基于全自動管式化學發光免疫檢測系統的人結核感染T細胞檢測方法的建立

陳鷺穎　林海軍　翁祖星　徐飛海　葛勝祥　張　軍★

基于全自動管式化學發光免疫檢測系統的人結核感染T細胞檢測方法的建立

陳鷺穎1林海軍2翁祖星2徐飛海2葛勝祥2張軍2★

［摘要］目的建立基于化學發光平臺的人結核感染T細胞檢測方法。方法將一株γ－干擾素單抗標記在磁微粒上，另一株γ－干擾素單抗標記在吖啶酯上，然后將檢測體系與已有的細胞刺激培養體系相結合。結果本研究成功建立基于化學發光平臺的人結核感染T細胞檢測方法。以ELISA平臺檢測試劑的檢測結果作為參考，該方法的靈敏度為98．3%，特異性為99．2%，總體符合率達到98．8%。結論該方法具有更高的分析靈敏度（可達0．27 pg／mL）、更寬的線性范圍（1 pg／mL～5 000 pg／mL）、更好的重復性（批內與批間變異系數均＜6．0%）及更易實現高通量檢測，為臨床診斷結核感染提供了有力的工具。

［關鍵詞］結核感染T細胞；γ-干擾素定量檢測；化學發光

作者單位：1．福建省食品藥品認證審評中心，福建，福州350003 2．廈門大學國家傳染病診斷試劑與疫苗工程技術研究中心，福建，廈門361102

Development of detection kit for T cell infected with Mycobacterium tuberculosis based on fullautomatic chemiluminescence immune analyzer

CHEN Luying1, LIN Haijun2, WENG Zuxing2, XU Feihai2, GE Shengxiang2, ZHANG Jun2★
(1. Fujian Food and Drug Administration, Fuzhou, Fujian, China, 350003; 2. National Institute of Diagnostics and Vaccine Development in Infectious Diseases, Xiamen University, Xiamen, Fujian, China, 361012)

[ABSTRACT] ObjectiveTo develop the detection kit for T cell infected with Mycobacterium tuberculosis based on full-automatic chemiluminescence immune analyzer. Mehtods An antiIFN-γ Mab was coated on the surface of microparticle. Another anti IFN-γ Mab was labelled to acridinium ester. After that, the detection system was combined with the in vitro cell culture system.ResultsThis research developed the detection kit for T cell infected with Mycobacterium tuberculosis based on full-automatic chemiluminescence immune analyzer. Compared with the testing results of the ELISA kit, the sensitivity, specificity and total matching ratio of the CLIA kit was 98.3%, 99.2% and 98.8%, respectively. ConclusionThe CLIA kit has better sensitivity (0.27 pg/mL), wilder linear range (1 pg/mL ～ 5 000 pg/mL), and better repeatability (intra and inter coefficient of variation < 6.0%). It makes a high throughput detection available. It will contribute to the clinical diagnosis of Mycobacterium tuberculosis.

[KEY WORDS] T cell infected with Mycobacterium tuberculosis; IFN-γ quantitative detection; Chemiluminescent immunoassay

目前全球約有三分之一的人感染了結核分枝桿菌，其中約10%會進一步發展成為活動性結核［1］。臨床上用于判斷結核分枝桿菌感染的最常見方法是結核菌素皮膚試驗（tuberculin skin test，TST）。但TST的特異性較差，陽性結果不能排除非結核分枝桿菌（Nontuberculous mycobacteria，NTM）感染，也無法區分是否由接種卡介苗引起［1－2］。因此，近年來在歐美發達國家，TST已逐漸被γ-干擾素釋放試驗（interferon gamma release assay，IGRA）所取代。IGRA的方法是利用結核分枝桿菌特異性抗原與受試者的新鮮外周全血中的T細胞共同孵育刺激培養，如果受試者受到過結核分枝桿菌感染，那么激活的T細胞會分泌大量的γ-干擾素，通過γ-干擾素的定量檢測可以判斷受試者是否存在結核菌感染?！?br>