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Breast Cancer Res. 2008;10(4):R69. doi: 10.1186/bcr2131. Epub 2008 Aug 7.

Enrichment with anti-cytokeratin alone or combined with anti-EpCAM antibodies significantly increases the sensitivity for circulating tumor cell detection in metastatic breast cancer patients.

Author information

1
Biology, Genetix USA Inc, 120 Baytech Drive, San Jose, CA 95134, USA. glenn.deng@genetix.com

Abstract

INTRODUCTION:

Circulating tumor cells (CTCs) are detectable in most cancer patients and they can meet an existing medical need to monitor cancer patients during a course of treatment and to help determine recurrent disease. CTCs are rarely found in the blood of cancer patients and enrichment is necessary for sensitive CTC detection. Most CTC enrichment technologies are anti-EpCAM antibody based even though CTC identification criteria are cytokeratin positive (CK+), CD45 negative (CD45-) and 4'6-diamidino-2-phenylindole (nuclear stain) positive (DAPI+). However, some tumor cells express low or no EpCAM. Here we present a highly sensitive and reproducible enrichment method that is based on binding to anti-CK alone or a combination of anti-CK and anti-EpCAM antibodies.

METHODS:

Blood samples from 49 patients with metastatic breast cancer were processed using the CellSearchtrade mark system (Veridex, LLC, Raritan, NJ, USA), in parallel with our CTC assay method. We used anti-CK alone or in combination with anti-EpCAM antibodies for CTC enrichment. Brightfield and fluorescence labeled anti-CK, anti-CD45 and DAPI (nuclear stain) images were used for CTC identification. The Ariol(R) system (Genetix USA Inc, San Jose, CA, USA) was used for automated cell image capture and analysis of CTCs on glass slides.

RESULTS:

Our method has the capability to enrich three types of CTCs including CK+&EpCAM+, CK+&EpCAM-/low, and CK-/low&EpCAM+ cells. In the blind method comparison, our anti-CK antibody enrichment method showed a significantly higher CTC positive rate (49% vs. 29%) and a larger dynamic CTC detected range (1 to 571 vs. 1 to 270) than that of the CellSearchtrade mark system in the total of 49 breast cancer patients. Our method detected 15 to 111% more CTCs than the CellSearchtrade mark method in patients with higher CTC counts (>20 CTCs per 7.5 ml of blood). The three fluorescent and brightfield images from the Ariol(R) system reduced the number of false-positive CTC events according to the established CTC criteria.

CONCLUSION:

Our data indicate that the tumor-specific intracellular CK marker could be used for efficient CTC enrichment. Enrichment with anti-CK alone or combined with anti-EpCAM antibodies significantly enhances assay sensitivity. The three fluorescent and brightfield superior images with the Ariol(R) system reduced false-positive CTC events.

PMID:
18687126
PMCID:
PMC2575542
DOI:
10.1186/bcr2131
[Indexed for MEDLINE]
Free PMC Article
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