PMID- 30424205
OWN - NLM
STAT- PubMed-not-MEDLINE
LR  - 20200930
IS  - 2072-666X (Print)
IS  - 2072-666X (Electronic)
IS  - 2072-666X (Linking)
VI  - 9
IP  - 6
DP  - 2018 May 30
TI  - Effects of Microchannel Shape and Ultrasonic Mixing on Microfluidic Padlock Probe 
      Rolling Circle Amplification (RCA) Reactions.
LID - 10.3390/mi9060272 [doi]
LID - 272
AB  - The fluorescence in situ hybridization (FISH)-based padlock probe and rolling 
      circle amplification (RCA) method allows for the detection of point mutations. 
      However, it requires multiple reaction steps and solution exchanges, making it 
      costly, labor-intensive, and time-consuming. In this study, we aimed to improve 
      the efficiency of padlock/RCA by determining the effects of microchannel shape 
      and ultrasonic solution mixing. Using a circular-shaped microchamber and 
      ultrasonic mixing, the efficiency of microfluidic padlock/RCA was improved, and 
      the consumption of the expensive probe solution was reduced from 10 microL to 
      approximately 3.5 microL. Moreover, the fluorescent probe hybridization time was 
      reduced to 5 min, which is four times faster than that of the standard protocol. 
      We used this method to successfully detect mitochondrial DNA and transcripts of 
      beta-actin and K-ras proto-oncogene codon 12 in cells. Our method offers 
      improvements over current padlock/RCA methods and will be helpful in optimizing 
      other microfluidics-based FISH-related analyses.
FAU - Ishigaki, Yuri
AU  - Ishigaki Y
AD  - Department of Chemical and Biological Sciences, Faculty of Science, Japan Women's 
      University, Bunkyo, Tokyo 112-8681, Japan. kaesato@icl.t.u-tokyo.ac.jp.
FAU - Sato, Kae
AU  - Sato K
AD  - Department of Chemical and Biological Sciences, Faculty of Science, Japan Women's 
      University, Bunkyo, Tokyo 112-8681, Japan. satouk@fc.jwu.ac.jp.
LA  - eng
GR  - JP15K15199/Japan Society for the Promotion of Science/
GR  - 09E51018a/New Energy and Industrial Technology Development Organization/
PT  - Journal Article
DEP - 20180530
PL  - Switzerland
TA  - Micromachines (Basel)
JT  - Micromachines
JID - 101640903
PMC - PMC6187661
OTO - NOTNLM
OT  - fluorescent hybridization
OT  - microfluidics
OT  - padlock probe
OT  - rolling circle amplification
COIS- The authors declare no conflicts of interest.
EDAT- 2018/11/15 06:00
MHDA- 2018/11/15 06:01
PMCR- 2018/05/30
CRDT- 2018/11/15 06:00
PHST- 2018/04/23 00:00 [received]
PHST- 2018/05/22 00:00 [revised]
PHST- 2018/05/29 00:00 [accepted]
PHST- 2018/11/15 06:00 [entrez]
PHST- 2018/11/15 06:00 [pubmed]
PHST- 2018/11/15 06:01 [medline]
PHST- 2018/05/30 00:00 [pmc-release]
AID - mi9060272 [pii]
AID - micromachines-09-00272 [pii]
AID - 10.3390/mi9060272 [doi]
PST - epublish
SO  - Micromachines (Basel). 2018 May 30;9(6):272. doi: 10.3390/mi9060272.