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Enrichment Of Rare Cancer Cells Through Depletion Of Normal Cells Using Density And Flow-through, Immunomagnetic Cell Separation.

Oscar R. Lara, Xiaodong Tong, Maciej Zborowski, J. Chalmers
Published 2004 · Biology, Medicine

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OBJECTIVE To develop a reliable technique to enrich for rare cells in blood suspensions using only negative selection steps including a flow-through immunomagnetic cell separations system and by optimizing variables normally encountered during such enrichment processes. METHODS A human breast cancer cell line was cultivated and spiked at a ratio of 1 cancer cell to 10(5) total leukocytes in buffy coat or 1 cancer cell to 10(8) total cells in whole blood samples. The final, optimized process consisted of: a red cell lysis step, immunomagnetically staining leukocytes with an anti-CD45 PE, anti- MACS sandwich, immunomagnetic sorting using a flow-through system (QMS), and a final cell analysis step using either an automated cell counter, filtration, and visual counting or a cytospin analysis. RESULTS The final, optimized process produced a final enrichment of the rare cancer cells of 5.17 log(10) and an average, final recovery of 46%. It should be noted that a negative depletion protocol was used (i.e., no labeling of the rare cancer cells was used). CONCLUSIONS To the authors' knowledge, no examples in the literature exist of a 5.17 log(10) enrichment of cancer cells in human blood using a negative depletion protocol. The closest example is a 4 log(10) enrichment in which two positive magnetic cell separation steps were used (none were used in this study). Ongoing studies are investigating further modifications of the precommercial, prototype flow-through immunmagnetic separation system to increase both the enrichment and recovery rate. However, even at current performance levels, the presented process could significantly improve visual and molecular analysis of rare cells in blood.
This paper references
Detection of rare circulating breast cancer cells by filtration cytometry and identification by DNA content: sensitivity in an experimental model.
P. Rostagno (1997)
10.1056/NEJME048163
Circulating tumor cells in metastatic breast cancer--toward individualized treatment?
Stephan Braun (2004)
10.1046/j.1537-2995.2000.40121489.x
Immunomagnetic tumor cell selection—implications for the detection of disseminated cancer cells
W. Krüger (2000)
Molecular detection of breast cancer cells in the peripheral blood of advanced-stage breast cancer patients using multimarker real-time reverse transcription-polymerase chain reaction and a novel porous barrier density gradient centrifugation technology.
Megan K. Baker (2003)
10.1634/THEONCOLOGIST.6-2-125
Clinical significance of occult metastatic cells in bone marrow of breast cancer patients.
S. Braun (2001)
Circulating tumor cell clusters in the peripheral blood of colorectal cancer patients.
B. Molnár (2001)
10.1080/0032472031000141282
Use of a tumor-cell enrichment column for the enhanced detection of minimal residual disease in the BM or apheresis peripheral blood transplant products of breast-cancer patients.
J. Shammo (1999)
10.1002/(SICI)1097-0215(20000120)89:1<8::AID-IJC2>3.0.CO;2-K
Molecular detection of blood‐borne epithelial cells in colorectal cancer patients and in patients with benign bowel disease
J. Hardingham (2000)
10.1016/S0021-9673(00)00879-7
Study of magnetic particles pulse-injected into an annular SPLITT-like channel inside a quadrupole magnetic field.
M. Hoyos (2000)
An instrument to determine the magnetophoretic mobility of paramagnetic particles and labeled, biological cells
JJ Chalmers (1999)
Magnetic cell separation is a function of antibody binding capacity (ABC)
K McCloskey (2003)
10.1016/S0002-9440(10)64706-2
Isolation by size of epithelial tumor cells : a new method for the immunomorphological and molecular characterization of circulatingtumor cells.
G. Vona (2000)
Changes in circulating carcinoma cells in patients with metastasis prostate cancer correlate with disease
JG Moreno (2001)
10.1016/S0002-9610(00)00518-3
Detection of carcinoma cells in the blood of breast cancer patients.
