荧光定量PCR仪光学校准方法与结果分析

Optical Calibration and Result Analysis of Real-time Fluorescence Quantitative PCR Instrument

实时荧光定量PCR仪特异性更强,自动化程度更高,且有效地解决了PCR污染的问题,应用领域及应用量都不断增加。但其设计更为复杂,温度模块和光学系统设计同时影响其性能和实验准确性,为定量PCR仪校准带来了巨大挑战。采用生物试剂等方式对定量PCR仪荧光部分校准缺乏溯源性,无法分析误差来源,存在较大缺陷。本文在对定量PCR仪影响因素和校准现状分析的基础上,采用Cyclertest 3D optical定量PCR仪光学校准系统对ABI 7500 Fast Real-Time定量PCR仪的温场部分和荧光系统进行了检测并对检测结果进行了分析,结果表明对温度模块和光学系统共同进行检测并分析相关性能够更科学全面地评估定量PCR仪性能,满足定量PCR仪校准需求。

The real-time fluorescence quantitative PCR instrument has stronger specificity,higher degree of automation,and effectively solves the problem of PCR contamination,the application field and application are increasing. However,its design is more complex. The design of temperature module and optical system simultaneously affects its performance and experimental accuracy,which brings great challenges to the calibration of quantitative PCR instrument. The fluorescence partial calibration of quantitative PCR instrument by means of biological reagents lacks traceability and cannot analyze the source of error. Based on quantitative PCR instrument calibration based on the analysis of the status quo,and the factors affecting the Cyclertest quantitative PCR optical calibration system for3 D optical ABI 7500 Fast Real-Time fluorescent quantitative PCR temperature field of the part and the system was tested and the test results are analyzed,the results show that the temperature of the module and the optical system for testing and analysis of correlation can be more scientific and comprehensive evaluation of quantitative PCR instrument performance,meet the demand of quantitative PCR instrument calibration.