Water channel efficiency & stopped flowLatest updated: August 26, 2020
Aquaporins are important and well-studied water channel membrane proteins. They form pores in the membranes of biological cells, facilitating the transport of water between cells. A particularly smart application of stopped-flow is the fast-tracking of transient kinetics of water and solute transport across the membrane of small vesicles.
Such a reaction can be generated by mixing purified membrane vesicles with a hyper-osmotic solution. The imposed concentration gradient induces a fast flow of water out of the vesicles. As a consequence, the volume of the vesicles is reduced and the concentration of the inner medium increases. This kinetics shrinkage will stop when the concentration of the inner medium has reached that of the surrounding one. The rapid change in the size of the vesicles can be detected by means of the scattered light intensity at 90°. The intensity of the light scattering signal will increase when the vesicle size decreases. Alternatively, vesicles can be loaded with a fluorescent dye with concentration-dependent quenching properties.
The reaction rates of the swelling or shrinking kinetics give direct information about cell permeability, but important experimental considerations should be considered for a successful experiment: which wavelength to use? How fast should I inject solutions? How much sample do I need?
An application note is available (Application Note #35) which outlines all such considerations as well as explaining in detail why BioLogic stopped-flow mixers are ideal instruments for membrane transport kinetics.
For more information, please see Application note #35