P. Beitsch (2000)
10.1081/SS-120000319
Visualizing patterns of protein uptake to porous media using confocal scanning laser microscopy
T. Linden (2002)
10.1002/(SICI)1097-0320(19990801)36:4<294::AID-CYTO3>3.0.CO;2-C
Detection of rare MCF-7 breast carcinoma cells from mixtures of human peripheral leukocytes by magnetic deposition analysis.
B. Fang (1999)
Antibody saturation studies of three antigens proposed for use in rare cancer cell detection
J Chosy (2003)
Continuous, flow-through immunomagnetic cell separation in a quadrupole field. Cytometry
L Sun (1998)
10.1002/(SICI)1096-9896(199911)189:3<368::AID-PATH441>3.0.CO;2-2
Immunomagnetic separation for enrichment and sensitive detection of disseminated tumour cells in patients with head and neck SCC
M. Partridge (1999)
Detection of occult metastatic breast cancer cells in blood by a multimolecular marker assay: correlation with clinical stage of disease.
B. Taback (2001)
10.1081/SS-120014809
SEPARATIONS BASED ON MAGNETOPHORETIC MOBILITY
Maciej Zborowski (2002)
10.1080/01496398508060702
A System Based on Split-Flow Lateral-Transport Thin (SPLITT) Separation Cells for Rapid and Continuous Particle Fractionation
J. Giddings (1985)
Detection of rare MCF7 breast cancer cells from mixtures of peripheral blood leucocytes by magnetic deposition analysis
B Fang (1999)
10.1021/bp010109q
Separation of a Breast Cancer Cell Line from Human Blood Using a Quadrupole Magnetic Flow Sorter
M. Nakamura (2001)
10.1021/bp020285e
Magnetophoretic Cell Sorting Is a Function of Antibody Binding Capacity
K. McCloskey (2003)
10.1007/s004320100239
Immunobead RT-PCR versus regular RT-PCR amplification of CEA mRNA in peripheral blood
Seung-Yoon Park (2001)
10.1002/1097-0320(20011201)45:4<285::AID-CYTO10018>3.0.CO;2-W
Effects of antibody concentration on the separation of human natural killer cells in a commercial immunomagnetic separation system.
K. Comella (2001)
10.1016/S0304-8853(00)01248-8
Comparison of different particles and methods for magnetic isolation of circulating tumor cells
S. Sieben (2001)
10.1073/PNAS.95.8.4589
Detection and characterization of carcinoma cells in the blood.
E. Racila (1998)
10.1002/(SICI)1098-2825(1997)11:6<346::AID-JCLA7>3.0.CO;2-3
Application of immunomagnetic beads in combination with RT‐PCR for the detection of circulating prostate cancer cells
A. Makarovskiy (1997)
10.1007/s004320050035
Sensitive detection of micrometastases in bone marrow from patients with breast cancer using immunomagnetic isolation of tumor cells in combination with reverse transcriptase/polymerase chain reaction for cytokeratin-19
X. Zhong (2000)
10.1021/AC034315J
Magnetic cell separation: characterization of magnetophoretic mobility.
K. McCloskey (2003)
10.1002/(SICI)1098-2825(1999)13:5<229::AID-JCLA7>3.0.CO;2-Y
Detection of circulating melanoma cells by immunomagnetic cell sorting
A. Benez (1999)
10.1016/S0304-8853(00)01251-8
Progenitor cell isolation with a high-capacity quadrupole magnetic flow sorter
L. Moore (2001)
10.1006/GYNO.2000.5781
Detection of micrometastasis by cytokeratin-20 (reverse transcription polymerase chain reaction) in lymph nodes of patients with endometrial cancer.
A. Fishman (2000)
10.1056/NEJMOA040766
Circulating tumor cells, disease progression, and survival in metastatic breast cancer.
M. Cristofanilli (2004)
10.1111/j.1572-0241.2000.02199.x
Bone marrow micrometastases and gastrointestinal cancer detection and significance
D. Maguire (2000)
10.1021/AC990284+
Flow rate optimization for the quadrupole magnetic cell sorter.
