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Cells 2018, 7(11), 183; https://doi.org/10.3390/cells7110183

Telomere Length Calibration from qPCR Measurement: Limitations of Current Method

1
Clinical Genetics Branch, Division of Cancer Epidemiology and Genetics, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA
2
Terry Fox Laboratory, British Columbia Cancer Agency, Vancouver, BC V5Z 1L3, Canada
3
Repeat Diagnostics Inc., North Vancouver, BC V7M 1A5, Canada
4
Cancer Genomics Research Laboratory, Division of Cancer Epidemiology and Genetics, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA
5
Leidos Biomedical Research, Inc., Frederick National Laboratory for Cancer Research, Frederick, MD 21701, USA
6
Center for International Blood and Marrow Transplant Research, Minneapolis, MN 55401, USA
7
Center for International Blood and Marrow Transplant Research, Medical College of Wisconsin, Milwaukee, WI 53226, USA
8
Clinical Research Division, Fred Hutchinson Cancer Research Center, Seattle, WA 98109, USA
9
Biostatistics Branch, Division of Cancer Epidemiology and Genetics, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA
*
Author to whom correspondence should be addressed.
Received: 29 August 2018 / Revised: 19 October 2018 / Accepted: 21 October 2018 / Published: 24 October 2018
(This article belongs to the Special Issue The Role of Telomere Biology in Aging and Human Disease)
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Abstract

Telomere length (TL) comparisons from different methods are challenging due to differences in laboratory techniques and data configuration. This study aimed to assess the validity of converting the quantitative polymerase chain reaction (qPCR) telomere/single copy gene (T/S) ratio to TL in kilobases (kb). We developed a linear regression equation to predict TL from qPCR T/S using flow cytometry with fluorescence in situ hybridization (flow FISH) TL data from 181 healthy donors (age range = 19–53) from the National Marrow Donor Program (NMDP) biorepository. TL measurements by qPCR and flow FISH were modestly correlated (R2 = 0.56, p < 0.0001). In Bland-Altman analyses, individuals with the shortest (≤10th percentile) or longest (≥90th) flow FISH TL had an over- or under-estimated qPCR TL (bias = 0.89 and −0.77 kb, respectively). Comparisons of calculated TL from the NMDP samples and 1810 age- and sex-matched individuals from the National Health and Nutrition Examination Survey showed significant differences (median = 7.1 versus 5.8 kb, respectively, p < 0.0001). Differences in annual TL attrition were also noted (31 versus 13 bp/year, respectively, p = 0.02). Our results demonstrate that TL calculated in kb from qPCR T/S may yield biased estimates for individuals with the shortest or longest TL, those often of high clinical interest. We also showed that calculated TL in kb from qPCR data are not comparable across populations and therefore are not necessarily useful. View Full-Text
Keywords: telomere length; qPCR; flow FISH; agreement telomere length; qPCR; flow FISH; agreement
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Wang, Y.; Savage, S.A.; Alsaggaf, R.; Aubert, G.; Dagnall, C.L.; Spellman, S.R.; Lee, S.J.; Hicks, B.; Jones, K.; Katki, H.A.; Gadalla, S.M. Telomere Length Calibration from qPCR Measurement: Limitations of Current Method. Cells 2018, 7, 183.

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