Single-cell PCR of genomic DNA enabled by automated single-cell printing for cell isolation

Biosensors & Bioelectronics
F StumpfG Roth

Abstract

Single-cell analysis has developed into a key topic in cell biology with future applications in personalized medicine, tumor identification as well as tumor discovery (Editorial, 2013). Here we employ inkjet-like printing to isolate individual living single human B cells (Raji cell line) and load them directly into standard PCR tubes. Single cells are optically detected in the nozzle of the microfluidic piezoelectric dispenser chip to ensure printing of droplets with single cells only. The printing process has been characterized by using microbeads (10µm diameter) resulting in a single bead delivery in 27 out of 28 cases and relative positional precision of ±350µm at a printing distance of 6mm between nozzle and tube lid. Process-integrated optical imaging enabled to identify the printing failure as void droplet and to exclude it from downstream processing. PCR of truly single-cell DNA was performed without pre-amplification directly from single Raji cells with 33% success rate (N=197) and Cq values of 36.3±2.5. Additionally single cell whole genome amplification (WGA) was employed to pre-amplify the single-cell DNA by a factor of >1000. This facilitated subsequent PCR for the same gene yielding a success rate of 64% (N=33) whi...Continue Reading

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Citations

Jul 28, 2015·International Journal of Molecular Sciences·Andre GrossPeter Koltay
Nov 17, 2015·Analytical Chemistry·Sanjin HosicAbigail N Koppes
Oct 23, 2016·Talanta·Hui DongJianping Zheng
Oct 8, 2016·Lab on a Chip·Christian D AhrbergBong Geun Chung
Dec 21, 2017·BMC Biotechnology·Jacob J MessnerDeirdre R Meldrum
Sep 5, 2018·The Analyst·Qiushi HuangJin-Ming Lin
Dec 22, 2019·Cellular and Molecular Neurobiology·Yuanyuan ZhuShengxi Wu
Sep 6, 2017·Nanoscale·Josephine Geertsen KellerBirgitta Ruth Knudsen
Sep 10, 2018·ACS Biomaterials Science & Engineering·Supeng DingRui Yao
Sep 10, 2020·Chemical Reviews·Xinda LiTao Xu

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