CD36 Antibody (1283D) - BSA Free

Immunocytochemistry/ Immunofluorescence: CD36 Antibody (1283D) - BSA Free [NBP2-54790]

Immunocytochemistry/Immunofluorescence: CD36 Antibody (1283D) [NBP2-54790] - HepG2 cells were fixed for 10 minutes using 10% formalin and then permeabilized for 5 minutes using 1X TBS + 0.5% Triton-X100. The cells were incubated with anti-CD36 (1283D) at 5.0 ug/ml overnight at 4C and detected with an anti-rabbit Dylight 488 (Green) at a 1:500 dilution. Alpha tubulin (DM1A) NB100-690 was used as a co-stain at a 1:1000 dilution and detected with an anti-mouse Dylight 550 (Red) at a 1:500 dilution. Nuclei were counterstained with DAPI (Blue). Cells were imaged using a 40X objective..

Western Blot: CD36 Antibody (1283D)BSA Free [NBP2-54790]

CD36-Antibody-1283D-BSA-Free-Western-Blot-NBP2-54790-img0024.jpg

Western Blot: CD36 Antibody (1283D) - BSA Free [NBP2-54790] -

Western Blot: CD36 Antibody (1283D) - BSA Free [NBP2-54790] - LYN phosphorylates DHHC5 & is required for the endocytosis of FAs.a 3T3-L1 adipocytes pretreated with serum-free medium for 4 h & PP2 (15 μM) for 1 h, followed by treatment with BSA or oleate (100 μM) for 5 min. Cells harvested & immunoprecipitated with anti-DHHC5 antibody to detect phosphorylation of Tyr91. b 3T3-L1 adipocytes transduced with indicated shRNA set up & treated with oleate for 5 min. Cells harvested to detect Tyr91 phosphorylation as in (a). c On day 0, HEK293T cells set up at 2.5 × 105 cells per 6-cm dish. On day 2, cells transfected with 0.5 μg of WT, Y91E, or Y91F of DHHC5-Flag/pCDH-puro and/or 0.5 μg LYN/pCDNA3.3. On day 3, cells harvested & immunoprecipitated with anti-Flag M2 beads to detect phosphorylation of DHHC5 (pY). Control & LYN knockdown adipocytes pretreated & treated with BSA or oleate (100 μM) for 1 h, followed by immunostaining (d), surface biotinylation (e), & Acyl-RAC (f) assays. Control & CD36 knockdown 3T3-L1 adipocytes pretreated & treated with oleate (100 μM) for indicated time (g) or 5 min (h). Cells harvested & subjected to immunoprecipitation of LYN to detect phosphorylation of LYN (Y396). i, j WT & Cd36−/− SVFs set up & subjected to FA uptake as in Fig. 3h, i, except that cells pretreated with PP2 (15 μM). Each value represents mean ± SEM obtained from 20 cells. Two-sided Student’s t test was performed between DMSO & PP2 treated cells. k WT & Cd36−/− adipocytes set up & subjected to 3H-oleate uptake as in Fig. 3j, except that cells pretreated with PP2 (15 μM). Each value represents mean ± SEM obtained from 3samples. Two-sided Student’s t test was performed between DMSO & PP2 treated cells. These experiments repeated at least twice. Source data provided as a Source Data file. Image collected & cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/32958780), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Western Blot: CD36 Antibody (1283D) - BSA Free [NBP2-54790] -

Western Blot: CD36 Antibody (1283D) - BSA Free [NBP2-54790] - LYN phosphorylates DHHC5 & is required for the endocytosis of FAs.a 3T3-L1 adipocytes pretreated with serum-free medium for 4 h & PP2 (15 μM) for 1 h, followed by treatment with BSA or oleate (100 μM) for 5 min. Cells harvested & immunoprecipitated with anti-DHHC5 antibody to detect phosphorylation of Tyr91. b 3T3-L1 adipocytes transduced with indicated shRNA set up & treated with oleate for 5 min. Cells harvested to detect Tyr91 phosphorylation as in (a). c On day 0, HEK293T cells set up at 2.5 × 105 cells per 6-cm dish. On day 2, cells transfected with 0.5 μg of WT, Y91E, or Y91F of DHHC5-Flag/pCDH-puro and/or 0.5 μg LYN/pCDNA3.3. On day 3, cells harvested & immunoprecipitated with anti-Flag M2 beads to detect phosphorylation of DHHC5 (pY). Control & LYN knockdown adipocytes pretreated & treated with BSA or oleate (100 μM) for 1 h, followed by immunostaining (d), surface biotinylation (e), & Acyl-RAC (f) assays. Control & CD36 knockdown 3T3-L1 adipocytes pretreated & treated with oleate (100 μM) for indicated time (g) or 5 min (h). Cells harvested & subjected to immunoprecipitation of LYN to detect phosphorylation of LYN (Y396). i, j WT & Cd36−/− SVFs set up & subjected to FA uptake as in Fig. 3h, i, except that cells pretreated with PP2 (15 μM). Each value represents mean ± SEM obtained from 20 cells. Two-sided Student’s t test was performed between DMSO & PP2 treated cells. k WT & Cd36−/− adipocytes set up & subjected to 3H-oleate uptake as in Fig. 3j, except that cells pretreated with PP2 (15 μM). Each value represents mean ± SEM obtained from 3samples. Two-sided Student’s t test was performed between DMSO & PP2 treated cells. These experiments repeated at least twice. Source data provided as a Source Data file. Image collected & cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/32958780), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Western Blot: CD36 Antibody (1283D) - BSA Free [NBP2-54790] -

