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Lymphocyte stimulation by concanavalin a studied by the fluorescent probe acridine orange

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Summary

Viable mouse thymocytes or spleen leucocytes stained with acridine orange (AO) were divided into one part used for stimulation, and the other part for control. Analysis of cellular green-fluorescence emission enabled physicochemical changes in lymphocytes to be detected after 30 min stimulation with the mitogens concanavalin A (Con A) and pokeweed mitogen (PWM). No change in fluorescence was observed with the nonmitogenic reagent wheat germ lectin (WGL) or with allogeneic cell stimulation (MLR). When green fluorescence intensity of individual cells was monitored by microfluorimetry, 30 min stimulation with Con A induced an increase, whereas PWM induced a decrease. When analysed by fluorescence spectrophotometry, Con A induced a 2 nm blue shift in emission maximum and a decrease in polarization values.

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References

  • Auer G, Zetterberg A, Killander D (1970) Changes in binding between DNA and arginine residues in histone induced by cell crowding. Exp Cell Res 62:32–38

    Google Scholar 

  • Azzi A (1975) The application of fluorescent probes in membrane studies. Q Rev Biophys 8:237–316

    Google Scholar 

  • Cercek L, Cercek B (1977) Application of the phenomenon of changes in the structure of cytoplasmic matrix (SCM) in the diagnosis of malignant disorders: a review. Eur J Cancer 13:903–915

    Google Scholar 

  • Cunningham AJ (1975) Why do so many cells take part in mixed lymphocyte reactions? Cellular Immunol 19:368–371

    Google Scholar 

  • Darzynkiewicz Z (1979) Acridine orange as a molecular probe in studies of nucleic acid in situ. In: Melamed MR, Mullaney PF, Mendelsohn ML (eds) Flow Cytometry and Sorting. John Wiley & Sons Inc, New York, pp 285–316

    Google Scholar 

  • Darzynkiewicz Z, Bolund L, Ringertz NR (1969) Nucleoprotein changes and initiation of RNA synthesis in PHA stimulated lymphocytes. Exp Cell Res 56:418–424

    Google Scholar 

  • Edelman GM (1974) Surface alterations and mitogenesis in lymphocytes. In: Clarkson B, Baserga R (eds) Control of Proliferation in Animal Cells. Cold Spring Harbor Press, New York, pp 357–377

    Google Scholar 

  • Greene WC, Parker CM, Parker CW (1976) Opposing effects of mitogenic and nonmitogenic lectins on lymphocyte activation: evidence that wheat germ agglutinin produced a negative signal. J Biol Chem 251:4017–4025

    Google Scholar 

  • Halliday GM, Nairn RC, Pallett MA, Rolland JM, Ward HA (1979) Detection of early lymphocyte activation by the fluorescent cell membrane probe N-phenyl-1-naphthylamine. J Immunol Methods 28:381–390

    Google Scholar 

  • Hirschhorn R, Brittinger G, Hirschhorn K, Weissmann G (1968) Studies on lysosomes. XII. Redistribution of acid hydrolases in human lymphocytes stimulated by phytohemagglutinin. J Cell Biol 37:412–423

    Google Scholar 

  • Killander D, Rigler R (1965) Initial changes of deoxyribonucleoprotein and synthesis of nucleic acid in phytohemagglutinine-stimulated human leucocytes in vitro. Exp Cell Res 39:701–704

    Google Scholar 

  • Killander D, Rigler R (1969) Activation of deoxyribonucleoprotein in human leucocytes stimulated by phytohemagglutinin. Exp Cell Res 54:163–170

    Google Scholar 

  • Liedeman RR, Matveyeva NP, Vostricova SA, Prilipko LL (1975) Extrinsic factors affecting the binding of acridine organe of the DNP complex of cell nuclei in different physiological states. Exp Cell Res 90:105–110

    Google Scholar 

  • Meissel MN, Zelenin AV (1973) Fluorescence cytochemistry of nucleic acids. Its modern state and prospects. In: Grey W, Rizzo ND (eds) Unity Through Diversity. Gordon & Breach, New York, pp 701–738

