Measurement of mitochondrial volume independent of refractive index by light scattering

Using latex spheres of accurately determined dimensions, it was found that, although the scattered light intensity at an angle within the forward scattering lobe varies with the refractive index of the medium, the ratio of the scattered light intensity at a pair of angles is independent of the refractive index. This finding is consistent with calculations from the Mie theory by Hodkinson that the shape of the forward scattering lobe depends only on the size of the sphere and is independent of the refractive index of the sphere or of the suspending medium. This method was applied to the measurement of volume changes in mitochondria induced either by suspending the mitochondria in solutions of different osmolality or by addition of ionophores to stimulate ion uptake. The volume changes measured were independent of the refractive index of the suspending medial. It is shown that measuring the ratio of the scattered light intensity at selected pair of angles within the forward scattering lobe is a good method to measure the average size of the mitochondria.