Standardization of Quantitative PCR for Human T-Cell Leukemia Virus Type 1 in Japan: a Collaborative Study.

Abstract

Quantitative PCR (qPCR) analysis of human T-cell leukemia virus type 1 (HTLV-1) was used to assess the amount of HTLV-1 provirus DNA integrated into the genomic DNA of host blood cells. Accumulating evidence indicates that a high proviral load is one of the risk factors for the development of adult T-cell leukemia/lymphoma and HTLV-1-associated myelopathy/tropical spastic paraparesis. However, interlaboratory variability in qPCR results makes it difficult to assess the differences in reported proviral loads between laboratories. To remedy this situation, we attempted to minimize discrepancies between laboratories through standardization of HTLV-1 qPCR in a collaborative study. TL-Om1 cells that harbor the HTLV-1 provirus were serially diluted with peripheral blood mononuclear cells to prepare a candidate standard. By statistically evaluating the proviral loads of the standard and those determined using in-house qPCR methods at each laboratory, we determined the relative ratios of the measured values in the laboratories to the theoretical values of the TL-Om1 standard. The relative ratios of the laboratories ranged from 0.84 to 4.45. Next, we corrected the proviral loads of the clinical samples from HTLV-1 carriers using the relative ratio. As expected, the overall differences between the laboratories were reduced by half, from 7.4-fold to 3.8-fold on average, after applying the correction. HTLV-1 qPCR can be standardized using TL-Om1 cells as a standard and by determining the relative ratio of the measured to the theoretical standard values in each laboratory.

• Authors: Madoka Kuramitsu, Kazu Okuma, Tadanori Yamochi, Tomoo Sato, Daisuke Sasaki, Hiroo Hasegawa, Kazumi Umeki, Ryuji Kubota, Rieko Sobata, Chieko Matsumoto, Noriaki Kaneko, Isao Naruse, Makoto Yamagishi, Makoto Nakashima, Haruka Momose, Kumiko Araki, Takuo Mizukami, Saeko Mizusawa, Yoshiaki Okada, Masaki Ochiai, Atae Utsunomiya, Ki-Ryang Koh, Masao Ogata, Kisato Nosaka, Kaoru Uchimaru, Masako Iwanaga, Yasuko Sagara, Yoshihisa Yamano, Masahiro Satake, Akihiko Okayama, Manabu Mochizuki, Shuji Izumo, Shigeru Saito, Kazuo Itabashi, Shimeru Kamihira, Kazunari Yamaguchi, Toshiki Watanabe, Isao Hamaguchi
• Journal: Journal of clinical microbiology (J Clin Microbiol) Vol. 53 Issue 11 Pg. 3485-91 (Nov 2015) ISSN: 1098-660X [Electronic] United States
• PMID: 26292315 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
• Copyright: Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Chemical References

• DNA, Viral

Topics

• Cell Line, Tumor
• DNA, Viral (analysis, genetics)
• HTLV-I Infections (genetics, virology)
• Human T-lymphotropic virus 1 (genetics)
• Humans
• Japan
• Jurkat Cells
• Leukemia, T-Cell (genetics, virology)
• Leukocytes, Mononuclear (virology)
• Proviruses (genetics)
• Real-Time Polymerase Chain Reaction (methods, standards)
• Viral Load (genetics)
• Virus Integration (genetics)