Quantification Endocytosis

Abstract

The study of nanoparticle internalization is significant for the purpose of its delivery and associated cell studies. Incorporation into endosomes is one of the common pathways of nanoparticle uptake by individual cells. However, very little is known, so far, about endosomal uptake, especially with respect to its quantification. Here we formulated malleable, polymer-free and emulsified nanosensors (nanogoo) comprising fluorescent dyes chromoionophore III and rhodamine. This formulation is selectively and effectively responsive to sodium in solution. Sodium ion fluxes are an integral component of action potentials and closely related to blood pressure regulation among other physiological functions. When this formulation is loaded onto the HEK-PN1 cells, endocytosis is the major pathway of uptake. Using this nanogoo, the endocytozed amount is quantified. Method is developed to indicate and differentiate the intracellular, extracellular and endosomal locations of the nanogoo with respect to the mammalian cell. Using confocal microscopy the spatial appearance of the nanogoo in solutions mimicking the three conditions is obtained. This information is then analyzed by a custom-written matlab code, which distinguishes the regions of interest in a quantitative representation. When extrapolated to images of spatial distribution after cell uptake, the presence of the nanogoo within the cell can be quantified. This cellular analysis would prove to be a landmark for sodium sensing and enable the detection of its fluxes across the cell membrane; which is the end goal of this ongoing research.