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![Western Blot: Rabex5 Antibody [NBP1-49938] - Detection of Human and Mouse RABGEF1 by Western Blot (h&m) and Immunoprecipitation (h). Samples: Whole cell lysate from HeLa (5, 15 and 50 mcg for WB; 1 mg for IP, 20% of IP loaded), 293T (T; 50 mcg) and mouse NIH3T3 (M; 50 mcg) cells. Antibodies: Affinity purified rabbit anti-RABGEF1 antibody used for WB at 0.1 mcg/ml (A) and 1 mcg/ml (B) and used for IP at 3 mcg/mg lysate. RABGEF1 was also immunoprecipitated by rabbit anti-RABGEF1 antibody which recognizes a downstream epitope. For blotting immunoprecipitated RABGEF1 was used. Detection: Chemiluminescence with exposure times of 3 minutes (A) and 30 seconds (B).](http://images.novusbio.com/fullsize/Rabex5-Antibody-Western-Blot-NBP1-49938-img0001.jpg "<b>Independent Antibodies Validation and Biological Strategies Validation.</b>Western Blot: Rabex5 Antibody [NBP1-49938] - Detection of Human and Mouse RABGEF1 by Western Blot (h&m) and Immunoprecipitation (h). Samples: Whole cell lysate from HeLa (5, 15 and 50 mcg for WB; 1 mg for IP, 20% of IP loaded), 293T (T; 50 mcg) and mouse NIH3T3 (M; 50 mcg) cells. Antibodies: Affinity purified rabbit anti-RABGEF1 antibody used for WB at 0.1 mcg/ml (A) and 1 mcg/ml (B) and used for IP at 3 mcg/mg lysate. RABGEF1 was also immunoprecipitated by rabbit anti-RABGEF1 antibody which recognizes a downstream epitope. For blotting immunoprecipitated RABGEF1 was used. Detection: Chemiluminescence with exposure times of 3 minutes (A) and 30 seconds (B).")
![<b>Independent Antibodies Validation and Biological Strategies Validation.</b>Western Blot: Rabex5 Antibody [NBP1-49938] - RABGEF1 is important for mitochondrial clearance. YFP-Parkin stably expressing WT and RABGEF1-mAID HCT116 cells pre-treated with IAA were treated with valinomycin for the indicated times. Total cell lysates were analyzed by immunoblotting. Image collected and cropped by CiteAb from the following publication (//elifesciences.org/articles/31326), licensed under a CC-BY license.](http://images.novusbio.com/fullsize/Rabex5-Antibody-Western-Blot-NBP1-49938-img0005.jpg)
![Immunohistochemistry-Paraffin: Rabex5 Antibody [NBP1-49938] - Section of human lung carcinoma. Antibody: Affinity purified rabbit anti- RABGEF1 used at a dilution of 1:1,000 (1ug/ml). Detection: DAB](http://images.novusbio.com/fullsize/Rabex5-Antibody-Immunohistochemistry-Paraffin-NBP1-49938-img0004.jpg)
![Western Blot: Rabex5 Antibody [NBP1-49938] - RABGEF1 recruitment to mitochondria during mitophagy.(A) The indicated cells were treated with DMSO or valinomycin for 3 hr followed by immunostaining. Bars, 10 μm. Graphs for quantification of RABGEF1 recruitment to mitochondria were shown below the images. None, partial & complete denote that GFP-mRABGEF1 signals were overlapped with no, some of, & all mitochondria, respectively. The error bars represent mean ±SE & over 100 cells were counted in each of three separate wells. (B) WT & TBC1D15/17 DKO HCT116 cells stably expressing mCherry-Parkin & GFP-mRABGEF1 were treated with DMSO or valinomycin for 3 hr. GFP-mRABGEF1 signals were enhanced by immunostaining with anti-GFP antibody. Bars, 10 μm. (C) Total cell lysates in (B) were analyzed by immunoblotting. * & # denote ubiquitinated forms & truncated forms, respectively.10.7554/eLife.31326.031Figure 8—figure supplement 1—source data 1.This excel file contains quantification of RABGEF1 (WT & Y26A/A58D mutant) recruitment to mitochondria in HCT116 (WT & TBC1D15/17 DKO) cells.This excel file contains quantification of RABGEF1 (WT & Y26A/A58D mutant) recruitment to mitochondria in HCT116 (WT & TBC1D15/17 DKO) cells. Image collected & cropped by CiteAb from the following publication (//elifesciences.org/articles/31326), licensed under a CC-BY license. Not internally tested by Novus Biologicals.](http://images.novusbio.com/fullsize/nbp1-49938_rabbit-polyclonal-rabex5-antibody-310202416182628.jpg)
