Detection of circulating tumor cells in blood using an optimized density gradient centrifugation.
The aim of the study was to compare the new density gradient centrifugation system OncoQuick with the standard density gradient centrifugation system Ficoll for improved tumor cell enrichment in blood of tumor patients. Evaluation of OncoQuick and Ficoll density gradient centrifugation was performed by flow-cytometry and immunocytochemistry using 10 ml unspiked and tumor cell-spiked blood samples of tumor-free probands. From 10 ml blood, OncoQuick density gradient centrifugation separated a cell fraction which consisted of a mean cell number of 9.5x10(4) mononuclear cells compared to 1.8x10(7) cells by Ficoll. Density gradient centrifugation of tumor cell-spiked blood samples with OncoQuick and Ficoll led to similar tumor cell recovery rates, between 70% and 90% for both methods. The improved depletion of mononuclear blood cells by OncoQuick simplified further immunocytochemical evaluation of the enriched cell fraction, which could be spun onto 1-2 glass slides by cytocentrifugation. In comparison, the mononuclear cells separated by Ficoll had to be spun onto more than 50 glass slides for complete immunocytochemical evaluation. Consequently, tumor cell density on each cytospin was higher after OncoQuick preparation compared to Ficoll. Density gradient centrifugation with OncoQuick results in higher relative tumor cell enrichment than Ficoll density gradient centrifugation. This simplifies further immunocytochemical tumor cell detection and is a promising tool for the detection of circulating tumor cells in blood of tumor patients.
Gertler
Ralf
R
Chirurgische Klinik und Poliklinik, Klinikum rechts der Isar der Technischen Universität München, 81675 Munich, Germany.
Rosenberg
Robert
R
Fuehrer
Katrin
K
Dahm
Michael
M
Nekarda
Hjalmar
H
Siewert
Joerg Ruediger
JR
D002499:Centrifugation, Density Gradient / Q000379:methods*; D005434:Flow Cytometry; D006801:Humans; D007150:Immunohistochemistry; D007963:Leukocytes, Mononuclear / Q000378:metabolism; D009360:Neoplastic Cells, Circulating / Q000378:metabolism*; D014407:Tumor Cells, Cultured