PMID- 12565993
OWN - NLM
STAT- MEDLINE
DCOM- 20030428
LR  - 20190718
IS  - 0959-8049 (Print)
IS  - 0959-8049 (Linking)
VI  - 39
IP  - 3
DP  - 2003 Feb
TI  - Reliability of quantitative reverse-transcriptase-PCR-based detection of tumour 
      cells in the blood between different laboratories using a standardised protocol.
PG  - 388-96
AB  - Differences in methods of reverse-transcriptase (RT)-polymerase chain reaction 
      (PCR)-based detection of tumour cells in the blood gives rise to conflicting 
      results, and standardisation is urgently needed. This pilot study aimed to assess 
      the variation of RT-PCR-based detection of tumour cells in blood between four 
      different laboratories using a commercially available kit with a standardised 
      protocol. This kit allows comparison of results from different laboratories and 
      facilitates the investigation of the influence of pre-analytical parameters. All 
      laboratories analysed identical sets of blood samples spiked with tumour cells in 
      a concentration range of 1-100 tumour cells/ml. To study at which level variation 
      was introduced, three kinds of sample sets were generated in which (i) tumour 
      cell RNA was spiked in the RNA of mononuclear cells (MNC), (ii) tumour cells were 
      spiked in isolated MNC, and (iii) tumour cells were spiked in blood. Real-time 
      quantitative RT-PCR was used to detect and quantify cytokeratin 20 (CK20) 
      expression, which is indicative for the presence of epithelial tumour cells. All 
      laboratories were able to detect CK20 expression in all spiked-RNA samples with 
      limited variation in expression levels between laboratories. There was a positive 
      correlation between the amount of spiked tumour cell RNA and CK20 expression 
      level. RT-PCR analysis of spiked-MNC samples resulted in more variation in the 
      CK20 expression levels between laboratories, however again all spiked samples 
      were reported to be positive by all of the laboratories. The evaluation of 
      spiked-blood samples gave rise to considerable quantitative and qualitative 
      variation between the laboratories. Our results underline the importance and need 
      for standardisation and extended quality control studies in the field of 
      pre-analytics.
FAU - Vlems, F A
AU  - Vlems FA
AD  - Department of Surgery, University Medical Centre Nijmegen, PO-box 9101, 6500HB, 
      Nijmegen, The Netherlands. f.vlems@pathol.azn.nl
FAU - Ladanyi, A
AU  - Ladanyi A
FAU - Gertler, R
AU  - Gertler R
FAU - Rosenberg, R
AU  - Rosenberg R
FAU - Diepstra, J H S
AU  - Diepstra JH
FAU - Roder, C
AU  - Roder C
FAU - Nekarda, H
AU  - Nekarda H
FAU - Molnar, B
AU  - Molnar B
FAU - Tulassay, Z
AU  - Tulassay Z
FAU - van Muijen, G N P
AU  - van Muijen GN
FAU - Vogel, I
AU  - Vogel I
LA  - eng
PT  - Comparative Study
PT  - Journal Article
PL  - England
TA  - Eur J Cancer
JT  - European journal of cancer (Oxford, England : 1990)
JID - 9005373
RN  - 0 (Intermediate Filament Proteins)
RN  - 0 (KRT20 protein, human)
RN  - 0 (Keratin-20)
RN  - 0 (RNA, Neoplasm)
SB  - IM
MH  - HT29 Cells
MH  - Humans
MH  - Intermediate Filament Proteins/metabolism
MH  - Keratin-20
MH  - Laboratories/*standards
MH  - Monocytes
MH  - Neoplasms/*diagnosis
MH  - *Neoplastic Cells, Circulating
MH  - Pilot Projects
MH  - Quality Control
MH  - RNA, Neoplasm/analysis
MH  - Reproducibility of Results
MH  - Reverse Transcriptase Polymerase Chain Reaction/*standards
MH  - Sensitivity and Specificity
EDAT- 2003/02/05 04:00
MHDA- 2003/04/29 05:00
CRDT- 2003/02/05 04:00
PHST- 2003/02/05 04:00 [pubmed]
PHST- 2003/04/29 05:00 [medline]
PHST- 2003/02/05 04:00 [entrez]
AID - S0959804902006317 [pii]
AID - 10.1016/s0959-8049(02)00631-7 [doi]
PST - ppublish
SO  - Eur J Cancer. 2003 Feb;39(3):388-96. doi: 10.1016/s0959-8049(02)00631-7.