Quantitation of lymphocyte intracellular free calcium signals using indo-1.
The calcium-responsive fluorescent dye indo-1 has been used in lymphocyte suspensions to measure changes in internal free calcium concentration, [Ca2+]i, in response to crosslinking of cell surface immunoglobulin. The quantitation of [Ca2+]i requires that indo-am ester used to load the cells be completely hydrolyzed to the indo-1 form inside the cells. This is shown to be greatly facilitated in the lymphocyte by the detergent Pluronic F-127. The quantitation of [Ca2+]i transients also requires an estimate of the fraction of the cells which contribute to the observed changes. The use of excessive amounts of intracellular dye can buffer [Ca2+]i transients and this effect has been used to estimate the size of the pool of calcium which is available for release when the B cell is stimulated by anti-immunoglobulin.
Owen
C S
CS
Department of Biochemistry, Jefferson Medical College, Philadelphia.
D000818:Animals; D002118:Calcium / Q000032:analysis* / Q000378:metabolism; D004952:Esters / Q000378:metabolism; D005260:Female; D005456:Fluorescent Dyes; D006801:Humans; D007211:Indoles; D008214:Lymphocytes / Q000032:analysis*; D051379:Mice; D011060:Poloxalene / Q000494:pharmacology; D013050:Spectrometry, Fluorescence; D013154:Spleen / Q000166:cytology
D004952:Esters; D005456:Fluorescent Dyes; D007211:Indoles; D011060:Poloxalene; C048960:indo-1; D002118:Calcium