Detection of circulating cytokeratin-positive cells in the blood of breast cancer patients using immunomagnetic enrichment and digital microscopy

Thomas Elmer Witzig, Blaise Bossy, Teresa Kimlinger, Patrick C. Roche, James N. Ingle, Clive Grant, John Donohue, Vera Jean Suman, Douglas Harrington, Jose Torre-Bueno, Kenneth D. Bauer

Research output: Contribution to journalArticle

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Abstract

Purpose: To examine the feasibility for identifying and enumerating cytokeratin positive (CK+) cells in the peripheral blood of breast cancer patients. Experimental Design: Blood specimens from 34 normal donors (negative controls), 15 samples to which carcinoma cells were added (positive controls), and 84 breast cancer patients [27 node-negative (N-), 29 node-positive (N+), and 28 metastatic] were studied. RBCs were lysed with ammonium chloride and the resulting cell suspension incubated with anti-EpCAM-conjugated immunomagnetic beads for carcinoma cell enrichment. Immunomagnetically selected cells were placed on slides; stained for CKs 8, 18, and 19; and evaluated with an automated digital microscopy system that rapidly scanned the slide and collected images of cells meeting predefined staining and cytomorphological criteria. A montage of the CK+ cells was reviewed to confirm tumor cell morphology. Results: Eighteen specimens (9 normal, 2 N-, 4 N+, and 3 metastatic) were excluded because of poor cytomorphology or staining artifact. All 15 of the positive controls [95% confidence interval (CI), 78-100%] and none of the 25 negative controls (95% CI, 0-14%) demonstrated CK+ cells. Twenty-one of the 75 (28%; 95% CI, 18-40%) samples from breast cancer patients demonstrated CK+ cells including 76% of patients with metastatic disease (95% CI, 55-91%), 8% with N+ disease (95% CI, 1-26%), and none of those with N- disease (95% CI, 0-14). The mean number of CK+ cells detected in the 21 CK+ patients was 18.4 (range, 1-120). Conclusions: Breast carcinoma cells can be detected in the blood from a significant fraction of metastatic breast cancer patients using immunomagnetic cell enrichment and digital microscopy. The incidence of CK+ cells was low in those with resected N+ disease (at most 26%) and those with resected N- breast cancer (at most 14%). This technique could be used in large prospective studies of patients with breast cancer to learn whether the detection of rare carcinoma cells is a useful predictive or prognostic factor.

Original languageEnglish (US)
Pages (from-to)1085-1091
Number of pages7
JournalClinical Cancer Research
Volume8
Issue number5
StatePublished - 2002

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Keratins
Microscopy
Blood Cells
Breast Neoplasms
Confidence Intervals
Carcinoma
Staining and Labeling
Ammonium Chloride
Artifacts
Suspensions
Research Design
Cell Count
Tissue Donors

ASJC Scopus subject areas

  • Cancer Research
  • Oncology

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Detection of circulating cytokeratin-positive cells in the blood of breast cancer patients using immunomagnetic enrichment and digital microscopy. / Witzig, Thomas Elmer; Bossy, Blaise; Kimlinger, Teresa; Roche, Patrick C.; Ingle, James N.; Grant, Clive; Donohue, John; Suman, Vera Jean; Harrington, Douglas; Torre-Bueno, Jose; Bauer, Kenneth D.

In: Clinical Cancer Research, Vol. 8, No. 5, 2002, p. 1085-1091.

Research output: Contribution to journalArticle

Witzig, TE, Bossy, B, Kimlinger, T, Roche, PC, Ingle, JN, Grant, C, Donohue, J, Suman, VJ, Harrington, D, Torre-Bueno, J & Bauer, KD 2002, 'Detection of circulating cytokeratin-positive cells in the blood of breast cancer patients using immunomagnetic enrichment and digital microscopy', Clinical Cancer Research, vol. 8, no. 5, pp. 1085-1091.
Witzig, Thomas Elmer ; Bossy, Blaise ; Kimlinger, Teresa ; Roche, Patrick C. ; Ingle, James N. ; Grant, Clive ; Donohue, John ; Suman, Vera Jean ; Harrington, Douglas ; Torre-Bueno, Jose ; Bauer, Kenneth D. / Detection of circulating cytokeratin-positive cells in the blood of breast cancer patients using immunomagnetic enrichment and digital microscopy. In: Clinical Cancer Research. 2002 ; Vol. 8, No. 5. pp. 1085-1091.
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title = "Detection of circulating cytokeratin-positive cells in the blood of breast cancer patients using immunomagnetic enrichment and digital microscopy",
abstract = "Purpose: To examine the feasibility for identifying and enumerating cytokeratin positive (CK+) cells in the peripheral blood of breast cancer patients. Experimental Design: Blood specimens from 34 normal donors (negative controls), 15 samples to which carcinoma cells were added (positive controls), and 84 breast cancer patients [27 node-negative (N-), 29 node-positive (N+), and 28 metastatic] were studied. RBCs were lysed with ammonium chloride and the resulting cell suspension incubated with anti-EpCAM-conjugated immunomagnetic beads for carcinoma cell enrichment. Immunomagnetically selected cells were placed on slides; stained for CKs 8, 18, and 19; and evaluated with an automated digital microscopy system that rapidly scanned the slide and collected images of cells meeting predefined staining and cytomorphological criteria. A montage of the CK+ cells was reviewed to confirm tumor cell morphology. Results: Eighteen specimens (9 normal, 2 N-, 4 N+, and 3 metastatic) were excluded because of poor cytomorphology or staining artifact. All 15 of the positive controls [95{\%} confidence interval (CI), 78-100{\%}] and none of the 25 negative controls (95{\%} CI, 0-14{\%}) demonstrated CK+ cells. Twenty-one of the 75 (28{\%}; 95{\%} CI, 18-40{\%}) samples from breast cancer patients demonstrated CK+ cells including 76{\%} of patients with metastatic disease (95{\%} CI, 55-91{\%}), 8{\%} with N+ disease (95{\%} CI, 1-26{\%}), and none of those with N- disease (95{\%} CI, 0-14). The mean number of CK+ cells detected in the 21 CK+ patients was 18.4 (range, 1-120). Conclusions: Breast carcinoma cells can be detected in the blood from a significant fraction of metastatic breast cancer patients using immunomagnetic cell enrichment and digital microscopy. The incidence of CK+ cells was low in those with resected N+ disease (at most 26{\%}) and those with resected N- breast cancer (at most 14{\%}). This technique could be used in large prospective studies of patients with breast cancer to learn whether the detection of rare carcinoma cells is a useful predictive or prognostic factor.",
author = "Witzig, {Thomas Elmer} and Blaise Bossy and Teresa Kimlinger and Roche, {Patrick C.} and Ingle, {James N.} and Clive Grant and John Donohue and Suman, {Vera Jean} and Douglas Harrington and Jose Torre-Bueno and Bauer, {Kenneth D.}",
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T1 - Detection of circulating cytokeratin-positive cells in the blood of breast cancer patients using immunomagnetic enrichment and digital microscopy

