目的验证无酚红M199培养基的贮存效期，同时探索无酚红M199效期对无酚红Sabin株脊髓灰质炎灭活疫苗（inactivated Sabin strain poliovirus vaccine，sIPV）关键质量属性的影响。
方法对贮存0、6、12、18、24、30、36及42个月的3批次无酚红M199干粉培养基进行外观、澄清度、pH、干燥失重、细菌内毒素、微生物限度等物理化学性质及微生物性质长期监测；检测用贮存36个月并检定合格的无酚红M199制备的sIPV成品关键质量属性；将近效期和远效期无酚红M199制备的各3批sIPV分别贮存于37 ℃和25 ℃进行稳定性监测，检测D抗原含量，评价其稳定性；用2组sIPV免疫大鼠，检测大鼠血清抗体效价，评价其免疫原性。
结果无酚红M199干粉培养基于2~8 ℃贮存42个月后的外观、澄清度、pH、干燥失重、细菌内毒素、微生物限度等均合格；用贮存36个月无酚红M199所制备sIPV成品的pH、D抗原、蛋白含量、Vero细胞DNA残留量、牛血清白蛋白残留量、Vero细胞蛋白残留量、游离甲醛含量等各项检定结果均符合中国药典2020年版三部和企业标准要求；用贮存36个月无酚红M199制备的sIPV效价合格、稳定性良好且与远效期无酚红sIPV免疫原性趋势一致，效力（ t =-0.15~2.17， P＞0.05）和诱导中和抗体水平（ t =-1.46~2.02， P＞0.05）差异均无统计学意义。
结论用贮存至36个月的无酚红M199制备的sIPV成品稳定性较好，近效期和远效期无酚红M199制备的sIPV产品质量属性、免疫原性及稳定性趋势一致，且对产品质量无影响，无酚红培养基的效期初步可定为36个月。
ObjectiveTo validate the storage potency of phenol red-free M199 medium and explore the effect of phenol red-free M199 potency on the key quality attributes of phenol red-free Sabin strain inactivated poliovirus vaccine (sIPV).
MethodsThe appearance, clarity, pH, weight loss on drying, bacterial endotoxin, microbial limit and other physical and chemical properties of 3 batches of phenol red-free M199 powder culture medium stored for 0, 6, 12, 18, 24, 30, 36 and 42 months were monitored. Key quality attributes of final sIPV products prepared with phenol red-free M199 calibration qualified after 36 months of storage were detected. The sIPVs prepared with near- and far-period phenol red-free M199 were stored at 37 ℃ and 25 ℃, respectively, for stability monitoring and evaluation by detecting the D antigen content.The 2 groups of sIPVs were used to immunize rats, and the immunogenicity was evaluated by detecting the antibody potency of rat serum.
ResultsThe appearance, clarity, pH, loss on drying, bacterial endotoxin, microbial limit of phenol red-free M199 dry powder culture medium stored at 2~8 ℃ for 42 months were all qualified. The pH, D antigen content, protein content, Vero cell DNA residue, bovine serum albumin residue, Vero cell protein residue, free formaldehyde content and other test results of sIPV produced using phenol red-free M199 stored for 36 months were in line with the requirements of the Chinese pharmacopoeia 2020 edition volume Ⅲ and the enterprise standards. The sIPV prepared with phenol red-free M199 stored up to 36 months was potent with good stability, and the trend of immunogenicity was consistent with that of the sIPV prepared with far-period phenol red-free M199, with no statistically significant differences in potency（ t =-0.15-2.17, P＞0.05）or neutralizing antibody level（ t =-1.46-2.02, P＞0.05）.
ConclusionsThe sIPV final product prepared with phenol red-free M199 stored up to 36 months has good stability. The product quality attributes, immunogenicity and stability of sIPV products prepared with near- and far-period M199 show the same trend, and near-period M199 has no effect on the product quality. Phenol red-free media can be valid for 36 months.
刘英微,刘哲宇,田东阳,等. 用于Sabin株脊髓灰质炎灭活疫苗生产的无酚红M199培养基贮存效期验证[J]. 国际生物制品学杂志,2025,48(01)：20-25.
DOI:10.3760/cma.j.cn311962-20240223-00009版权归中华医学会所有。
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刘英微：实验设计、研究实施、数据采集、数据分析、统计分析、论文撰写；刘哲宇、李建新：研究实施、数据采集、数据分析、统计分析、技术支持、研究指导；田东阳：研究实施、数据采集、数据分析、技术支持、研究指导；张萌：研究实施、数据采集、数据分析；李娜：实验设计、研究实施、统计分析、论文撰写、论文修改、支持性贡献
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