目的 探討凝膠法測定人臍帶間充質(zhì)干細胞培養(yǎng)上清中細菌內(nèi)毒素的可行性。方法 收集人臍帶間充質(zhì)干細胞培養(yǎng)72 h的上清為供試品。參照《中國藥典》2020年版第三部通則1143細菌內(nèi)毒素檢查法中凝膠法的具體要求和步驟對鱟試劑進行靈敏度復核，對供試品進行干擾初篩試驗、干擾試驗和供試品細菌內(nèi)毒素的測定。結果 鱟試劑靈敏度測定值為0.125 EU/mL，在0.5λ～2.0λ符合規(guī)定，可用于后續(xù)試驗。供試品1、2、4倍稀釋對鱟試劑均無干擾；干擾試驗驗證2倍稀釋供試品對細菌內(nèi)毒素凝膠法檢測無干擾作用；采用凝膠法測定2倍稀釋供試品的細菌內(nèi)毒素含量均≤0.5 EU/mL，判定供試品中細菌內(nèi)毒素含量合格。結論 凝膠法可用于人臍帶間充質(zhì)干細胞培養(yǎng)上清中細菌內(nèi)毒素的檢測。

Objective To investigate the feasibility of sol-gel method to detect bacterial endotoxin in supernatant of human umbilical cord mesenchymal stem cells (hUC-MSCs). Methods The supernatants of hUC-MSCs cultured for 72 h were collected as the test substance. According to incoterms 1143 on sol-gel method of bacterial endotoxin test (BET) method in Chinese Pharmacopoeia Ver. 2020, sensitivity tests of tachypleus amebocyte lysate (TAL) was studied, then interference initial screening test, interference verification test, and the endotoxin content in samples were determined by sol-gel method. Results The TAL with the sensitivity of 0.125 EU/mL coincided with the regulation in the range of 0.5λ-2.0λ, and could be used in the tests. The samples diluted 1, 2, and 4 times, interference initial screening test weren’t observed the effect of inhibition. The interference verification test with 2 times dilution of samples had no impact on TAL. Sol-gel method was used to determine the endotoxin, and indicated the TAL in samples with 2 times dilution was less than 0.5 EU/mL qualified to regulation. Conclusion It is suitable for bacterial endotoxin in supernatant of hUC-MSCs by sol-gel method.

R286.02

云南省衛(wèi)生健康委員會內(nèi)設機構資助項目（2017NS228，2018NS261）；云南省臨床病毒學重點實驗室項目（202205AG070053）；云南省衛(wèi)生科技計劃項目（L-2019003）；云南省血液疾病臨床醫(yī)學中心開放課題（2020LCZXKF-XY04）