霍乱弧菌核酸检测试剂国家参考品的建立

目的：

建立霍乱弧菌核酸检测试剂国家参考品并制定质量标准。

方法：

收集并筛选霍乱弧菌核酸阳性和阴性细菌培养物样本，建立霍乱弧菌核酸检测试剂国家参考品，并对参考品稳定性和均匀性进行评价；经 7 个实验室的协作标定，依据标定结果制定国家参考品质量标准。

结果：

霍乱弧菌核酸检测试剂国家参考品包括阳性 8 份、阴性 10 份、重复性 2 份和检测限 2 份。参考品均匀性符合要求。冻融 1 次、2–8 ℃及室温（25 ℃）环境中放置 7 天均不影响参考品的稳定性。国家参考品的质量标准为阳性符合率 8/8，阴性符合率 10/10，重复两个浓度水平 Ct 值的变异系数均不高于 5.0 %，L1 和 L2 要求 1:10⁴ 稀释或更高浓度时应均为霍乱弧菌阳性。检测限和重复性参考品 L1、L2、R1 和 R2 分别赋值为 3.9×10⁷ U/mL、2.4×10⁷ U/mL、7.3×10⁷ U/mL 和 6.3×10⁷ U/mL。

结论：

建立了霍乱弧菌核酸检测试剂国家参考品，可用于相关试剂的质量控制及评价。

Objective: 

To develop a national reference panel for Vibrio cholerae nucleic acid tests and establish the relative quality requirements.

Methods: 

The bacterium culture samples positive and negative for Vibrio cholerae nucleic acid were collected and screened to establish a national reference panel for Vibrio cholerae nucleic acid tests. The stability and homogeneity of the reference panel were evaluated, and the quality criteria were determined via collaborative calibration conducted by seven laboratories.

Results: 

The national reference panel consists of 8 positive, 10 negative, 2 precision and 2 lower detection limit samples. The homogeneity of the reference panel meets the requirements, and the stability of the reference panel was not affected by freezing and thawing once, storing for 7 days at 2–8 ℃ and room temperature (25 ℃). The quality requirements of the national reference panel are that the positive coincidence rate is 8/8, and the negative coincidence rate is 10/10. For precision, the coefficient of variation (CV) of the Ct value at the two concentration levels is not higher than 5.0 %. For the limit of minimum detection, a dilution ratio of 1∶10⁴ or higher concentrations on L1 and L2 should be positive for nucleic acid of Vibrio cholerae. Precision and lower detection limit samples L1, L2, R1 and R2 are assigned as 9×10⁷ U/mL, 2.4×10⁷ U/mL, 7.3×10⁷ U/mL, and 6.3×10⁷ U/mL.

Conclusion: 

A national reference panel for Vibrio cholerae nucleic acid tests has been established. This panel can be used for quality control and evaluation of related diagnostics.