Korean J Blood Transfus.  2019 Apr;30(1):57-64. 10.17945/kjbt.2019.30.1.57.

Comparison of the Effectiveness in the Application of Competitive and Noncompetitive Internal Control for the Laboratory Developed Polymerase Chain Reaction

• Shin S
• Kang JW
• Kang JW
• Seo YI
• Min H

• KMID: 2457203
• DOI: http://doi.org/10.17945/kjbt.2019.30.1.57

Abstract

BACKGROUND
A nucleic acid amplification test was adopted to detect transfusion-transmitted infectious agents. In the case of HTLV, however, there was no internal control (IC) because the laboratory developed polymerase chain reaction (laboratory-developed PCR) was used. In this study, noncompetitive IC was constructed for the laboratory-developed PCR of HTLV and the effectiveness was compared with the competitive test that was constructed in a previous study.
METHODS
As a competitive IC, plasmid DNA, including the primer recognition sequence for the amplification of the HTLV pX region, was constructed. As a noncompetitive IC, an additional primer was constructed for the amplification of the housekeeping gene, the glyceraldehyde 3-phosphate dehydrogenase (GAPDH) gene. The performance of the competitive and noncompetitive IC was verified and compared using 10 HTLV positive samples and 10 negative samples. In addition, the detection limits in the assay adopting competitive IC and noncompetitive IC were compared.
RESULTS
In the case of competitive IC applications, all 10 positive samples were positive and all 10 negative samples were negative. In the case of noncompetitive IC applications, however, one positive sample was not detected. The detection limit of the assay using competitive IC was 100 pg and that of the assay using noncompetitive IC was 1 ng.
CONCLUSION
Although the manufacturing processes is not required using noncompetitive IC, the adoption of competitive IC is more effective to ensure the assay results because the ability of detection of the assay adopting competitive IC was better than that using noncompetitive IC.

Keyword

Human T-cell lymphotropic virus; Laboratory-developed PCR; Competitive internal control; Noncompetitive internal control

MeSH Terms

DNA
Genes, Essential
Glyceraldehyde 3-Phosphate
Limit of Detection
Nucleic Acid Amplification Techniques
Oxidoreductases
Plasmids
Polymerase Chain Reaction*
DNA
Glyceraldehyde 3-Phosphate
Oxidoreductases