Cancer Diagnosis Program | Biorepositories and Biospecimen Research Branch

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Comparison of two methods for enumerating circulating tumor cells in carcinoma patients.

Author(s): Balic M, Dandachi N, Hofmann G, Samonigg H, Loibner H, Obwaller A, van der Kooi A, Tibbe AG, Doyle GV, Terstappen LW, Bauernhofer T

Publication: Cytometry B Clin Cytom, 2005, Vol. 68, Page 25-30

PubMed ID: 16142788 PubMed Review Paper? No

Purpose of Paper

Conclusion of Paper

Studies

1. Study Purpose

   This study compared two methods for the enumeration of CTCs in peripheral blood collected from patients diagnosed with a carcinoma and healthy volunteers. A total of 22.5 ml of blood was collected in CellSave tubes from each of 15 healthy donors and 61 patients with a newly diagnosed carcinoma (14 breast, 11 colon, 6 rectal, 6 gastric, 5 ovarian, 5 prostate, 2 liver, 2 esophageal, 2 lung, 2 testicular, 2 pancreatic, 2 carcinomas of unknown primary, 1 kidney, and 1 uterine) of which 53 (87%) had metastatic disease that had not been treated with chemotherapy, radiation, or hormonal therapy for at least 6 weeks and eight (13%) were undergoing adjuvant therapy. Blood was pooled for each person then aliquoted such that 15 ml was analyzed by the OncoQuick assay and 7.5 ml was analyzed by the CellSearch assay the day following collection. Blood specimens analyzed by OncoQuick, which is based on density gradient centrifugation, were centrifuged for 20 min at 1600 x g at 4°C, interphase cells were washed, and then centrifuged at 200 x g for 10 min at 4°C prior to incubation with a fluorescently-labeled anti-cytokeratin (CK) antibody A45-B/B3 (recognizes CK8, CK18, and CK19) and visualization with an automated fluorescence image analysis system. Blood specimens analyzed by CellSearch, which utilizes antibody-labeled ferrofluid nanoparticles for magnetic separation, were centrifuged at 800 x g for 10 min prior to analysis with a CellPrep semi-automated instrument, incubation with anti-EpCAM antibody-labeled ferrofluids and visualization with a semi-automated four-color fluorescence microscope.

   Summary of Findings:

   Peripheral blood specimens were considered positive for CTCs if at least one cell was detected. No CTCs were detected in blood from healthy donors by either the CellSave or OncoQuick method. CTCs were detected in 14/61 (23%) of peripheral blood specimens from carcinoma patients using OncoQuick and 33/61 (54%) specimens using CellSearch (P<0001). Eleven of the 14 specimens that were positive with OncoQuick were also positive by CellSearch, but 22 of the 33 specimens that were positive with CellSearch were negative with OncoQuick (P<0.0001). CellSearch detected significantly higher mean numbers of CTCs in blood from carcinoma patients than OncoQuick (20 vs. 3, P<0001) and higher percentages of blood specimens positive for CTCs than OncoQuick (31% vs. 15% with one to four CTCs and 23% vs. 8% with at least five CTCs). Further, a higher percentage of blood specimens collected from carcinoma patients tested negative for CTCs when determined by OncoQuick than with CellSearch (77% vs. 46%). More CTCs were detected by CellSearch than by OncoQuick in the 53 patients with metastatic carcinoma (60% vs. 26%). Of the eight nonmetastatic patients, CTCs were detected in one specimen (12%) by each method, although it was not the same patient. A higher percentage of blood specimens were positive for CTCs when detected by Cell Search versus OncoQuick for patients with breast cancer (71% vs. 23%), colon cancer (64% vs. 45%), stomach cancer (33% vs. 0%), rectal cancer (66% vs. 33%), ovarian cancer (60% vs. 40%), and prostate cancer (20% vs.  0%).

   Biospecimens

   • Bodily Fluid - Blood

   Preservative Types

   • None (Fresh)

   Diagnoses:

   • Neoplastic - Carcinoma
   • Normal

   Platform:

   Analyte	Technology Platform
   Cell count/volume	H-and-E microscopy
   Cell count/volume	Magnetic-activated cell sorting

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   Biospecimen Aliquots and Components	Cell number	OncoQuick CellSearch
   Preaquisition	Duration of anesthesia	Metastatic disease Nonmetastatic Healthy Breast cancer Colon cancer Rectal cancer Gastric cancer Ovarian cancer Prostate cancer Liver cancer Lung cancer Testicular cancer Pancreatic cancer Carcinoma of unknown primary Kidney carcinoma Uterine carcinoma

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