Establishment of internally controlled Real-time PCR for the detection of human parvovirus B19 DNA in serum
10.3760/cma.j.issn.1003-9279.2010.06.028
- VernacularTitle:建立设有内对照的Real-time PCR方法检测人血清中细小病毒B19
- Author:
Meng LI
1
;
Xiao-Hui ZOU
;
Min WANG
;
Xiu-Ping YU
;
Zhuo-Zhuang LU
;
Tao HONG
Author Information
1. 山东大学医学院
- Keywords:
Polymerase Chain reaction;
Parvovirus B19,human;
Serologic tests;
Internal
- From:
Chinese Journal of Experimental and Clinical Virology
2010;24(6):479-481
- CountryChina
- Language:Chinese
-
Abstract:
Objective To establish a TaqMan-based Real-time PGR assay with internal control for the detection of parvovirus B19 DNA in human serum. Methods Two DNA fragments in length of 113 bp each were artificially synthesized, cloned into T vector and used as standard DNA or internal control,respectively. One pair of primers and two probes were included in the Real-time PGR. The probes were labeled with different fluoresceins and could bind B19 DNA or internal control, respectively. Precision and specificity of the method were evaluated. Specimens of human serum were examined by this assay to find B19DNA. Results The standard curve was constructed using the quantified standard B19 DNA. The Real-time PGR method was established. It was stable according to precision evaluation by the intra- and inter-assay and specific without any evident cross-reaction with human hepatitis B virus (HBV). Among 160 samples of human serum, B19 DNA was detected in 2 with a concentration of 2. 1 × 105 Geq/ml and 3.6 × 103 Geq/ml,respectively. Conclusion The Real-time PGR for B19 DNA detection was developed successfully, in which the internal control was helpful to exclude false-negative results.
