Abstract
Application of four fluorescence techniques to the measurement of the passive ionic permeability in biomembrane was demonstrated with the examples in the investigation of sarcoplasmic reticulum vesicles, in which fluorescence technique has widely been employed as well as other techniques, e. g., radio isotope tracer method, volume change-light scattering method, and conductance measurement using artifitial planar bilayer membrane.
First, the chlortetracycline method was described. The change in the amount of the intravesicular divalent cations can be followed by measuring the fluorescence of the dye, which increases upon binding to diamagnetic cations such as Ca2+ and Mg2+ on membrane surface. Secondly explained was the technique using anilinonaphthalenesulfonate, whose fluorescence change arises from its binding to the membrane that depends on the membrane surface potential, and indicates the change in the ionic compositions outside and inside the vesicles. Thirdly, application of the slow response of the potential-sensitive dyes was described. Permeability of ions can be determined from the amplitude and its dissipation rate of transmembrane diffusion potential due to ionic concentration gradient. Lastly, the fluorescence quenching method was shown to give the highest time resolution. The decrease of the fluorescence of the water soluble dyes entrapped in the vesicles due to the influx of ionic quenchers is analyzed to yield the ionic permeability of the vesicles.
