Screening microarrays of novel monoclonal antibodies for binding to T-, B- and myeloid leukaemia cells.
We have developed a microarray (DotScan) that enables rapid immunophenotyping and classification of leukaemias and lymphomas by measuring the capture of cells by immobilized dots of 82 CD antibodies [Belov, L., de la Vega, O., dos Remedios, C.G., Mulligan, S.P., 2001. Immunophenotyping of leukemia using a cluster of differentiation antibody microarray. Cancer Res. 61, 4483; Belov, L., Huang, P., Barber, N., Mulligan, S.P., Christopherson, R.I., 2003. Identification of repertoires of surface antigens on leukemias using an antibody microarray. Proteomics 3, 2147]. The DotScan technology has been used to investigate the properties of 498 new antibodies submitted to the HLDA8 Workshop. These antibodies have been applied as 10 nl dots to a film of nitrocellulose on a microscope slide to make an HLDA8 microarray. After blocking the remaining nitrocellulose surface, individual arrays were incubated with each of 7 cell types from a human leukaemia cell panel consisting of three cell lines, CCRF-CEM (a T-cell acute lymphocytic leukaemia), MEC-1 (derived from B-cell chronic lymphocytic leukaemia) and HL-60 (a promyelocytic leukaemia), and four leukaemias from patients: a T-cell prolymphocytic leukaemia, a B-cell chronic lymphocytic leukaemia, and two acute myeloid leukaemias. Leukaemia cells were captured by those immobilized antibodies for which they expressed the corresponding surface molecule. Unbound cells were gently washed off, bound cells were fixed to the arrays and dot patterns were recorded using a DotScan array reader and quantified using DotScan data analysis software. The data obtained show the unique expression profiles of the 7 cell types in the leukaemia cell panel obtained with the DotScan microarray, and the differential capture patterns for these 7 cell types screened against the 498 antibodies in the HLDA8 microarray constructed for this study.
Belov
Larissa
L
Medsaic Pty. Ltd., Suite 145, National Innovation Centre, Australian Technology Park, Garden Street, Eveleigh, NSW 1430, Australia.
Huang
Pauline
P
Chrisp
Jeremy S
JS
Mulligan
Stephen P
SP
Christopherson
Richard I
RI
D000911:Antibodies, Monoclonal / Q000737:chemistry / Q000276:immunology*; D015703:Antigens, CD / Q000032:analysis*; D045744:Cell Line, Tumor; D003101:Collodion / Q000737:chemistry; D005434:Flow Cytometry; D006801:Humans; D007938:Leukemia / Q000145:classification* / Q000276:immunology; D015448:Leukemia, B-Cell / Q000145:classification / Q000276:immunology; D007951:Leukemia, Myeloid / Q000145:classification / Q000276:immunology; D015458:Leukemia, T-Cell / Q000145:classification / Q000276:immunology; D040081:Protein Array Analysis / Q000379:methods*
D000911:Antibodies, Monoclonal; D015703:Antigens, CD; D003101:Collodion