Get access

Real-time amplification of glyceraldehyde-3-phosphate dehydrogenase gene for quality control of leukopoor platelets

Authors

  • Ding-Ping Chen,

    1. Department of Laboratory Medicine, Chang Gung Memorial Hospital
    2. Department of Medical Biotechnology and Laboratory Science, College of Medicine, Department of Pathology, School of Medicine, Molecular Medicine Research Center, Chang Gung University, Taoyuan County, Taiwan
    Search for more papers by this author
  • Chien-Feng Sun,

    1. Department of Laboratory Medicine, Chang Gung Memorial Hospital
    2. Department of Medical Biotechnology and Laboratory Science, College of Medicine, Department of Pathology, School of Medicine, Molecular Medicine Research Center, Chang Gung University, Taoyuan County, Taiwan
    Search for more papers by this author
  • Hsiao-Chen Ning,

    1. Department of Laboratory Medicine, Chang Gung Memorial Hospital
    2. Department of Medical Biotechnology and Laboratory Science, College of Medicine, Department of Pathology, School of Medicine, Molecular Medicine Research Center, Chang Gung University, Taoyuan County, Taiwan
    Search for more papers by this author
  • Chien-Ting Peng,

    1. Department of Laboratory Medicine, Chang Gung Memorial Hospital
    2. Department of Medical Biotechnology and Laboratory Science, College of Medicine, Department of Pathology, School of Medicine, Molecular Medicine Research Center, Chang Gung University, Taoyuan County, Taiwan
    Search for more papers by this author
  • Wei-Ting Wang,

    1. Department of Laboratory Medicine, Chang Gung Memorial Hospital
    2. Department of Medical Biotechnology and Laboratory Science, College of Medicine, Department of Pathology, School of Medicine, Molecular Medicine Research Center, Chang Gung University, Taoyuan County, Taiwan
    Search for more papers by this author
  • Ching-Ping Tseng

    Corresponding author
    1. Department of Laboratory Medicine, Chang Gung Memorial Hospital
    2. Department of Medical Biotechnology and Laboratory Science, College of Medicine, Department of Pathology, School of Medicine, Molecular Medicine Research Center, Chang Gung University, Taoyuan County, Taiwan
    • Address reprint requests to: Ching-Ping Tseng, Department of Medical Biotechnology and Laboratory Science, Chang Gung University, Taoyuan County, 333, Taiwan; e-mail: ctseng@mail.cgu.edu.tw.

    Search for more papers by this author

  • This work was partly supported by the NSC 98-2320-B-182A-006 and NSC 99-2320-B-182A-006MY3 from the National Science Council, Taiwan.

Abstract

Background

Leukoreduction of blood products is crucial to prevent white blood cell (WBC)-associated complications during transfusion. Of the widely accepted methods for quantifying WBCs in blood components, Nageotte hemocytometry is time-consuming and laborious whereas a specialized instrument is required for flow cytometry. A reliable and affordable method to assess WBC count in blood products is of particular interest.

Study Design and Methods

Real-time polymerase chain reaction (PCR) of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) gene was developed for quantifying WBCs in leukopoor platelets (LPPs). After normalization by the cell-free prefiltrated and postfiltrated plasma DNA, the relative copy number of GAPDH gene in the platelet (PLT) concentrate and its corresponding LPPs was calculated according to the equation of 2−ΔΔCt of which Ct is defined as the threshold cycle. The percentage and the number of WBCs that remained in LPPs were consequently determined. This method was compared to Nageotte hemocytometry and was validated by using serially diluted PLT concentrate and 10 pairs of PLT concentrate-LPP samples.

Results

Consistent with the removal of WBCs after filtration, the Ct values for the LPP samples were increased when compared to their corresponding PLT concentrate. As revealed by real-time PCR of GAPDH gene, there is a correlation between the calculated and theoretical WBC count in the serially diluted PLT concentrate (correlation coefficient, 0.9532). The WBC counts for the 10 LPP samples were comparable between Nageotte and real-time PCR method and were all below 3.3 × 106 WBCs/L.

Conclusion

The real-time PCR method we report in this study is applicable for routine quality assurance during leukoreduction process.

Get access to the full text of this article

Ancillary