Influence of Blood Storage and Sample Processing on Molecular Detection of Circulating Prostate Cells in Cancer

Abstract

Although not in routine use, molecular techniques appear able to detect small numbers of solid tumor cells in blood, lymph nodes, and bone marrow (for review see 1 ). Provided that a given mRNA is expressed exclusively in tumor tissue and not by nucleated blood cells, lymph node cells, or bone marrow cells, its presence outside the organ identifies disseminated metastatic cells. In prostate cancer (CaP), early detection of metastases by molecular staging could help in selecting which patients would benefit most from surgical therapy (2). Several reverse transcriptase-polymerase chain reaction (RT-PCR) protocols are able to amplify the mRNAs encoding prostate-specific antigen (PSA) or prostate-specific membrane antigen (PSMA) in blood (1), lymph nodes (3), or bone marrow (4). Nevertheless, the results reported have varied so widely (e.g., positive PSA RT-PCR results ranging from 25% (5) to 80% (6) for metastatic CaP patients) that definitive conclusions about the clinical relevance of this new molecular tool have not yet been established. The heterogeneity of both preanalytical and analytical steps of PCR protocols may explain the discrepancies observed between studies. To our knowledge, no systematic studies of the successive steps of the RT-PCR method have yet been performed. To better standardize the assay, we have assessed the effects of the storage conditions of blood samples and delay before processing. To compare the effect of delayed vs immediate sample processing, we collected 250-mL blood samples in 25 10-mL EDTA-treated tubes (Becton Dickinson) from 4 …