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Trend of telomerase activity change during human iPSC self-renewal and differentiation revealed by a quartz crystal microbalance based assay.

Zhou Y, Zhou P, Xin Y, Wang J, Zhu Z, Hu J, Wei S, Ma H - Sci Rep (2014)

Bottom Line: Telomerase plays an important role in governing the life span of cells for its capacity to extend telomeres.The pluripotency of hiPSCs during differentiation could be estimated through monitoring telomerase activity and compared with the expression levels of markers of pluripotency gene via quantitative real time PCR.Regular assessment for factors associated with pluripotency or stemness was expensive and requires excessive sample consuming, thus TREAQ could be a promising alternative technology for routine monitoring of telomerase activity and estimate the pluripotency of stem cells.

View Article: PubMed Central - PubMed

Affiliation: Division of Nanobiomedicine, Suzhou Institute of Nano-Tech and Nano-Bionics, Chinese Academy of Sciences, Suzhou 215123, P. R. China.

ABSTRACT
Telomerase plays an important role in governing the life span of cells for its capacity to extend telomeres. As high activity of telomerase has been found in stem cells and cancer cells specifically, various methods have been developed for the evaluation of telomerase activity. To overcome the time-consuming procedures and complicated manipulations of existing methods, we developed a novel method named Telomeric Repeat Elongation Assay based on Quartz crystal microbalance (TREAQ) to monitor telomerase activity during the self-renewal and differentiation of human induced pluripotent stem cells (hiPSCs). TREAQ results indicated hiPSCs possess invariable telomerase activity for 11 passages on Matrigel and a steady decline of telomerase activity when differentiated for different periods, which is confirmed with existing golden standard method. The pluripotency of hiPSCs during differentiation could be estimated through monitoring telomerase activity and compared with the expression levels of markers of pluripotency gene via quantitative real time PCR. Regular assessment for factors associated with pluripotency or stemness was expensive and requires excessive sample consuming, thus TREAQ could be a promising alternative technology for routine monitoring of telomerase activity and estimate the pluripotency of stem cells.

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Illustration of TREAQ on polymer-coated chips.(A) Amido modified primers were immobilized on the polymer surface and extended by telomerase in reaction buffer. (B) The schematic of monitoring of telomerase activity during the self-renewal and differentiation of hiPSCs by QCM.
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f1: Illustration of TREAQ on polymer-coated chips.(A) Amido modified primers were immobilized on the polymer surface and extended by telomerase in reaction buffer. (B) The schematic of monitoring of telomerase activity during the self-renewal and differentiation of hiPSCs by QCM.

Mentions: In this study, we introduce TREAQ detection, a rapid and PCR-free method, easily to detect telomerase activity. By mimicking telomere elongation in vivo, we can monitor the behavior of telomerase extending the primer in real time on different surfaces (gold chips and polymer-coated chips) by FT (the absolute value of QCM resonance frequency shift (Δf) caused by extension of primers), which mirrors the activity of telomerase (Fig. 1A). Our results suggest that TREAQ is a helpful tool to rapidly carry out high-throughput detections of telomerase activity (using off-line detection method introduced in Results section) and has wide application prospects in life science and biomedical research.


Trend of telomerase activity change during human iPSC self-renewal and differentiation revealed by a quartz crystal microbalance based assay.

Zhou Y, Zhou P, Xin Y, Wang J, Zhu Z, Hu J, Wei S, Ma H - Sci Rep (2014)

Illustration of TREAQ on polymer-coated chips.(A) Amido modified primers were immobilized on the polymer surface and extended by telomerase in reaction buffer. (B) The schematic of monitoring of telomerase activity during the self-renewal and differentiation of hiPSCs by QCM.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4225532&req=5

f1: Illustration of TREAQ on polymer-coated chips.(A) Amido modified primers were immobilized on the polymer surface and extended by telomerase in reaction buffer. (B) The schematic of monitoring of telomerase activity during the self-renewal and differentiation of hiPSCs by QCM.
Mentions: In this study, we introduce TREAQ detection, a rapid and PCR-free method, easily to detect telomerase activity. By mimicking telomere elongation in vivo, we can monitor the behavior of telomerase extending the primer in real time on different surfaces (gold chips and polymer-coated chips) by FT (the absolute value of QCM resonance frequency shift (Δf) caused by extension of primers), which mirrors the activity of telomerase (Fig. 1A). Our results suggest that TREAQ is a helpful tool to rapidly carry out high-throughput detections of telomerase activity (using off-line detection method introduced in Results section) and has wide application prospects in life science and biomedical research.

Bottom Line: Telomerase plays an important role in governing the life span of cells for its capacity to extend telomeres.The pluripotency of hiPSCs during differentiation could be estimated through monitoring telomerase activity and compared with the expression levels of markers of pluripotency gene via quantitative real time PCR.Regular assessment for factors associated with pluripotency or stemness was expensive and requires excessive sample consuming, thus TREAQ could be a promising alternative technology for routine monitoring of telomerase activity and estimate the pluripotency of stem cells.

View Article: PubMed Central - PubMed

Affiliation: Division of Nanobiomedicine, Suzhou Institute of Nano-Tech and Nano-Bionics, Chinese Academy of Sciences, Suzhou 215123, P. R. China.

ABSTRACT
Telomerase plays an important role in governing the life span of cells for its capacity to extend telomeres. As high activity of telomerase has been found in stem cells and cancer cells specifically, various methods have been developed for the evaluation of telomerase activity. To overcome the time-consuming procedures and complicated manipulations of existing methods, we developed a novel method named Telomeric Repeat Elongation Assay based on Quartz crystal microbalance (TREAQ) to monitor telomerase activity during the self-renewal and differentiation of human induced pluripotent stem cells (hiPSCs). TREAQ results indicated hiPSCs possess invariable telomerase activity for 11 passages on Matrigel and a steady decline of telomerase activity when differentiated for different periods, which is confirmed with existing golden standard method. The pluripotency of hiPSCs during differentiation could be estimated through monitoring telomerase activity and compared with the expression levels of markers of pluripotency gene via quantitative real time PCR. Regular assessment for factors associated with pluripotency or stemness was expensive and requires excessive sample consuming, thus TREAQ could be a promising alternative technology for routine monitoring of telomerase activity and estimate the pluripotency of stem cells.

Show MeSH
Related in: MedlinePlus