Simple and rapid measurement of human T lymphocytes and their subclasses in peripheral blood.

Hoffman, R. A.; Kung, Patrick; Hansen, W.; Goldstein, Gideon

doi:10.1073/pnas.77.8.4914

Cited by 387 publications

(145 citation statements)

References 14 publications

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“…CD4 + and CD8 + T cells were isolated from PBMC by depletion of monocytes by plastic adherence for 2 h at 37°C. Non-adherent cells were stained with supernatants from hybridomas A9 (anti-CD16, kindly provided by M. Pfreundschuh, Homburg/Saar, Germany), HD37 (anti-CD19) [35] and HP2/6 (anti-CD4) [36] to obtain CD8 + cells or OKT8 (anti-CD8) [37] to isolate CD4 + T cells. Negative isolation of T cells was performed with BioMag goat antimouse IgG Beads (Paesel + Lorei, Hanau, Germany).…”

Section: Induction and Analysis Of Apoptosismentioning

confidence: 99%

Induction of apoptosis and modulation of activation and effector function in T cells by immunosuppressive drugs

Strauß

Osen

Debatin

2002

Clinical and Experimental Immunology

149

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SUMMARYImmunosuppressive drugs (ISD) are used for the prevention and treatment of graft rejection, graftversus-host-disease (GVHD) and autoimmune disorders. The precise mechanisms by which ISD interfere with T cell activation and effector function or delete antigen-specific T cells are defined only partially. We analysed commonly used ISD such as dexamethasone (DEX), mycophenolic acid (MPA), FK506, cyclosporin A (CsA), rapamycin (RAP), methotrexate (MTX) and cyclophosphamide (CP) for apoptosis-induction and modulation of activation and effector function in human peripheral T cells, cytotoxic T cell lines (CTL) and Jurkat T cells. Of all drugs tested only CP and MTX prevented antigenspecific proliferation of T cells and decreased cytotoxicity of alloantigen specific CTL lines by direct induction of apoptosis. MTX and CP also slightly increased activation-induced cell death (AICD) and CD95-sensitivity. In contrast, all other drugs tested did not induce T cell apoptosis, increase CD95-sensitivity or AICD. CsA and FK506 even prevented AICD by down-modulation of CD95L. DEX, MPA, CsA, FK506 and RAP inhibited activation of naive T cells, but were not able to block proliferation of activated T cells nor decrease cytotoxic capacity of CTL lines. These results show that ISD can be classified according to their action on apoptosis-induction and inhibition of proliferation and would favour a rational combination therapy to delete existing reactive T cells and prevent further T cell activation.

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Section: Induction and Analysis Of Apoptosismentioning

confidence: 99%

Induction of apoptosis and modulation of activation and effector function in T cells by immunosuppressive drugs

Strauß

Osen

Debatin

2002

Clinical and Experimental Immunology

149

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“…(c) The ratio of helper (CD4) to suppressor (CD8) T cells was measured by automated flow cytometry at Diaclin Laboratories (Nashville, TN) (The normal CD4:CD8 ratio found in our laboratory is 2.06 * 0.70, mean 2 SD.) (17). (d) Complete HLA typing for A, B, C and DR loci was performed for most of the patients by Dr. Peter Stastny at the University of Texas Health Science Center in Dallas and, for some of the VAMC patients, by Dr. Gary Niblack at the Nashville VAMC, using a microcytotoxicity assay (18).…”

mentioning

confidence: 99%

In vitro rheumatoid factor synthesis in patients taking second‐line drugs for rheumatoid arthritis

Olsen

Callahan

Pincus

1988

Arthritis & Rheumatism

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Rheumatoid arthritis (RA) patients whose unstimulated peripheral blood mononuclear cells produce high levels of IgM rheumatoid factor (IgM‐RF) in vitro have more severe disease activity. RA patients being treated with second‐line agents, including gold salts, penicillamine, or methotrexate, tend to be low producers or nonproducers of IgM‐RF in vitro. The possibility that low production or nonproduction of IgM‐RF in vitro may be explained by treatment with second‐line agents alone, irrespective of disease activity, was analyzed in 133 RA patients whose disease status was assessed by multiple laboratory and clinical measures. The results indicate that treatment with second‐line agents and in vitro IgM‐RF synthesis are independently associated with disease activity.

