PMID- 23711823
OWN - NLM
STAT- MEDLINE
DCOM- 20131018
LR  - 20230216
IS  - 1530-0307 (Electronic)
IS  - 0023-6837 (Linking)
VI  - 93
IP  - 8
DP  - 2013 Aug
TI  - Integrated RNA extraction and RT-PCR for semi-quantitative gene expression 
      studies on a microfluidic device.
PG  - 961-6
LID - 10.1038/labinvest.2013.76 [doi]
AB  - This paper describes the development of a microfluidic methodology, using RNA 
      extraction and reverse transcription PCR, for investigating expression levels of 
      cytochrome P450 genes. Cytochrome P450 enzymes are involved in the metabolism of 
      xenobiotics, including many commonly prescribed drugs, therefore information on 
      their expression is useful in both pharmaceutical and clinical settings. RNA 
      extraction, from rat liver tissue or primary rat hepatocytes, was performed using 
      a silica-based solid-phase extraction technique. Following elution of the 
      purified RNA, amplification of target sequences for the housekeeping gene, 
      glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and the cytochrome P450 gene 
      CYP1A2, was carried out using a one-step reverse transcription PCR. Once the 
      microfluidic methodology had been optimized, analysis of control and 
      3-methylcholanthrene-induced primary rat hepatocytes were used to evaluate the 
      system. As expected, GAPDH was consistently expressed, whereas CYP1A2 levels were 
      found to be raised in the drug-treated samples. The proposed system offers an 
      initial platform for development of both rapid throughput analyzers for 
      pharmaceutical drug screening and point-of-care diagnostic tests to aid provision 
      of drug regimens, which can be tailor-made to the individual patient.
FAU - Shaw, Kirsty J
AU  - Shaw KJ
AD  - School of Science and the Environment, Manchester Metropolitan University, 
      Manchester, UK. k.shaw@mmu.ac.uk
FAU - Hughes, Elizabeth M
AU  - Hughes EM
FAU - Dyer, Charlotte E
AU  - Dyer CE
FAU - Greenman, John
AU  - Greenman J
FAU - Haswell, Stephen J
AU  - Haswell SJ
LA  - eng
PT  - Journal Article
DEP - 20130527
PL  - United States
TA  - Lab Invest
JT  - Laboratory investigation; a journal of technical methods and pathology
JID - 0376617
RN  - 63231-63-0 (RNA)
RN  - EC 1.14.14.1 (Cytochrome P-450 CYP1A2)
RN  - EC 1.2.1.- (Glyceraldehyde-3-Phosphate Dehydrogenases)
SB  - IM
MH  - Animals
MH  - Cytochrome P-450 CYP1A2/genetics
MH  - Gene Expression
MH  - Gene Expression Regulation, Enzymologic
MH  - Glyceraldehyde-3-Phosphate Dehydrogenases/genetics
MH  - Hepatocytes/chemistry
MH  - Liver/chemistry
MH  - Male
MH  - Microfluidic Analytical Techniques/instrumentation/*methods
MH  - RNA/*isolation & purification
MH  - Rats
MH  - Rats, Wistar
MH  - Reverse Transcriptase Polymerase Chain Reaction/*methods
EDAT- 2013/05/29 06:00
MHDA- 2013/10/19 06:00
CRDT- 2013/05/29 06:00
PHST- 2013/04/08 00:00 [received]
PHST- 2013/05/06 00:00 [revised]
PHST- 2013/05/07 00:00 [accepted]
PHST- 2013/05/29 06:00 [entrez]
PHST- 2013/05/29 06:00 [pubmed]
PHST- 2013/10/19 06:00 [medline]
AID - S0023-6837(22)00917-5 [pii]
AID - 10.1038/labinvest.2013.76 [doi]
PST - ppublish
SO  - Lab Invest. 2013 Aug;93(8):961-6. doi: 10.1038/labinvest.2013.76. Epub 2013 May 
      27.