Surface antigen profiling of colorectal cancer using antibody microarrays with fluorescence multiplexing

• Published in: Journal of immunological methods Vol. 355; no. 1; pp. 40 - 51
• Main Authors: Zhou, Jerry, Belov, Larissa, Huang, Pauline Y., Shin, Joo-Shik, Solomon, Michael J., Chapuis, Pierre H., Bokey, Leslie, Chan, Charles, Clarke, Candice, Clarke, Stephen J., Christopherson, Richard I.
• Format: Journal Article
• Language: English
• Published: Amsterdam Elsevier B.V 15.04.2010; Elsevier
• Subjects: Adult; Aged; Aged, 80 and over; Antibodies, Monoclonal - chemistry; Antibodies, Neoplasm - chemistry; Antibody microarrays; Antigens, Neoplasm - metabolism; Biological and medical sciences; CD antigens; Cell Line, Tumor; Colorectal cancer; Colorectal Neoplasms - metabolism; Female; Fluorescence; Fluorescent Dyes - chemistry; Fundamental and applied biological sciences. Psychology; Fundamental immunology; Humans; Leukocytes; Male; Middle Aged; Molecular immunology; Multiplexing; Protein Array Analysis - methods; Quantum Dots; Techniques; Multiplexing; CD; QDs; EpCAM; IHC; Colorectal cancer; Antibody microarrays; Fluorescence; Leukocytes; ACP; EGFR; CD antigens; BSA; TAMs; CRC; FCS; TILs; HBSS; CEA; MeV; Rectal disease; Antibody; Leukocyte; Malignant tumor; Immunological method; Colonic disease; Antigen; Digestive diseases; Intestinal disease; Cancer
• ISSN: 0022-1759; 1872-7905
• DOI: 10.1016/j.jim.2010.01.015

A procedure is described for the disaggregation of colorectal cancers (CRC) and normal intestinal mucosal tissues to produce suspensions of viable single cells, which are then captured on customized antibody microarrays recognising 122 different surface antigens (DotScan™ CRC microarray). Cell binding patterns recorded by optical scanning of microarrays provide a surface profile of antigens on the cells. Sub-populations of cells bound on the microarray can be profiled by fluorescence multiplexing using monoclonal antibodies tagged with Quantum Dots or other fluorescent dyes. Surface profiles are presented for 6 CRC cell lines (T84, LIM1215, SW480, HT29, CaCo and SW620) and surgical samples from 40 CRC patients. Statistical analysis revealed significant differences between profiles for CRC samples and mucosal controls. Hierarchical clustering of CRC data identified several disease clusters that showed some correlation with clinico-pathological stage as determined by conventional histopathological analysis. Fluorescence multiplexing using Phycoerythrin- or Alexa Fluor 647-conjugated antibodies was more effective than multiplexing with antibodies labelled with Quantum Dots. This relatively simple method yields a large amount of information for each patient sample and, with further application, should provide disease signatures and enable the identification of patients with good or poor prognosis.