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“…Flow cytometric differential blood cell counting is based on various principles, including cytochemical stains (9,15), fluorescent stains such as acridine orange (1,2,18), and differences in electronic cell volume (19) or in orthogonal light scattering (7,8). Differences in P12 uptake, demonstrated in the present report, provide a potential additional parameter for differential cell counting.…”

Section: Resultsmentioning

confidence: 70%

Flow cytofluorometric analysis of the uptake of the fluorescent fatty acid pyrene‐dodecanoic acid by human peripheral blood cells

et al. 1988

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The fluorescence activated cell sorter (FACS) was used for measuring the uptake of the fluorescent fatty acid derivative U-(l-pyrene) dodecanoic acid (Pa) by human peripheral blood cells. The results indicate that blood cells differ widely in their ability to take up Pl2, with polymorphonuclear cells showing the greatest uptake, followed by lymphocytes, platelets, and RBCs. These differences in Pl2 uptake provide a potential additional parameter for differential cell counting. Using the ability of the FACS to "gate out" nonrelevant cells, it was possible to measure the rate of Pl2 uptake by each respective cell type even when admixed with other cells. Thus elaborate physical separation procedures could be avoided, and contaminating cells did not influence the results. Differences in Pl2 uptake were also utilized to separate blood cells into pure subpopulations of specific cell types. Key terms: Fluorescence activated cell sorter, fatty acid derivativesA major obstacle in studying biological and biochemical properties of specific cells is the fact that, in vivo, cells are admixed with a variety of other cell types. For example, the study of hemopoietic cells is limited by the heterogeneity of a population consisting of cells belonging to several lineages that are at various stages of maturation. Analysis of such cells requires complete separation into discrete, homogenous subpopulations, since even a small contamination by cells that differ considerably in the property examined from the subpopulation studied may affect the results.We have demonstrated that the fluorescent, mediumchain fatty acid lZ-(l-pyrene) dodecanoic acid (P12) is effectively transported across the membranes of several cultured cell types and incorporated into their neutrallipids and phospholipids (10,ll). Using in vitro established leukemic cell lines, we have shown that cells of different hemopoietic lineages and stages of maturation differ in the rate and extent of PI2 uptake as well as in their metabolism of this acid (43). Recently, we have shown that the cellular uptake of P12 can be measured by the fluorescence activated cell sorter (FACS) (13,141, The present study describes the uptake of P12 by various cell types present in human peripheral blood and shows that blood cells differ widely in their ability to take up the compound. By taking advantage of the abilpossible to measure the rate of P12 uptake by each respective cell type, even when admixed with other cells. Thus elaborate separation procedures were avoided, and contaminating cells did not influence the results. Differences in P12-uptake were also utilized to separate blood cells into pure subpopulations of specific cell types. MATERIALS AND METHODS CellsPeripheral blood from normal individuals cells was collected in preservative-free heparin or citrate. Buffy coat cells were separated by mixing 2 volumes of blood with 1 volume of 1% dextran in saline (Pharmacia, Uppsala, Sweden) and allowing it to sediment at unit gravity for about 1 hour at room temperature. The cellcontai...

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Simple and rapid measurement of human T lymphocytes and their subclasses in peripheral blood.

Cited by 387 publications

References 14 publications

Induction of apoptosis and modulation of activation and effector function in T cells by immunosuppressive drugs

Induction of apoptosis and modulation of activation and effector function in T cells by immunosuppressive drugs

In vitro rheumatoid factor synthesis in patients taking second‐line drugs for rheumatoid arthritis

Flow cytofluorometric analysis of the uptake of the fluorescent fatty acid pyrene‐dodecanoic acid by human peripheral blood cells

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