Determination of lymphocyte division by flow cytometry.

Techniques currently available for determining cell division are able to show one or, at best, a limited number of cell divisions. Other methods exist which can quantify overall division, but tell nothing about the division history of individual cells. Here we present a new technique in which an intracellular fluorescent label is divided equally between daughter cells upon cell division. The technique is applicable to in vitro cell division, as well as in vivo division of adoptively transferred cells, and can resolve multiple successive generations using flow cytometry. The label is fluorescein derived, allowing monoclonal antibodies conjugated to phycoerythrin or other compatible fluorochromes to be used to immunophenotype the dividing cells.

[1]  T. Mosmann.  Rapid colorimetric assay for cellular growth and survival: application to proliferation and cytotoxicity assays. , 1983, Journal of immunological methods.

[2]  D. V. van Bockstaele,et al.  Improved BrdUrd-Hoechst bivariate cell kinetic analysis by helium-cadmium single laser excitation. , 1992, Cytometry.

[3]  C. Parish,et al.  New fluorescent dyes for lymphocyte migration studies. Analysis by flow cytometry and fluorescence microscopy. , 1990, Journal of immunological methods.

[4]  K. Rajewsky,et al.  Flow cytometric analysis of cell proliferation dynamics in the B cell compartment of the mouse. , 1989, International immunology.

[5]  C. Janeway,et al.  Microbial induction of co-stimulatory activity for CD4 T-cell growth. , 1991, International immunology.

[6]  H. Gratzner,et al.  Monoclonal antibody to 5-bromo- and 5-iododeoxyuridine: A new reagent for detection of DNA replication. , 1982, Science.

[7]  R. M. Bohmer,et al.  FLOW CYTOMETRIC CELL CYCLE ANALYSIS USING THE QUENCHING OF 33258 HOECHST FLUORESCENCE BY BROMODEOXYURIDINE INCORPORATION , 1979 .

[8]  C. Parish,et al.  A procedure for removing red cells and dead cells from lymphoid cell suspensions. , 1975, Journal of immunological methods.

[9]  D. Houck,et al.  Simultaneous analysis of cell surface antigens, bromodeoxyuridine incorporation and DNA content. , 1985, Cytometry.

[10]  A. Freitas,et al.  Lymphocyte population kinetics during the development of the immune system. B cell persistence and life-span can be determined by the host environment. , 1989, International immunology.

[11]  I. Stamenkovic,et al.  A 39-kDa protein on activated helper T cells binds CD40 and transduces the signal for cognate activation of B cells. , 1992, Proceedings of the National Academy of Sciences of the United States of America.