Evaluating minimally invasive sample collection methods for telomere length measurement

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Abstract

<div class="section"> <a class="named-anchor" id="S1">  </a> <h5 class="section-title" id="d1014641e170">Objectives</h5> <p id="P1">Telomere length (TL) is a biomarker of aging and age-related decline. Although venous blood is considered the “gold standard” for TL measurement, its collection is often not feasible or desired in non-clinical settings. Saliva and dried blood spots (DBS) have been used as alternatives when venipuncture cannot be performed. However, it is not known whether these sample types yield TL measurements comparable to those obtained from venous blood. We sought to determine whether different samples from the same individual yield comparable TL measurements. </p> </div><div class="section"> <a class="named-anchor" id="S2">  </a> <h5 class="section-title" id="d1014641e175">Methods</h5> <p id="P2">We extracted DNA from matched buffy coat, saliva (Oragene and Oasis), and DBS (venous and capillary) samples from 40 women aged 18 to 77 years. We used the monochrome multiplex qPCR (MMQPCR) assay to measure TL in all sample types for each participant and applied quality control measures to retain only high-quality samples for analysis. We then compared TL from buffy coat and saliva to examine how these measurements differ and to test if TL is correlated across sample types. </p> </div><div class="section"> <a class="named-anchor" id="S3">  </a> <h5 class="section-title" id="d1014641e180">Results</h5> <p id="P3">TL differed significantly across buffy coat, Oragene saliva, and Oasis saliva samples. TL from buffy coat and Oragene saliva was moderately correlated (ρ = 0.48, <i>p =</i> 0.002) and the most similar in size. Oasis saliva TL was not correlated with buffy coat or Oragene saliva TL, and was the shortest. DBS DNA yields were inadequate for TL measurement using the MMQPCR assay. </p> </div><div class="section"> <a class="named-anchor" id="S4">  </a> <h5 class="section-title" id="d1014641e188">Conclusions</h5> <p id="P4">Using a matched dataset we demonstrate that sample type significantly influences the TL measurement obtained using the MMQPCR assay. </p> </div>

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What a drop can do: dried blood spots as a minimally invasive method for integrating biomarkers into population-based research.

Thomas McDade, Sharon Williams, James Snodgrass (2007)

Logistical constraints associated with the collection and analysis of biological samples in community-based settings have been a significant impediment to integrative, multilevel bio-demographic and biobehavioral research. However recent methodological developments have overcome many of these constraints and have also expanded the options for incorporating biomarkers into population-based health research in international as well as domestic contexts. In particular using dried blood spot (DBS) samples-drops of whole blood collected on filter paper from a simple finger prick-provides a minimally invasive method for collecting blood samples in nonclinical settings. After a brief discussion of biomarkers more generally, we review procedures for collecting, handling, and analyzing DBS samples. Advantages of using DBS samples-compared with venipuncture include the relative ease and low cost of sample collection, transport, and storage. Disadvantages include requirements for assay development and validation as well as the relatively small volumes of sample. We present the results of a comprehensive literature review of published protocols for analysis of DBS samples, and we provide more detailed analysis of protocols for 45 analytes likely to be of particular relevance to population-level health research. Our objective is to provide investigators with the information they need to make informed decisions regarding the appropriateness of blood spot methods for their research interests.

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Evidence for a mitotic clock in human hematopoietic stem cells: loss of telomeric DNA with age.

Rebecca Allsopp, Peter M. Lansdorp, L. Thomas … (1994)

The proliferative life-span of the stem cells that sustain hematopoiesis throughout life is not known. It has been proposed that the sequential loss of telomeric DNA from the ends of human chromosomes with each somatic cell division eventually reaches a critical point that triggers cellular senescence. We now show that candidate human stem cells with a CD34+CD38lo phenotype that were purified from adult bone marrow have shorter telomeres than cells from fetal liver or umbilical cord blood. We also found that cells produced in cytokine-supplemented cultures of purified precursor cells show a proliferation-associated loss of telomeric DNA. These findings strongly suggest that the proliferative potential of most, if not all, hematopoietic stem cells is limited and decreases with age, a concept that has widespread implications for models of normal and abnormal hematopoiesis as well as gene therapy.

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Impartial comparative analysis of measurement of leukocyte telomere length/DNA content by Southern blots and qPCR

Abraham Aviv, Steven Hunt, Jue Lin … (2011)

Telomere length/DNA content has been measured in epidemiological/clinical settings with the goal of testing a host of hypotheses related to the biology of human aging, but often the conclusions of these studies have been inconsistent. These inconsistencies may stem from various reasons, including the use of different telomere length measurement techniques. Here, we report the first impartial evaluation of measurements of leukocyte telomere length by Southern blot of the terminal restriction fragments and quantitative PCR (qPCR) of telomere DNA content, expressed as the ratio of telomeric product (T)/single copy gene (S) product. Blind measurements on the same samples from 50 donors were performed in two independent laboratories on two different occasions. Both the qPCR and Southern blots displayed highly reproducible results as shown by r values > 0.9 for the correlations between results obtained by either method on two occasions. The inter-assay CV measurement for the qPCR was 6.45%, while that of the Southern blots was 1.74%. The relation between the results generated by Southern blots versus those generated by qPCR deviated from linearity. We discuss the ramifications of these findings with regard to measurements of telomere length/DNA content in epidemiological/clinical circumstances.