    Google Scholar 

  • Menter JM, Hurst RE, West SS (1979) Photochemistry of heparin-acridine orange complexes in solution: photochemical changes occurring in the dye and polymer on fluorescence fading. Photochem Photobiol 29:473–478

    Google Scholar 

  • Mysliwska J, Mysliwski A, Witkowski J (1976) Comparison of the early chroma tin reaction of the mouse thymocytes incubated by phytohemagglutinin and concanavalin A. Cell Tissue Res 166:553–561

    Google Scholar 

  • Nairn RC (1976) Fluorescent protein tracing, 4th edn. Churchill Livingstone, Edinburgh

    Google Scholar 

  • Nairn RC, Rolland JM (1980) Review. Fluorescent probes to detect lymphocyte activation. Clin Exp Immunol 39:1–13

    Google Scholar 

  • Nairn RC, Rolland JM, Halliday GM, Jablonka IM, Ward HA (1978) Fluorescent probes to monitor early lymphocyte activation. In: Knapp W, Holubar K, Wick G (eds) Immunofluorescence and Related Staining Techniques. Elsevier/North-Holland, Amsterdam, pp 57–66

    Google Scholar 

  • Ohba Y (1966) Structure of nucleohistone 1. Hydrodynamic behaviour. Biochim Biophys Acta 123:76–83

    Google Scholar 

  • Pritchard JAV, Sutherland WH, Seaman JE, Deeley TJ, Evans IH, Kerby IJ, James KW, Paterson ICM, Davies BH (1978) Cancer-specific density changes in lymphocytes after stimulation with phytohemagglutinin. Lancet 2:1275–1277

    Google Scholar 

  • Rigler R (1966) Microfluorometric characterization of intracellular nucleic acids and nucleoproteins by acridine orange. Acta Physiol Scand 67: Suppl 267, 1–122

    Google Scholar 

  • Rigler R (1969) Acridine orange in nucleic acid analysis. Ann NY Acad Sci 157:211–224

    Google Scholar 

  • Rigler R, Killander D (1969) Activation of deoxyribonucleoprotein in human leucocytes stimulated by phytohemagglutinin. Exp Cell Res 54:171–180

    Google Scholar 

  • Ringertz NR, Bolund L, Darzynkiewicz Z (1970) AO binding of intracellular nucleic acid in fixed cells in relation to cell growth. Exp Cell Res 63:233–238

    Google Scholar 

  • Rolland JM, Betts RL, Halliday GM, Hocking GR, Nairn RC (1980) Early changes in concanavalin A-stimulated lymphocytes detected by the fluorescent probe N-phenyl-l-naphthylamine. Cell Tissue Res 214:119–128

    Google Scholar 

  • Segel GB, Lichtman MA (1977) Transmembrane K+ turnover by phytohemagglutinin (PHA)stimulated human lymphocytes. In: Lucas DO (ed) Regulatory Mechanisms in Lymphocyte Activation. Academic Press, New York, pp 432–434

    Google Scholar 

  • Sokal RR, Rohlf FJ (1969) Biometry. W.H. Freedman & Company, San Francisco

    Google Scholar 

  • Watson JD (1970) Molecular biology of the gene. 2nd edn. W.A. Benjamin Inc, New York

    Google Scholar 

  • Zubay G, Doty P (1959) The isolation and properties of deoxyribonucleoprotein particles containing single nucleic acid molecules. J Molec Biol 1:1–20

    Google Scholar 

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Supported by grants from the Anti-Cancer Council of Victoria. We are grateful to Dr. H.A. Ward for helpful discussion

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Halliday, G.M., Nairn, R.C. & Rolland, J.M. Lymphocyte stimulation by concanavalin a studied by the fluorescent probe acridine orange. Cell Tissue Res. 217, 117–126 (1981). https://doi.org/10.1007/BF00233831

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