EB病毒核酸检测质控品制备及其应用

doi:10.3969/j.issn.1673-8640.2015.11.005

摘要/Abstract

摘要： 目的

制备可模拟真实临床标本的EB病毒(EBV)核酸检测质控品用于上海地区临床实验室室间质评,以评估其EBV DNA的检测能力。

方法

选择整合有EBV基因组的淋巴细胞(NAMALWA)进行培养,收集细胞后观察细胞原液对国内常用商品化试剂盒的适用性,随后用EBV国际标准品对其定值,经血浆稀释制得含有阴、阳性共5份样本的样本盘,随机编码后发送至参加EBV DNA室间质评的临床实验室,并对回报结果进行分析。

结果

采用不同公司EBV PCR检测试剂盒检测NAMALWA细胞原液,结果全部阳性。EBV国际标准品对NAMALWA细胞原液定值浓度约为2.13×10⁵ IU/mL,部分参评实验室出现假阳性和假阴性问题,高浓度样本(10⁵和10⁴ IU/ml)的符合率均为100%,低浓度样本(10³和10² IU/mL)的符合率分别为92%和40%,阴性样本的符合率为90%。重复样本实验室内检测结果的平均变异系数(CV)为4.1%(1.4%~11.3%),实验室间CV为6%;大多数实验室(75%)的检测结果与预期值呈良好的线性相关。

结论

成功制备可模拟临床标本的EBV PCR检测质控品,并证实其可有效用于临床实验室室间质评。质评结果显示上海地区参评实验室EBV DNA检测质量整体较好,但个别实验室检测能力尚需提高。

关键词: EB病毒, 核酸, 质控品, 室间质量评价

Abstract: Objective

To develop quality control materials for Epstein-Barr virus (EBV) nucleic acid determination, and to apply it into the laboratories participating Shanghai external quality assessment, in order to evaluate the performance of EBV DNA determination.

Methods

The cultured lymphocytes (NAMALWA) known to contain EBV genome were collected as internal control. The applicability of the internal control with commercial EBV kits was evaluated, and the amounts in international units by tracing the internal control to the international standard were established. The panel consisted of 5 blindly coded samples diluted with human plasma and distributed to external quality assessment participates in Shanghai. The results from the participates were summarized and analyzed.

Results

The original NAMALWA cells were all positive with different commercial EBV PCR kits. The concentration of internal control valued by international standard was 2.13×10⁵ IU/mL. False positivity and false negativity errors were found in parts of participating laboratories. The coincidence rates were 100% for high-concentration samples (10⁵ and 10⁴ IU/mL), 92% and 40% for low-concentration samples (10³ and 10² IU/mL) and 90% for negative samples, respectively. For samples replicated in the panels, the average coefficient of variation (CV) for intra-laboratory was 4.1% (1.4%-11.3%), and that for inter-laboratory was 6%. Good overall linearity was observed for most of the laboratories (75%).

Conclusions

The quality control materials for EBV PCR kits have been established successfully, and the applicability in external quality assessment is proved. These results indicate that most of participating laboratories show good performance in determining EBV DNA, and it is necessary for individual laboratories to improve their operation.

Key words: Epstein-Barr virus, Nucleic acid, Quality control material, External quality assessment

中图分类号:

R373.1

王雪亮, 刘芬, 蒋玲丽, 肖艳群, 王华梁. EB病毒核酸检测质控品制备及其应用[J]. 检验医学, 2015, 30(11): 1078-1082.

WANG Xueliang, LIU Fen, JIANG Lingli, XIAO Yanqun, WANG Hualiang. Development and application of quality control materials for EBV nucleic acid determination[J]. Laboratory Medicine, 2015, 30(11): 1078-1082.

图/表 4

图1 细胞原液荧光PCR扩增曲线注:(a)中山达安PCR试剂;(b)湖南圣湘PCR试剂

表1 NAMALWA细胞原液定值检测结果 (IU/mL)

样本	第1次	第2次	第3次
1	2.43×10⁵	1.93×10⁵	2.34×10⁵
2	2.06×10⁵	2.26×10⁵	1.96×10⁵
3	2.11×10⁵	2.12×10⁵	2.03×10⁵

表2 2014年上半年EBV DNA室间质量评价定性和定量结果

样本	预期浓度[IU/mL(Log₁₀)]	定性结果	定量结果[( $x ?$ ±S) copies/mL(Log₁₀)]	实验室内CV (%)	实验室间CV (%)
1	3.69	+	3.80±0.12
2	3.69	+	3.80±0.07
3	3.69	+	3.84±0.13	0.01~0.11	0.06
4	3.69	+	3.80±0.12
5	-	-	/	/	/

表2 2014年上半年EBV DNA室间质量评价定性和定量结果

样本	预期浓度[IU/mL(Log₁₀)]	定性结果	定量结果[( $x ?$ ±S) copies/mL(Log₁₀)]	实验室内CV (%)	实验室间CV (%)
1	3.69	+	3.80±0.12
2	3.69	+	3.80±0.07
3	3.69	+	3.84±0.13	0.01~0.11	0.06
4	3.69	+	3.80±0.12
5	-	-	/	/	/

表3 2014年下半年EBV DNA室间质量评价定性和定量结果

样本	预期浓度 [IU/mL(Log₁₀)]	定性结果	定量结果[( $x ?$ ±S), copies/mL(Log₁₀)]
1	5.33	+	5.46±0.16
2	4.33	+	4.45±0.24
3	3.33	+	3.23±0.31
4	2.33	-	2.26±0.20
5	-	-	/

表3 2014年下半年EBV DNA室间质量评价定性和定量结果

样本	预期浓度 [IU/mL(Log₁₀)]	定性结果	定量结果[( $x ?$ ±S), copies/mL(Log₁₀)]
1	5.33	+	5.46±0.16
2	4.33	+	4.45±0.24
3	3.33	+	3.23±0.31
4	2.33	-	2.26±0.20
5	-	-	/

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