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中国防痨杂志 ›› 2014, Vol. 36 ›› Issue (5): 380-384.doi: 10.3969/j.issn.1000-6621.2014.05.016

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结核分枝杆菌感染T细胞斑点试验的质量控制与质量保证

蒋俊   

  1. 110044  沈阳市胸科医院检验科结核病实验室 沈阳市结核病重点实验室
  • 收稿日期:2013-04-21 出版日期:2014-05-10 发布日期:2014-06-07
  • 通信作者: 蒋俊 E-mail:xkyyjykjj@sina.com

Quality control and quality assurance of T-SPOT.TB

JIANG Jun   

  1. TB Laboratory of Shenyang Chest Hospital,Shenyang TB Key Laboratory,Shenyang 110044,China
  • Received:2013-04-21 Online:2014-05-10 Published:2014-06-07
  • Contact: JIANG Jun E-mail:xkyyjykjj@sina.com

摘要: 近几年来,结核分枝杆菌感染T细胞斑点试验(T-SPOT.TB)在国内外已较多地应用于结核分枝杆菌感染检测或相关性研究。T-SPOT.TB作为集细胞生物学和免疫学检测技术为一体的新方法,虽然操作并不复杂,但对试验条件、操作技术和质量控制要求很高,其试验体系中外周血单个核细胞(PBMCs)的分离、加入活体细胞的数量、细胞孵育条件和结果判定等的全程质量控制,既是决定试验成败的关键,又是常常易于忽略的“潜在性”问题。作者依据其试验原理和临床实践对试验中常见的PBMCs分离呈现的不同状态进行了分析和描述;对PBMCs的分离、悬液制备、体外孵育和结果判定的质量控制进行了重点论述,其中就人工PBMCs计数误差对试验结果的影响进行了详细阐述。除此,还依据传统的改良牛鲍血细胞计数板白细胞计数法和溶液浓度稀释法溶液浓度与体积成反比的基本原理,结合活体PBMCs计数和标准细胞悬液终浓度2.5×105/100μl的特殊要求,建立了简易的细胞计数“一步法”和“计算式”,使标准细胞悬液的制备趋于简化。

关键词: 结核/诊断, &, gamma, 干扰素释放试验, 白细胞, 单核, 细胞计数, 质量控制

Abstract: T-SPOT.TB has been widely used to assay the Mycobacterium tuberculosis infection all around the world. As a new cell biological and immunological detection technology which its operation isn’t complex, T-SPOT.TB also raises a claim strictly on experimental condition, operation technique and quality control. The overall quality control including PBMCs separation, adjusting the number of PBMCs, cell incubation condition and result determination is the key steps to succeed as well as to be ignored. According to the test principle and clinical practice, we described and analyzed the different states of PBMCs separation including PBMCs separation, suspension preparation, incubation and result determination, especially, the error of PBMCs count affected experimental result. In addition, we established “one-step” and “calculation formula” method for PBMCs count according to traditional improved NiuBao leukocyte count plate and solution concentration dilution method, PBMCs count and standard cell suspension final concentration of 2.5×105/100μl.

Key words: Tuberculosis/diagnosis, Interferon-gamma release tests, Leukocytes, mononuclear, Cell count, Quality control