OBJECTIVETo establish and evaluate the methods of internal quality control in blood donor screening by nucleic acid test (NAT).

METHODSAfter HBV-DNA standard quality control (QC) sample (60 IU/ml) was diluted by pooling 6 samples, the concentration was 10.0 IU/ml, which was approach twice of the low limit. When the pooling result turned out reactive, the pooling samples need to be split into single sample to process. Meanwhile, the standard QC samples were tested as well. The same batch QC samples were tested 20 times respectively, calculate the mean (x̄), standard deviation (SD) and CV. Make Levey-Jennings QC curves by setting x̄±2SD as warning, x̄±3SD as rejected. The Levey-Jennings quality controls chart was mapped by using Microsoft Excel.

RESULTSAfter 20 times test of mixed/split samples, the x̄±2SD were 33.03±1.47 and 30.08±0.98, the x̄±3SD were 33.03±2.20 and 30.08±1.47, the CV were 2.22% and 1.63%, respectively. The P value of t test was 0.08 and 0.17 respectively, there was no statistically significant difference between the 2 group.

CONCLUSIONWhen establish an internal QC system in the screening laboratory by nucleic acid testing, the concentration of the QC samples should be equal to normal specimens. This type of QC system may validate the extraction and amplification of the nucleic acid, and improve the stability of the test results.

Blood Donors ; Donor Selection ; Humans ; Nucleic Acid Amplification Techniques ; standards ; Quality Control

OBJECTIVESTo establish a Chinese national standard for a nucleic acid test (NAT) for HBV DNA.

METHODSThe candidate sample of HBV DNA positive plasma was diluted with HBV-negative human plasma. The sample was lyophilised with a concentration of approximately 300,000 copies/ml. The measurement methods used included Roche Amplicor assay (version 2.0) and real-time PCR. The lyophilised preparation was calibrated by the international standard (NIBSC code: 97/746) from NIBSC.

RESULTSThe quantity of this lyophilised preparation was (1.29+/-0.24) x 10(5)IU/ml in comparison with the international standard for HBV DNA 97/746. The stability test indicated that the sample was stable at room temperature (20 to 25 degrees C) for 2 weeks and at 37 degrees C for at least 1 week. Long-term stability was observed at 2 to 8 degrees C for 6 months and at -20 degrees C for more than 2 years with no significant changes. The vial-to-vial imprecision rate was 3.53%.

CONCLUSIONBased on the results of this study, our lyophilized sample can be used as a standard in China for the nucleic acid test (NAT) for HBV DNA.

DNA, Viral ; blood ; Hepatitis B virus ; genetics ; Humans ; Nucleic Acid Amplification Techniques ; standards ; Plasma ; chemistry

Objective To evaluate the efficiency of REPLI-g® Single Cell Kit for sample DNA amplification, and explore its application value in forensic trace DNA amplification. Methods Three DNA extraction kits were selected to extract DNA from peripheral blood of 10 unrelated individuals. The DNA yield and purity of the three DNA extraction kits were compared. According to the results of comparison, one DNA sample was selected to concentrate and dilute, then used as the initial sample of whole genome amplification （WGA）. REPLI-g® Single Cell Kit was used to amplify the initial sample at the whole genome level. The amplification yield and amplification times were calculated, and the distribution of DNA fragments was detected by agarose gel electrophoresis. Goldeneye® DNA ID System 20A Kit was used to perform the STR typing of the initial sample and DNA samples amplified at the whole genome level to evaluate the performance of REPLI-g® Single Cell Kit in trace DNA amplication in terms of purity and yield as well as the success rate of STR typing. Results After comparison, one DNA sample was selected from QIAsymphony® DNA Investigator® Kit extracts to concentrate and dilute as the initial sample of WGA. After amplifying the whole genome of a series of initial samples by REPLI-g® Single Cell Kit, the lowest average of amplification yield reached 8.77×103 ng, while the average of the corresponding amplification times reached 1.40×106. DNA fragments were large and concentrated. The STR typing success rate of WGA samples became lower with the decrease of initial samples used, but when the amount of samples was lower than 0.5 ng, the STR typing success rate of samples after DNA WGA was higher than that of samples without DNA WGA. Conclusion REPLI-g® Single Cell Kit can increase the yield of template DNA. Especially for trace DNA, the STR typing success rate can be improved to a certain extent.
DNA ; DNA Fingerprinting ; Humans ; Microsatellite Repeats ; Nucleic Acid Amplification Techniques/standards* ; Sequence Analysis, DNA/methods*