Simultaneous determination of intracellular calcium concentration and histamine secretion in rat basophilic leukemia cells (RBL-2H3).
In rat basophilic leukemia cells (RBL-2H3), a tumor analogue of mast cells, the aggregation of IgE receptors initiates increase in the intracellular concentration of calcium ([Ca2+]i), monitored with the fluorescent Ca probe fura-2, and finally results in histamine secretion. In cell suspensions, however, the fluorescence gradually increases due to leakage and exocytosis of the dye. A superfusion system was developed to overcome these problems and [Ca2+]i was calculated from the ratio of fluorescence intensities at 505 nm of fura-2 excited at 340 and 380 nm. Histamine and beta-N-acetylglucosaminidase in granules are released during exocytosis, and both substances in the superfusates were determined simultaneously. This system is useful for studies on the relationships of cell stimulation, changes in second messengers, and final responses.
Maeyama
K
K
Department of Pharmacology I, Tohoku University School of Medicine, Sendai, Japan.
Sasaki
M
M
Watanabe
T
T
D000118:Acetylglucosaminidase / Q000378:metabolism; D000818:Animals; D000944:Antigens, Differentiation, B-Lymphocyte / Q000378:metabolism; D002118:Calcium / Q000378:metabolism*; D003600:Cytosol / Q000378:metabolism; D016257:Fura-2; D006636:Histamine Release; D015471:Leukemia, Basophilic, Acute; D008239:Lysine / Q000031:analogs & derivatives / Q000494:pharmacology; D051381:Rats; D011961:Receptors, Fc / Q000378:metabolism; D017455:Receptors, IgE; D013050:Spectrometry, Fluorescence; D014407:Tumor Cells, Cultured
D000944:Antigens, Differentiation, B-Lymphocyte; D011961:Receptors, Fc; D017455:Receptors, IgE; C007493:epsilon-dinitrophenyllysine; D000118:Acetylglucosaminidase; D008239:Lysine; D002118:Calcium; D016257:Fura-2