CBER Expertise
Flow cytometry: a way to characterize cell populations
Principal Investigator: Gerald E. Marti, MD
Office / Division / Lab: OCTGT / DCGT / CTTB
Overview
Public Health Issue: New treatments techniques are needed for many diseases, including forms of cancer. Cellular therapies for these diseases are being developed, and must be characterized properly in order to be safe and effective. In addition, clinical trials of gene therapies and tumor vaccines include testing of patient cells for biomarkers. These efforts require sensitive and quantitative methods for measuring markers on cell surfaces. Another public health need is early detection of cancer, which is important for successful treatment. We have found a new approach to the discovery of biomarkers that can be used for early cancer detection.
Regulatory Contribution: Multi-color flow cytometry is a very sensitive method that permits the detection of cells expressing combinations of surface markers. Testing for such markers is necessary for identity, purity, and potency testing of biological products such as cell therapy products. These methods are also necessary for monitoring immune cell subsets during preclinical studies and clinical trials. The expertise in this program and our contributions to standards for quantitative flow cytometry have been critical in regulating various products, e.g., review of flow cytometry-based potency assays in regulatory submissions.
Research Approach: We have developed quantitative methods for flow cytometry, an approach that allows reproducible measurements of cell populations and their surface biomarkers. Microbead standards are used to establish calibration curves, which, in turn allow development of instrument performance measures and comparison of results from one assay to another. This approach to standardization can be applied to quality control of cellular products, and also to discovery of biomarkers. Also, in the event of a bioterrorism act, such markers of abnormal cells could be used to monitor the state of blood cells in individuals after radiation, chemical or biological exposure. One example is our discovery of a biomarker in familial chronic lymphocytic leukemia (CLL), the most common hematological malignancy (blood cancer) in older adults. We used flow cytometry to detect small populations of abnormal blood cells whose presence provides a biomarker long before cancer symptoms develop. Our work can lead to earlier detection of disease, resulting in a better prognosis for individuals and families, and to new treatment strategies that can be tested in animal models and later in clinical trials.
Mission Relevance and Outcomes: Flow cytometry is often a key technique for product characterization, patient monitoring, and in vitro diagnostic testing. Our work has supported bringing multicolor flow cytometry from the research laboratory into the clinical laboratory and the product quality control arena. In addition, we have developed biomarkers that are needed in trials of numerous cellular therapy products and tumor vaccines. In addition, we are collaborating with CDC and NIST on an initiative to develop a national laboratory of fluorescent standards for use in these applications.
Publications
Cytotherapy 2007;9(2):123-32
CD69 expression as an index of T-cell function: assay standardization, validation and use in monitoring immune recovery.
Lindsey WB, Lowdell MW, Marti GE, Abbasi F, Zenger V, King KM, Lamb LS
Cytometry A 2007 Jun;71(6):414-37
International Society for Analytical Cytology biosafety standard for sorting of unfixed cells.
Schmid I, Lambert C, Ambrozak D, Marti GE, Moss DM, Perfetto SP; International Society of Analytical Cytology
Blood 2007 Jun 15;109(12):5079-86
Abnormal microRNA-16 locus with synteny to human 13q14 linked to CLL in NZB mice.
Raveche ES, Salerno E, Scaglione BJ, Manohar V, Abbasi F, Lin YC, Fredrickson T, Landgraf P, Ramachandra S, Huppi K, Toro JR, Zenger VE, Metcalf RA, Marti GE
Blood 2007 Feb 1;109(3):916-25
Identification of a novel chromosome region, 13q21.33-q22.2, for susceptibility genes in familial chronic lymphocytic leukemia.
Ng D, Toure O, Wei MH, Arthur DC, Abbasi F, Fontaine L, Marti GE, Fraumeni JF Jr, Goldin LR, Caporaso N, Toro JR
Cytometry B Clin Cytom 2006 Nov 15;70(6):410-5
Comparison of fluorescein and phycoerythrin conjugates for quantifying CD20 expression on normal and leukemic B-cells.
Wang L, Abbasi F, Gaigalas AK, Vogt RF, Marti GE
Cytometry B Clin Cytom 2006 Aug 11;70B(4):197-200
ZAP-70 in CLL: Towards standardization of a biomarker for patient management: History of clinical cytometry special issue.
Marti G, Orfao A, Goolsby C
Br J Haematol 2006 Apr;133(1):59-61
High-density mapping and follow-up studies on chromosomal regions 1, 3, 6, 12, 13 and 17 in 28 families with chronic lymphocytic leukaemia.
Ng D, Marti GE, Fontaine L, Toro JR, Caporaso N, Goldin LR
Leukemia 2005 Dec;19(12):2339-41
Identification of a new monoclonal B-cell subset in unaffected first-degree relatives in familial chronic lymphocytic leukemia.
Aurran-Schleinitz T, Telford W, Perfetto S, Caporaso N, Wilson W, Stetler-Stevenson MA, Zenger VE, Abbasi F, Marti GE
Br J Haematol 2005 Aug;130(3):325-32
Diagnostic criteria for monoclonal B-cell lymphocytosis.
Marti GE, Rawstron AC, Ghia P, Hillmen P, Houlston RS, Kay N, Schleinitz TA, Caporaso N, The International Familial CLL Consortium
Blood 2004 Sep 1;104(5):1428-34
Fludarabine treatment of patients with chronic lymphocytic leukemia induces a p53-dependent gene expression response.
Rosenwald A, Chuang EY, Davis RE, Wiestner A, Alizadeh AA, Arthur DC, Mitchell JB, Marti GE, Fowler DH, Wilson WH, Staudt LM
Semin Hematol 2004 Jul;41(3):201-6
Perspectives on familial chronic lymphocytic leukemia: genes and the environment.
Caporaso N, Marti GE, Goldin L
Leukemia 2004 Mar;18(3):597-606
Studies in NZB IL-10 knockout mice of the requirement of IL-10 for progression of B-cell lymphoma.
Czarneski J, Lin YC, Chong S, McCarthy B, Fernandes H, Parker G, Mansour A, Huppi K, Marti GE, Raveche E
Haematologica 2004 Mar;89(3):262-3
Familial lymphoid neoplasms in patients with mantle cell lymphoma.
Marti GE
Clin Cancer Res 2004 Feb 1;10(3):1047-56
Autoreactive, cytotoxic T lymphocytes specific for peptides derived from normal B-cell differentiation antigens in healthy individuals and patients with B-cell malignancies.
Grube M, Rezvani K, Wiestner A, Fujiwara H, Sconocchia G, Melenhorst JJ, Hensel N, Marti GE, Kwak LW, Wilson W, Barrett JA
Cytometry 2004 Jan;57B(1):1-6
Formalization of the MESF unit of fluorescence intensity.
Schwartz A, Gaigalas AK, Wang L, Marti GE, Vogt RF, Fernandez-Repollet E