Biomedical subjects

Larissa Belov

Publications and source records attributed to Larissa Belov.

7 recordsLinked to original sources

PubMed2006

Analysis of human leukaemias and lymphomas using extensive immunophenotypes from an antibody microarray.

A novel antibody microarray has been developed that provides an extensive immunophenotype of leukaemia cells. The assay is a solid phase cell-capture technique in which 82 antigens are studied simultaneously. This paper presents the analysis of 733 patients with a variety of leukaemias and lymphomas from peripheral blood and bone marrow. Discriminant Function Analysis of the expression profiles from these 733 patients and 63 normal subjects were clustered and showed high levels of consistency with diagnoses obtained using conventional clinical and laboratory criteria. The overall levels of consensus for classification using the microarray compared with established criteria were 93.9% (495/527 patients) for peripheral blood and 97.6% (201/206 patients) for bone marrow aspirates, showing that the extensive phenotype alone was frequently able to classify the disease when the leukaemic clone was the dominant cell population present. Immunophenotypes for neoplastic cells were distinguishable from normal cells when the leukaemic cell count was at least 5 x 10(9) cells/l in peripheral blood, or 20% of cells obtained from bone marrow aspirates. This technique may be a useful adjunct to flow cytometry and other methods when an extensive phenotype of the leukaemia cell is desired for clinical trials, research and prognostic factor analysis.

Acute Disease↗

PubMed2006

Multiplex detection of surface molecules on colorectal cancers.

A technique of fluorescence multiplexing is described for analysis of the plasma membrane proteome of colorectal cancer cells from surgically resected specimens, enabling detection and immunophenotyping when the cancer cells are in the minority. A single-cell suspension was prepared from a colorectal tumour, and the mixed population of cells was captured on a CD antibody microarray. The cancer cells were detected using a fluorescently tagged antibody for carcinoembryonic antigen (CEA-Alexa647) or epithelial cell adhesion marker (EpCAM-Alexa488). Using this multiplexing procedure, dot patterns from colorectal cancers were distinct from those of adjacent normal tissue. Subtraction of the expression levels for each antigen from normal tissue from those for the cancer shows differential expression in the cancer of CD66c, CD15s, CD55, CD45, CD71, CD45RO, CD11b and CEA, in descending order. Cells captured on the same microarray were also labelled with fluorescent CD3-phycoerythrin antibody revealing the presence of tumour-infiltrating lymphocytes. The immunophenotypes of T lymphocytes from the tumour samples showed differential expression of HLA-DR, TCR alpha/beta, CD49d, CD52, CD49e, CD5, CD95, CD28, CD38 and CD71, in descending order. Fluorescence multiplexing of mixed cell populations captured on a single antibody microarray enables expression profiling of multiple sub-populations of cells within a tumour sample.

CD4 Antigens↗

PubMed2006

Classification of AML using a monoclonal antibody microarray.

A cluster of differentiation (CD) antibody microarray called the DotScan microarray has been developed that enables an extensive immunophenotype to be obtained for a suspension of leukocytes in a single analysis. For a leukemia with a leukemia count of greater than 10 x 10(9)/L, the immunophenotype obtained is essentially that of the leukemic clone. The antibody microarray is printed as microscopic (10 nL) dots on a nitrocellulose film on a microscope slide. Cells are captured by the immobilized antibodies and a dot pattern is recorded with an optical array reader giving the immunophenotype of the leukemia. Procedures are being developed that should enable diagnosis of myeloid leukemias by comparison of the dot pattern obtained from an unknown blood sample with a library of consensus patterns for the common leukemias.

Antibodies, Neoplasm↗

PubMed2005

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.

Antibodies, Monoclonal↗

PubMed2005

Conservation of unique cell-surface CD antigen mosaics in HIV-1-infected individuals.

Cluster of differentiation (CD) antigens are expressed on cells of myeloid and lymphoid lineages. As most disease processes involve immune system activation or suppression, these antigens offer unique opportunities for monitoring host responses. Immunophenotyping using limited numbers of CD antigens enables differentiation states of immune system cells to be determined. Extended phenotyping involving parallel measurement of multiple CD antigens may help identify expression pattern signatures associated with specific disease states. To explore this possibility we have made a CD monoclonal antibody array and scanner, enabling the parallel immunophenotyping of leukocyte cell suspensions in a single and rapid analysis. To demonstrate this approach, we used the specific example of patients infected with human immunodeficiency virus type-1 (HIV-1). An invariant HIV-induced CD antigen signature has been defined that is both robust and independent of clinical outcome, composed of a unique profile of CD antigen expression levels that are both increased and decreased relative to internal controls. The results indicate that HIV-induced changes in CD antigen expression are disease specific and independent of outcome. Their invariant nature indicates an irreversible component to retroviral infection and suggests the utility of CD antigen expression patterns in other disease settings.

Antibodies, Monoclonal↗

PubMed2005

Immunophenotypic changes induced on human HL60 leukaemia cells by 1alpha,25-dihydroxyvitamin D3 and 12-O-tetradecanoyl phorbol-13-acetate.

1alpha,25-Dihydroxyvitamin D3 (1,25D3) induces HL60 cells to acquire a monocyte-like phenotype, while cells treated with 12-O-tetradecanoyl phorbol-13-acetate (TPA) resemble macrophages. Using a microarray of 82 CD antibodies, 24 cluster of differentiation (CD) antigens were detected on HL60 cells. 1,25D3 induced the following antigens in decreasing order of the change: CD14, CD11c, CD11b, CD54, CD86, CD38 and CD66c, with repression of CD117, CD71, CD95, CD45 and CD64. TPA induced the following antigens in decreasing order of the change: CD11c, CD9, CD11b, CD54, CD38, CD45RO and CD66c, with repression of CD4, CD117, CD95, CD71 and CD64. The results presented provide a basis for monitoring differentiation therapy of myeloid leukaemias in patients.

Antigens, CD↗

PubMed2003

Identification of repertoires of surface antigens on leukemias using an antibody microarray.

We have previously described a microarray of cluster of differentiation (CD) antibodies that enables concurrent determination of more than 60 CD antigens on leukocytes. This procedure does not require protein purification or labeling, or a secondary detection system. Whole cells are captured by a microarray of 10 nL antibody dots immobilized on a nitrocellulose film on a microscope slide. Distinct patterns of cell binding are observed for different leukemias or lymphomas. These haematological malignancies arise from precursor cells of T- or B-lymphocytic, or myeloid lineages of hematopoiesis. The dot patterns obtained from patients are distinct from those of peripheral blood leukocytes from normal subjects. This microarray technology has recently undergone a number of refinements. The microarray now contains more CD antibodies, and a scanner for imaging dot patterns and software for data analysis provide an extensive immunophenotype sufficient for diagnosis of common leukemias. The technology is being evaluated for diagnosis of leukemias with parallel use of conventional diagnostic criteria.

Antibodies↗