![Western Blot: Rabex5 Antibody [NBP1-49938] - RABGEF1 is important for mitochondrial clearance.(A) WT & RABGEF1-mAID HCT116 cells were treated with or without IAA for 16 hr. Total cell lysates were analyzed by immunoblotting. (B) Quantification of Parkin recruitment to mitochondria in WT & RABGEF1-mAID HCT116 cells after 3 hr of valinomycin treatment. Partial & complete denote that YFP-Parkin signals were overlapped with some of & all mitochondria, respectively. (C) YFP-Parkin stably expressing WT & RABGEF1-mAID HCT116 cells pre-treated with IAA were treated with valinomycin for the indicated times. Total cell lysates were analyzed by immunoblotting. (D) WT & RABGEF1-mAID HCT116 cells stably expressing YFP-Parkin & mt-mKeima were treated with IAA for 16 hr followed by DMSO or OAQ for 6 hr & subjected to FACS analysis. Plots are representative of n = 3 experiments. (E) Quantification of mitophagy in (D). Error bars represent mean ±SE of three independent experiments. Statistical differences were determined by student’s t-test. *p<0.05.10.7554/eLife.31326.035Figure 9—source data 1.Quantification of YFP-Parkin recruitment to mitochondria in RABGEF1-mAID HCT116 & the corresponding WT cells during mitophagy.10.7554/eLife.31326.036Figure 9—source data 2.Quantification of mitophagy using mt-mKeima & FACS analysis.Quantification of YFP-Parkin recruitment to mitochondria in RABGEF1-mAID HCT116 & the corresponding WT cells during mitophagy.Quantification of mitophagy using mt-mKeima & FACS analysis. Image collected & cropped by CiteAb from the following publication (//elifesciences.org/articles/31326), licensed under a CC-BY license. Not internally tested by Novus Biologicals.](http://images.novusbio.com/fullsize/nbp1-49938_rabbit-polyclonal-rabex5-antibody-310202416182635.jpg)
![Western Blot: Rabex5 Antibody [NBP1-76548] - Analysis in human liver tissue lysate with antibody at (A) 1 and (B) 2 ug/mL.](http://images.novusbio.com/fullsize/Rabex5-Antibody-Western-Blot-NBP1-76548-img0004.jpg "Western Blot: Rabex5 Antibody [NBP1-76548] - Analysis in human liver tissue lysate with antibody at (A) 1 and (B) 2 ug/mL.")
![Immunohistochemistry: Rabex5 Antibody - BSA Free [NBP1-76548] - Immunohistochemistry of Rabex5 in mouse brain tissue with Rabex5 antibody at 5 ug/mL.](http://images.novusbio.com/fullsize/nbp1-76548_rabbit-polyclonal-rabex5-antibody-27112024200411.jpg)
![Immunocytochemistry/ Immunofluorescence: Rabex5 Antibody - BSA Free [NBP1-76548] - Immunofluorescence of Rabex5 in mouse brain tissue with Rabex5 antibody at 20 ug/mL.](http://images.novusbio.com/fullsize/nbp1-76548_rabbit-polyclonal-rabex5-antibody-27112024192366.jpg)
![Western Blot: Rabex5 Antibody [H00027342-B01P] - Analysis of RABGEF1 expression in human liver.](http://images.novusbio.com/fullsize/Rabex5-Antibody-Western-Blot-H00027342-B01P-img0003.jpg "Western Blot: Rabex5 Antibody [H00027342-B01P] - Analysis of RABGEF1 expression in human liver.")
![Western Blot: Rabex5 Antibody [H00027342-B01P] - Analysis of RABGEF1 expression in transfected 293T cell line by RABGEF1 polyclonal antibody. Lane 1: RABGEF1 transfected lysate(54.01 KDa). Lane 2: Non-transfected lysate.](http://images.novusbio.com/fullsize/Rabex5-Antibody-Western-Blot-H00027342-B01P-img0004.jpg)
![Western Blot: Rabex5 Overexpression Lysate (Adult Normal) [NBL1-15095] Left-Empty vector transfected control cell lysate (HEK293 cell lysate); Right -Over-expression Lysate for Rabex5.](http://images.novusbio.com/fullsize/Rabex5-Overexpression-Lysate-Adult-Normal-Western-Blot-NBL1-15095-img0002.jpg "Western Blot: Rabex5 Overexpression Lysate (Adult Normal) [NBL1-15095] Left-Empty vector transfected control cell lysate (HEK293 cell lysate); Right -Over-expression Lysate for Rabex5.")
![Immunocytochemistry/Immunofluorescence: Rabex5 Antibody [NBP2-97267] - Immunofluorescence staining of Rabex5 in U251MG cells. Cells were fixed with 4% PFA, permeabilzed with 0.1% Triton X-100 in PBS, blocked with 10% serum, and incubated with rabbit anti-Human Rabex5 polyclonal antibody (dilution ratio 1:200) at 4C overnight. Then cells were stained with the Alexa Fluor(R)488-conjugated Goat Anti-rabbit IgG secondary antibody (green) and counterstained with DAPI (blue).Positive staining was localized to Cytoplasm.](http://images.novusbio.com/fullsize/Rabex5-Antibody-Immunocytochemistry-Immunofluorescence-NBP2-97267-img0001.jpg "Immunocytochemistry/Immunofluorescence: Rabex5 Antibody [NBP2-97267] - Immunofluorescence staining of Rabex5 in U251MG cells. Cells were fixed with 4% PFA, permeabilzed with 0.1% Triton X-100 in PBS, blocked with 10% serum, and incubated with rabbit anti-Human Rabex5 polyclonal antibody (dilution ratio 1:200) at 4C overnight. Then cells were stained with the Alexa Fluor(R)488-conjugated Goat Anti-rabbit IgG secondary antibody (green) and counterstained with DAPI (blue).Positive staining was localized to Cytoplasm.")