PMID- 25233459 OWN - NLM STAT- MEDLINE DCOM- 20160426 LR - 20211021 IS - 1932-6203 (Electronic) IS - 1932-6203 (Linking) VI - 9 IP - 9 DP - 2014 TI - Parallel single cancer cell whole genome amplification using button-valve assisted mixing in nanoliter chambers. PG - e107958 LID - 10.1371/journal.pone.0107958 [doi] LID - e107958 AB - The heterogeneity of tumor cells and their alteration during the course of the disease urges the need for real time characterization of individual tumor cells to improve the assessment of treatment options. New generations of therapies are frequently associated with specific genetic alterations driving the need to determine the genetic makeup of tumor cells. Here, we present a microfluidic device for parallel single cell whole genome amplification (pscWGA) to obtain enough copies of a single cell genome to probe for the presence of treatment targets and the frequency of its occurrence among the tumor cells. Individual cells were first captured and loaded into eight parallel amplification units. Next, cells were lysed on a chip and their DNA amplified through successive introduction of dedicated reagents while mixing actively with the help of integrated button-valves. The reaction chamber volume for scWGA 23.85 nl, and starting from 6-7 pg DNA contained in a single cell, around 8 ng of DNA was obtained after WGA, representing over 1000-fold amplification. The amplified products from individual breast cancer cells were collected from the device to either directly investigate the amplification of specific genes by qPCR or for re-amplification of the DNA to obtain sufficient material for whole genome sequencing. Our pscWGA device provides sufficient DNA from individual cells for their genetic characterization, and will undoubtedly allow for automated sample preparation for single cancer cell genomic characterization. FAU - Yang, Yoonsun AU - Yang Y AD - Medical Cell BioPhysics Group, MIRA Institute for Biomedical Technology and Technical Medicine, University of Twente, Enschede, The Netherlands. FAU - Swennenhuis, Joost F AU - Swennenhuis JF AD - Medical Cell BioPhysics Group, MIRA Institute for Biomedical Technology and Technical Medicine, University of Twente, Enschede, The Netherlands. FAU - Rho, Hoon Suk AU - Rho HS AD - Mesoscale Chemical Systems Group, MESA+ Institute for Nanotechnology, University of Twente, Enschede, The Netherlands. FAU - Le Gac, Severine AU - Le Gac S AD - BIOS Lab on a Chip Group, MESA+ Institute for Nanotechnology, University of Twente, Enschede, The Netherlands. FAU - Terstappen, Leon W M M AU - Terstappen LW AD - Medical Cell BioPhysics Group, MIRA Institute for Biomedical Technology and Technical Medicine, University of Twente, Enschede, The Netherlands. LA - eng PT - Journal Article PT - Research Support, Non-U.S. Gov't DEP - 20140918 PL - United States TA - PLoS One JT - PloS one JID - 101285081 SB - IM MH - Breast Neoplasms/*genetics MH - Escherichia coli/genetics MH - Female MH - Genome, Bacterial MH - Genome, Human MH - Humans MH - Lab-On-A-Chip Devices MH - MCF-7 Cells MH - Nucleic Acid Amplification Techniques MH - *Sequence Analysis, DNA MH - Single-Cell Analysis PMC - PMC4169497 COIS- Competing Interests: This study is funded by NanoNextNL, a micro and nanotechnology consortium of companies, universities, knowledge institutes and university medical centers. There are no patents, products in development or marketed products to declare. This does not alter the authors' adherence to all the PLOS ONE policies on sharing data and materials. EDAT- 2014/09/19 06:00 MHDA- 2016/04/27 06:00 PMCR- 2014/09/18 CRDT- 2014/09/19 06:00 PHST- 2014/04/30 00:00 [received] PHST- 2014/08/16 00:00 [accepted] PHST- 2014/09/19 06:00 [entrez] PHST- 2014/09/19 06:00 [pubmed] PHST- 2016/04/27 06:00 [medline] PHST- 2014/09/18 00:00 [pmc-release] AID - PONE-D-14-19393 [pii] AID - 10.1371/journal.pone.0107958 [doi] PST - epublish SO - PLoS One. 2014 Sep 18;9(9):e107958. doi: 10.1371/journal.pone.0107958. eCollection 2014.