title: Continuous collection and simultaneous detection of picoliter volume of nucleic acid samples using a mille-feuille probe.
abstract: Investigation of the positional heterogeneity of messenger RNA (mRNA) expression in tissues requires a technology that facilitates analysis of mRNA expression in the selected single cells. We developed a mille-feuille probe (MP) that allows the lamination of the aqueous and organic phases in a nanopipette under voltage control. The MP was used for continuous collection of different nucleic acid samples and sequential evaluation of gene expression with mRNA barcoding tags. First, we found that the aqueous phases could be laminated into five individual layers and separated by the plugs of the organic phases in a nanopipette when the salt (THATPBCl) concentration in the organic phase was 100 mM. Second, the aspiration rate of the MP was stabilized and the velocity of the aqueous phase in the MP was lowered at higher THATPBCl concentrations in the organic phase. This was because the force during ingression of the aqueous phase into the organic - phase-filled nanopipette induced an electro-osmotic flow between the inside wall of the nanopipette and THATPBCl in the organic phase. Third, inclusion of mRNA barcoding tags in the MP facilitated complementary DNA construction and sequential analysis of gene expression. This technique has potential to be applicable to RNA sequencing from different cell samples across the life sciences. Graphical abstract We developed a mille-feuille probe (MP) that allows the lamination of the aqueous and organic phases in a nanopipette under voltage control.
authors.affiliation: Graduate School of Environmental Studies, Tohoku University, Aramaki-aoba 6-6-11-604, Sendai, 980-8579, Japan.|Department of Tumor Cell Biology, Graduate School of Biological Sciences, Nara Institute of Science and Technology, Ikoma, 630-0192, Nara, Japan.
authors.forename: Hidenori
authors.initials: H
authors.lastname: Ito
authors.affiliation: Graduate School of Environmental Studies, Tohoku University, Aramaki-aoba 6-6-11-604, Sendai, 980-8579, Japan.
authors.forename: Motoki
authors.initials: M
authors.lastname: Tanaka
authors.affiliation: WPI-Advanced Institute for Materials Research, Tohoku University, 2-1-1 Katahira, Aoba-ku, Sendai, 980-8577, Japan.|Division of Electrical Engineering and Computer Science, Kanazawa University, Ishikawa, Kanazawa, 920-1192, Japan.
authors.forename: Yuanshu
authors.initials: Y
authors.lastname: Zhou
authors.affiliation: Graduate School of Environmental Studies, Tohoku University, Aramaki-aoba 6-6-11-604, Sendai, 980-8579, Japan.|Department of Microengineering, Graduate School of Engineering, Kyoto University, Kyoto, 615-8540, Japan.
authors.forename: Yuji
authors.initials: Y
authors.lastname: Nashimoto
authors.affiliation: Division of Electrical Engineering and Computer Science, Kanazawa University, Ishikawa, Kanazawa, 920-1192, Japan.|PRESTO, JST, 4-1-8, Honcho, Kawaguchi-shi, Saitama, 332-0012, Japan.
authors.forename: Yasufumi
authors.initials: Y
authors.lastname: Takahashi
authors.affiliation: Graduate School of Environmental Studies, Tohoku University, Aramaki-aoba 6-6-11-604, Sendai, 980-8579, Japan.
authors.forename: Kosuke
authors.initials: K
authors.lastname: Ino
authors.affiliation: Graduate School of Environmental Studies, Tohoku University, Aramaki-aoba 6-6-11-604, Sendai, 980-8579, Japan. matsue@bioinfo.che.tohoku.ac.jp.|WPI-Advanced Institute for Materials Research, Tohoku University, 2-1-1 Katahira, Aoba-ku, Sendai, 980-8577, Japan. matsue@bioinfo.che.tohoku.ac.jp.
authors.forename: Tomokazu
authors.initials: T
authors.lastname: Matsue
authors.affiliation: Department of Applied Chemistry, Graduate School of Engineering, Tohoku University, Sendai, 980-8579, Japan. shiku@bioinfo.che.tohoku.ac.jp.
authors.forename: Hitoshi
authors.initials: H
authors.lastname: Shiku
mesh_terms: D001483:Base Sequence; D018076:DNA, Complementary / Q000032:analysis*; D006801:Humans; D057230:Limit of Detection; D061986:MCF-7 Cells; D015335:Molecular Probes; D016133:Polymerase Chain Reaction / Q000379:methods; D012333:RNA, Messenger / Q000032:analysis*
chemical_list: D018076:DNA, Complementary; D015335:Molecular Probes; D012333:RNA, Messenger
keywords: Liquid/liquid interface; Microfluidics; PCR; mRNA