AU - Witzig, Thomas Elmer

AU - Bossy, Blaise

AU - Kimlinger, Teresa

AU - Roche, Patrick C.

AU - Ingle, James N.

AU - Grant, Clive

AU - Donohue, John

AU - Suman, Vera Jean

AU - Harrington, Douglas

AU - Torre-Bueno, Jose

AU - Bauer, Kenneth D.

PY - 2002

Y1 - 2002

N2 - Purpose: To examine the feasibility for identifying and enumerating cytokeratin positive (CK+) cells in the peripheral blood of breast cancer patients. Experimental Design: Blood specimens from 34 normal donors (negative controls), 15 samples to which carcinoma cells were added (positive controls), and 84 breast cancer patients [27 node-negative (N-), 29 node-positive (N+), and 28 metastatic] were studied. RBCs were lysed with ammonium chloride and the resulting cell suspension incubated with anti-EpCAM-conjugated immunomagnetic beads for carcinoma cell enrichment. Immunomagnetically selected cells were placed on slides; stained for CKs 8, 18, and 19; and evaluated with an automated digital microscopy system that rapidly scanned the slide and collected images of cells meeting predefined staining and cytomorphological criteria. A montage of the CK+ cells was reviewed to confirm tumor cell morphology. Results: Eighteen specimens (9 normal, 2 N-, 4 N+, and 3 metastatic) were excluded because of poor cytomorphology or staining artifact. All 15 of the positive controls [95% confidence interval (CI), 78-100%] and none of the 25 negative controls (95% CI, 0-14%) demonstrated CK+ cells. Twenty-one of the 75 (28%; 95% CI, 18-40%) samples from breast cancer patients demonstrated CK+ cells including 76% of patients with metastatic disease (95% CI, 55-91%), 8% with N+ disease (95% CI, 1-26%), and none of those with N- disease (95% CI, 0-14). The mean number of CK+ cells detected in the 21 CK+ patients was 18.4 (range, 1-120). Conclusions: Breast carcinoma cells can be detected in the blood from a significant fraction of metastatic breast cancer patients using immunomagnetic cell enrichment and digital microscopy. The incidence of CK+ cells was low in those with resected N+ disease (at most 26%) and those with resected N- breast cancer (at most 14%). This technique could be used in large prospective studies of patients with breast cancer to learn whether the detection of rare carcinoma cells is a useful predictive or prognostic factor.

AB - Purpose: To examine the feasibility for identifying and enumerating cytokeratin positive (CK+) cells in the peripheral blood of breast cancer patients. Experimental Design: Blood specimens from 34 normal donors (negative controls), 15 samples to which carcinoma cells were added (positive controls), and 84 breast cancer patients [27 node-negative (N-), 29 node-positive (N+), and 28 metastatic] were studied. RBCs were lysed with ammonium chloride and the resulting cell suspension incubated with anti-EpCAM-conjugated immunomagnetic beads for carcinoma cell enrichment. Immunomagnetically selected cells were placed on slides; stained for CKs 8, 18, and 19; and evaluated with an automated digital microscopy system that rapidly scanned the slide and collected images of cells meeting predefined staining and cytomorphological criteria. A montage of the CK+ cells was reviewed to confirm tumor cell morphology. Results: Eighteen specimens (9 normal, 2 N-, 4 N+, and 3 metastatic) were excluded because of poor cytomorphology or staining artifact. All 15 of the positive controls [95% confidence interval (CI), 78-100%] and none of the 25 negative controls (95% CI, 0-14%) demonstrated CK+ cells. Twenty-one of the 75 (28%; 95% CI, 18-40%) samples from breast cancer patients demonstrated CK+ cells including 76% of patients with metastatic disease (95% CI, 55-91%), 8% with N+ disease (95% CI, 1-26%), and none of those with N- disease (95% CI, 0-14). The mean number of CK+ cells detected in the 21 CK+ patients was 18.4 (range, 1-120). Conclusions: Breast carcinoma cells can be detected in the blood from a significant fraction of metastatic breast cancer patients using immunomagnetic cell enrichment and digital microscopy. The incidence of CK+ cells was low in those with resected N+ disease (at most 26%) and those with resected N- breast cancer (at most 14%). This technique could be used in large prospective studies of patients with breast cancer to learn whether the detection of rare carcinoma cells is a useful predictive or prognostic factor.

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