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目的探讨流感网络实验室病毒分离各环节的关键影响因素,提高实验室工作质量与监测效率。方法按照目前湖北省流感监测实施方案,通过对近5年的相关文献综述与2013-2014年笔者所在实验室的所得数据,对其中涉及标本采集、保存运输、狗肾细胞、病毒分离等过程中的重要环节的影响因素进行分析,使用SPSS 17.0进行Distances程序分析并探索各因素的关系。结果在病毒分离过程中有以下参考:标本采集发病1~2 d内且体温达到39℃的病例标本,标本种类根据流行株的型别进行重点部位采样,新甲(H1N1)亚型流感标本采集咽拭子,B型流感标本采集鼻拭子;标本采集后48 h内无法送达实验时放置-80℃保存;细胞代数选择较低的,生长液中谷氨酰胺浓度维持在1.5%~2.0%,培养时间不超过48 h。标本48 h内接种效果较好,维持液中加入适量胰酶(约2 ug/m1),吸附时间采用37℃加1 h组合较宜,收获病毒前冻融1~2次有助于病毒分离;CT值在22~24之间的标本容易分离。结论在流感监测工作中要注意标本采集时间、部位、保存位置、细胞代数、谷氨酰胺浓度、培养时间、胰酶浓度、吸附温度与时间、CT值、冻融次数等重要环节与重要参考值,以期提高流感工作效率,保证工作质量。
Objective To explore the key factors influencing the isolation of influenza virus in laboratory and improve the quality of laboratory work and monitoring efficiency. Methods According to the current implementation plan of influenza surveillance in Hubei Province, through the literature review of the past five years and the data obtained in the laboratory of the author from 2013 to 2014, we collected the data of samples collected, preserved and transported, isolated dog kidney cells and virus Of the important factors of the impact of analysis, the use of SPSS 17.0 Distances program analysis and explore the relationship between the various factors. The results of the virus in the process of separation of the following reference: specimens collected within 1 ~ 2 d and the temperature reached 39 ℃ case specimens, specimen types according to the type of epidemic strain sampling of key parts, the new H1N1 subtype influenza virus samples collected Nasopharyngeal swabs and influenza B specimens were collected. The specimens were stored for 48 hours and then stored at -80 ° C. The cell algebra was selected to be low, and the concentration of glutamine in the growth liquid was maintained at 1.5% -2.0% , Culture time does not exceed 48 h. The inoculation effect was good within 48 h, and the amount of trypsin (about 2 ug / ml) was added to the maintenance solution. The adsorption time was preferably set at 37 ° C for 1 h. Frozen or thawed for 1 to 2 times ; Specimens with CT values between 22 and 24 were easily separated. Conclusions During the surveillance of influenza, attention should be paid to the important links such as sample collection time, location, storage location, cell algebra, glutamine concentration, culture time, trypsin concentration, adsorption temperature and time, CT value, , In order to improve the work efficiency of the flu and ensure the quality of work.