Intracellular Ion Measurement Protocols

This protocol describes a method to assess concentrations of free cytoplasmic calcium (Ca2+) in cultured RAW 264.7 cells. This objective is accomplished with the Ca2+-sensitive fluorescent dye, fluo-3, which permeates cells as an ester and is hydrolyzed in the cell to its Ca2+-sensitive acidic form. Fluorescence is measured over time with adherent cells that have been washed free of extracellular dye. - [Read Assay of Intracellular Free Calcium in RAW 264.7 Cells for Ligand Screen Protocol]

This protocol describes a method to assess concentrations of free cytoplasmic calcium, [Ca2+]i, in cultured adherent RAW 264.7 cells, using a 96- well plate format. This objective is accomplished by using the Ca2+-sensitive fluorescent dye, fluo-3, which permeates cell membranes as an ester and is hydrolyzed in the cell to its Ca2+-sensitive acidic form. Fluorescence for the adherent cells is measured over time by using a bottom read of a 96-well plate, with cells that have been washed. - [Read Assay of Intracellular Free Calcium in RAW 264.7 Cells Loaded with Fluo-3 Protocol]

This protocol describes a method to assess concentrations of free cytoplasmic calcium, [Ca2+]i , in cultured adherent RAW 264.7 cells, using a 96- well plate format. This objective is accomplished by using the Ca2+-sensitive fluorescent dye, fura-2, which permeates cell membranes as an ester and is hydrolyzed in the cell to its Ca2+-sensitive acidic form. - [Read Assay of Intracellular Free Calcium in RAW 264.7 Cells Loaded with Fura-2 Protocol]

This protocol describes a method to assess concentrations of free cytoplasmic calcium (Ca2+) in mouse splenic B cells in the absence and presence of ligands for cell surface receptors. This objective is accomplished with the Ca2+-sensitive fluorescent dye, Fluo-3, which permeates cells as an ester and is hydrolyzed in the cell to its Ca2+- sensitive acidic form. - [Read Assay of Intracellular Free Calcium in Suspended B Cells Protocol]

Protocol for calcium flux analysis for flow cytometry. - [Read Calcium Flux Analysis Protocol for Flow Cytometry]

Protocol for calcium flux using Indo-1 on the FACSVantage. - [Read Calcium Flux Using Indo-1 on the FACSVantage Protocol]

Protocol for Calcium Flux: Indo-1 loading and sample staining procedure for simultaneous measurement of intracellular Ca2+. Includes: Preparation of INDO-1; Preparation of Ionomycin. - [Read Calcium Flux: Indo-1 Loading and Sample Staining Procedure for Simultaneous Measurement of Intra Ca2]

This protocol describes a method to assess concentrations of free cytoplasmic calcium, [Ca2+]i, for cultured adherent RAW 264.7 cells in an 8-well coverglass. This objective is accomplished using the Ca2+-sensitive fluorescent dye, fura-2 acetoxymethyl (AM), which permeates cell membranes as an ester and is hydrolyzed in the cell to its Ca2+-sensitive acidic form. - [Read Live Single-Cell Fura-2 Measurements to Determine the Intracellular Free Calcium Protocol]

Describes flow cytometric protocols using the dyes Indo-1 AM, Fluo-3, and Fura Red AM to measure intracellular calcium concentration. Support protocols detail the use of calcium buffers to calibrate a flow cytometric calcium assay, and methods to facilitate dye loading; an alternate protocol describes the use of a spectrofluorimeter to measure intracellular calcium for those investigators without access to a flow cytometer. - [Read Measurement of Intracellular Ions by Flow Cytometry Protocol]

INDO-1 loading and sample staining protocol for simultaneous measurement of intracellular CA2+ and cell surface antigen expression. Includes: Preparation of INDO-1; Preparation of Ionomycin. - [Read Simultaneous Measurement of Intracellular CA2+ and Cell Surface Antigen Expression Protocol]

Protocol for sodium and potassium analysis by flow cytometry. Includes: Preparation of the dyes; Loading of the dyes; Flow Cytometric Analysis. - [Read Sodium and Potassium Analysis by Flow Cytometry Protocol]