The effect of centrifugation on circulating mRNA quantitation opens up a new scenario in expression profiling from patients with metastatic colorectal cancer.
OBJECTIVE
To investigate the effect of centrifugation in circulating mRNA quantitation and to design a new, consistent protocol for it.


METHODS
The concentrations of beta-catenin mRNA, P-selectin mRNA, cytokeratin 20 mRNA, and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) mRNA in the plasma, the mononuclear and red cells (MARC) from metastatic colorectal (CRC) patients whose blood samples were centrifuged with 2 different forces, were measured. A new protocol to measure expression profiles in MARC was studied.


RESULTS
When the centrifugal force was increased: (1) the concentrations of all transcripts were dramatically decreased in the plasma samples; (2) only the concentrations of P-selectin mRNA and GAPDH mRNA were significantly increased in the mononuclear cell samples; and (3) their concentrations in the red cells did not change significantly. The new protocol found 9 differential expressed genes.


CONCLUSIONS
Centrifugal force causes inconsistent mRNA quantitation in the plasma. The new protocol discovered 9 differential expressed genes for metastatic CRC.
Wong
S C Cesar
SC
State Key Laboratory in Oncology in South China, Sir Y K Pao Centre for Cancer, Department of Clinical Oncology, Hong Kong Cancer Institute and Prince of Wales Hospital, The Chinese University of Hong Kong. cesar01@netvigator.com
Ma
Brigette B Y
BB
Lai
Paul B S
PB
Ng
Simon S M
SS
Lee
Janet F Y
JF
Hui
Edwin P
EP
Lam
Money Y Y
MY
Chan
Charles M L
CM
Chan
Anthony T C
AT
D002498:Centrifugation / Q000379:methods*; D015179:Colorectal Neoplasms / Q000097:blood* / Q000235:genetics* / Q000473:pathology; D020869:Gene Expression Profiling; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000235:genetics; D006801:Humans; D053554:Keratin-20 / Q000235:genetics; D009362:Neoplasm Metastasis; D019007:P-Selectin / Q000235:genetics; D012333:RNA, Messenger / Q000097:blood*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D051176:beta Catenin / Q000235:genetics
D053554:Keratin-20; D019007:P-Selectin; D012333:RNA, Messenger; D051176:beta Catenin; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases