VPS45 Antibody

Immunocytochemistry/ Immunofluorescence: VPS45 Antibody [NB100-2431] -

mVps45 binding to Sx16 controls GLUT4 trafficking.HeLa cells expressing HA-GLUT4-GFP were transfected with plasmids encoding either HA-mVps45 (wild-type) or HA-mVps45-V107R (a mutant which prevents the interaction of the Sx16 N-terminus with mVps45 –see text). 48 h after transfection, cells were incubated in serum-free media for 2 h, fixed and cell surface GLUT4 immuno-stained using anti-HA (pseudo-coloured blue) prior to permeabilization. Subsequently, cells were permeabilised and stained using anti-mVps45 which detects both endogenous and over-expressed mVps45 (pseudo-coloured red; note that the use of HA-tagged mVps45 constructs and HA-tagged GLUT4 precluded this as a means to distinguish cells over-expressing mVps45 species). Signal from GFP is pseudo-coloured green. (A) Data from a typical experiment; white asterisk represent cells expressing higher than average anti-mVps45 immunoreactivity and which are therefore assumed to be over-expressing the indicated species. Data from a typical experiment is shown. (B) Levels of HA-GLUT4-GFP were not significantly altered upon over-expression of either wild-type or mVps45-V107R, which were expressed at similar levels (both are recognised by anti-HA); the approximate positions of molecular weight markers are shown (in kDa). (C) Quantification of the HA/GFP ratio from fields of cells such as those shown in (A). Fields of cells transfected with mVps45-V107R exhibited increased HA/GFP ratios compared to cells transfected with wild-type mVps45 (∗p < 0.001 ANOVA). Wild-type transfected cells were indistinguishable from non-transfected controls (ns; p = 0.33 ANOVA). Each point on the graph is from a single field of cells; data from three independent biological experiments is presented. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/37520260), licensed under a CC-BY license. Not internally tested by Novus Biologicals.