氧化氢溶液处理1 h后,样品VSV和PRV的病毒灭活值均大于4 Logs;经低p H孵放处理14 d后,样品中已检测不到VSV和PRV,病毒灭活值分别为6.55和5.59 Logs;经25 k Gy剂量~(60)Coγ射线辐照处理后,样品中VSV、PRV、Reo3和PPV均被有效灭活,病毒灭活值分别为5.64、4.35、4.87和5.36 Logs。结论医用胶原修复膜制备工艺中的3种灭活/去除病毒步骤均有效,经灭活/去除病毒后的产品质量安全可靠。
Objective To verify and evaluate the feasibility of hydrogen peroxide solution in medical collagen repair membrane preparation process, low p H incubation and γ-ray irradiation inactivation / removal of virus. Methods Vesicular stomatitis virus (VSV), pseudorabies virus (PRV), respiratory enteric orphan virus Ⅲ (Reo 3) and porcine parvovirus (PPV) The viruses were used to detect the virus. Vero and porcine kidney cells (PK-15) were used to evaluate the virus multiplication and infectivity. Karber assay was used to measure the virus titer. Samples of tendonoid were treated with 3% hydrogen peroxide p H incubation (pH 3.4 ± 0.3), and the inactivation / removal efficiency of the virus was tested. The medical collagen repair membrane intermediate was irradiated with ~ (60) Co γ-ray (dose of 25 k Gy) effect. Results The virus inactivation values of VSV and PRV were both higher than 4 Logs after treated with 3% hydrogen peroxide solution for 1 h. VSV and PRV were not detected in the samples after 14 days incubation with low p H, The live values were 6.55 and 5.59 Logs, respectively. VSV, PRV, Reo3 and PPV in the samples were effectively inactivated after irradiation with 25 Gy dose of ~ (60) Coγ-rays, the inactivated virus values were 5.64 and 4.35, 4.87 and 5.36 Logs. Conclusion The three steps of inactivation / removal of virus in the preparation of medical collagen repair membrane are effective. The quality of products after inactivation / removal of virus is safe and reliable.