P. S. Williams (1999)
RL.Detection of rare MCF-7breast cancer cells from mixtures of peripheral blood leucocytes by magnetic deposition analysis
B Fang (1999)
10.1081/SS-120002215
Pulse-injection studies of blood progenitor cells in a quadrupole magnet flow sorter
M. Hoyos (2002)
10.1097/00008390-200208000-00002
Magnetic bead RT-PCR: establishment of a new method for detecting circulating melanoma cells
Juliana Georgieva (2002)
10.1002/1097-0215(20000720)89:4<337::AID-IJC4>3.0.CO;2-R
Detection of tumor cells in blood using CD45 magnetic cell separation followed by nested mutant allele‐specific amplification of p53 and K‐ras genes in patients with colorectal cancer
H. Iinuma (2000)
10.1016/S0090-4295(01)01191-8
Changes in circulating carcinoma cells in patients with metastatic prostate cancer correlate with disease status.
J. Moreno (2001)
10.1056/NEJM200002243420801
Cytokeratin-positive cells in the bone marrow and survival of patients with stage I, II, or III breast cancer.
S. Braun (2000)
10.1016/S0304-8853(00)01254-3
Optimization of ferrofluids and protocols for the enrichment of breast tumor cells in blood
P. Liberti (2001)
10.1016/S0304-8853(98)00557-5
An instrument to determine the magnetophoretic mobility of labeled, biological cells and paramagnetic particles
J. Chalmers (1999)
10.1093/JNCI/91.13.1113
Detection and clinical importance of micrometastatic disease.
K. Pantel (1999)
10.1002/(SICI)1097-0320(19981201)33:4<469::AID-CYTO11>3.0.CO;2-6
Continuous, flow-through immunomagnetic cell sorting in a quadrupole field.
L. Sun (1998)
10.1021/bp010079r
Evaluation of Eluents from Separations of CD34+ Cells from Human Cord Blood Using a Commerical, Immunomagnetic Cell Separation System
K. Melnik (2001)
Circulating tumor cells in the peripheral blood of colorectal cancer patients
B Molnar (2001)
Immunomagnetic enrichment of disseminated epithelial tumor cells from peripheral blood by MACS.
V. Martin (1998)
Magnetic bead-PCR: establishment of a new method for detecting circulating melanoma cells. Melanoma Res
J Georgieva (2002)
10.1016/S0022-5347(05)67116-9
Immunomagnetic cell enrichment detects more disseminated cancer cells than immunocytochemistry in vitro.
R. Zigeuner (2000)
10.1002/BIT.10581
Characterization of antibody binding to three cancer-related antigens using flow cytometry and cell tracking velocimetry.
E. J. Chosy (2003)
10.1200/JCO.2002.02.152
Immunomagnetic enrichment and detection of micrometastases in colorectal cancer: correlation with established clinical parameters.
M. Weihrauch (2002)
10.1002/ijc.1217
Detection and enrichment of disseminated renal carcinoma cells from peripheral blood by immunomagnetic cell separation
U. Bilkenroth (2001)
10.1373/CLINCHEM.2003.028563
Multigene reverse transcription-PCR profiling of circulating tumor cells in hormone-refractory prostate cancer.
S. M. O'hara (2004)
Immunobead-PCR: a technique for the detection of circulating tumor cells using immunomagnetic beads and the polymerase chain reaction.
J. Hardingham (1993)
10.1002/(SICI)1097-0215(19981123)78:5<556::AID-IJC5>3.0.CO;2-G
Increased sensitivity for detection of micrometastases in bone‐marrow/peripheral‐blood stem‐cell products from breast‐cancer patients by negative immunomagnetic separation
B. Naume (1998)



This paper is referenced by
10.1371/journal.pone.0026925
Screening of Peptide Libraries against Protective Antigen of Bacillus anthracis in a Disposable Microfluidic Cartridge
Joshua M. Kogot (2011)
10.1016/J.SNB.2013.04.030
Cancer cell separator using size-dependent filtration in microfluidic chip
T. Masuda (2013)
Comparing Sequential Steps For Detection Of Circulating Tumor Cells : More Specific Or Just Less Sensitive ?
Nadine Rüdiger (2011)
10.1158/1078-0432.CCR-10-1105
Portable Filter-Based Microdevice for Detection and Characterization of Circulating Tumor Cells
H. Lin (2010)
10.1371/journal.pone.0172697
In situ single cell detection via microfluidic magnetic bead assay
F. Liu (2017)
10.1007/s10311-020-01072-z
Methods of detection of food-borne pathogens: a review
A. Saravanan (2020)
10.1016/j.ymeth.2013.09.006
Isolation and analysis of rare cells in the blood of cancer patients using a negative depletion methodology.
Y. Wu (2013)
10.5772/INTECHOPEN.84366
CTCs as Liquid Biopsy: Where Are We Now?
Laure Cayrefourcq (2019)
10.21037/atm-20-2751
Analysis of preoperative circulating tumor cells for recurrence in patients with hepatocellular carcinoma after liver transplantation.
Zhitao Chen (2020)
10.1016/j.jbiomech.2019.01.010
Numerical simulations of cell flow and trapping within microfluidic channels for stiffness based cell isolation.
Mutabe Aljaghtham (2019)
10.3389/fonc.2012.00131
Isolation and characterization of circulating tumor cells in prostate cancer
Elan Diamond (2012)
10.1201/B12138-3
Cancer and the Use of Biosensors for Cancer Clinical Testing
R. Chuaqui (2012)
10.1007/S10404-011-0798-1
Size-dependent microparticles separation through standing surface acoustic waves
J. Nam (2011)
10.1186/bcr3622
Heterogeneous atypical cell populations are present in blood of metastatic breast cancer patients
M. Lustberg (2013)
10.1021/ac3037766
Negative enrichment of circulating tumor cells using a geometrically activated surface interaction chip.
Kyung-A Hyun (2013)
10.1039/c9lc00575g
Hybrid negative enrichment of circulating tumor cells from whole blood in a 3D-printed monolithic device.
Chia-Heng Chu (2019)
10.5772/20492
Bacterial Display Peptides for Use in Biosensing Applications
Dimitra N. Stratis-Cullum (2011)
10.1364/OE.18.006396
Optical chromatography using a photonic crystal fiber with on-chip fluorescence excitation.
P. Ashok (2010)
10.1002/bit.22635
Quantification of non‐specific binding of magnetic micro‐ and nanoparticles using cell tracking velocimetry: Implication for magnetic cell separation and detection
J. Chalmers (2010)
10.1109/MEMSYS.2015.7050919
Cluster sizing of cancer cells by rail-based serial gap filtration in stopped-flow, continuous sedimentation mode
Macdara T. Glynn (2015)
10.1007/978-3-030-26439-0_2
Circulating Tumor Cell Enrichment Technologies.
Mert Boya (2020)
Integration methods for enhanced trapping and spectroscopy in optofluidics
P. Ashok (2011)
10.3389/fphys.2017.00381
The Potential for Circulating Tumor Cells in Pancreatic Cancer Management
Michael Pimienta (2017)
10.1016/J.SNB.2015.02.107
Portable Coulter counter with vertical through-holes for high-throughput applications
Y. Chen (2015)
10.1002/smll.201502120
Quantitative Magnetic Separation of Particles and Cells Using Gradient Magnetic Ratcheting.
C. Murray (2016)
10.14288/1.0073460
Automated microfluidic device for the separation of cancer cells from blood
B. K. Lin (2012)
10.1039/c6cs00803h
Current detection technologies for circulating tumor cells.
Z. Shen (2017)
Detecting uterine cervical cancer cells using molecular biomarkers
Ahmed Mousa (2015)
10.1063/1.3678303
An integrated micro-chip for rapid detection of magnetic particles
C. Gooneratne (2012)
10.1007/s12645-010-0007-z
Micro- and nanotechnology approaches for capturing circulating tumor cells
B. Panchapakesan (2010)
10.1007/978-3-540-92841-6_190
Microdevice for Trapping Circulating Tumor Cells for Cancer Diagnostics
S. J. Tan (2009)
10.1016/J.JMMM.2006.10.1160
Engineering quadrupole magnetic flow sorting for the isolation of pancreatic islets
D. Kennedy (2007)
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