Western Blot: CD36 Antibody (1283D) - BSA Free [NBP2-54790] - FAs trigger internalization of CD36.a, b On day 8 of differentiation, 3T3-L1 adipocytes were pretreated with serum-free medium for 4 h, & then treated with BSA-conjugated oleate (100 μM) for indicated time. a One set of cells was subjected to immunostaining with anti-CD36 & anti-CAV1 antibodies. LipidTOX was used to label lipid droplets. Images were taken under a Zeiss LSM-780 microscope in a 3D Z-stack mode & reconstructed using Imaris 9.2.0. b The other set of cells was subjected to surface biotinylation assay & blotted with indicated antibodies. c, d On day 4 of differentiation, 3T3-L1 cells were infected with lentivirus encoding scrambled shRNA or shRNAs against CD36 or CAV1. On day 5, cells were selected with 5 μg/ml puromycin. On day 8, cells were pretreated as in (a) & treated with oleate (100 μM) for 4 h, followed by immunostaining with anti-CD36 & anti-CAV1 antibodies (c), or surface biotinylation assay (d). e, f 3T3-L1 adipocytes were pretreated as in (a) & treated with BSA-conjugated FAs with different chain lengths or saturation (100 μM) for 4 h. Cells were subjected to immunostaining with anti-CD36 antibody (e), or surface biotinylation assay (f). After oleate treatment for 4 h, 3T3-L1 adipocytes were switched to serum-free medium for indicated time & harvested for immunostaining (g) & surface biotinylation (h). The scale bars were as indicated in each figure. These experiments were repeated at least three times. Source data are provided as a Source Data file. Image collected & cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/32958780), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Western Blot: CD36 Antibody (1283D) - BSA Free [NBP2-54790] -

Western Blot: CD36 Antibody (1283D) - BSA Free [NBP2-54790] - FAs trigger internalization of CD36.a, b On day 8 of differentiation, 3T3-L1 adipocytes were pretreated with serum-free medium for 4 h, & then treated with BSA-conjugated oleate (100 μM) for indicated time. a One set of cells was subjected to immunostaining with anti-CD36 & anti-CAV1 antibodies. LipidTOX was used to label lipid droplets. Images were taken under a Zeiss LSM-780 microscope in a 3D Z-stack mode & reconstructed using Imaris 9.2.0. b The other set of cells was subjected to surface biotinylation assay & blotted with indicated antibodies. c, d On day 4 of differentiation, 3T3-L1 cells were infected with lentivirus encoding scrambled shRNA or shRNAs against CD36 or CAV1. On day 5, cells were selected with 5 μg/ml puromycin. On day 8, cells were pretreated as in (a) & treated with oleate (100 μM) for 4 h, followed by immunostaining with anti-CD36 & anti-CAV1 antibodies (c), or surface biotinylation assay (d). e, f 3T3-L1 adipocytes were pretreated as in (a) & treated with BSA-conjugated FAs with different chain lengths or saturation (100 μM) for 4 h. Cells were subjected to immunostaining with anti-CD36 antibody (e), or surface biotinylation assay (f). After oleate treatment for 4 h, 3T3-L1 adipocytes were switched to serum-free medium for indicated time & harvested for immunostaining (g) & surface biotinylation (h). The scale bars were as indicated in each figure. These experiments were repeated at least three times. Source data are provided as a Source Data file. Image collected & cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/32958780), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Western Blot: CD36 Antibody (1283D) - BSA Free [NBP2-54790] -

Western Blot: CD36 Antibody (1283D) - BSA Free [NBP2-54790] - FAs trigger internalization of CD36.a, b On day 8 of differentiation, 3T3-L1 adipocytes were pretreated with serum-free medium for 4 h, & then treated with BSA-conjugated oleate (100 μM) for indicated time. a One set of cells was subjected to immunostaining with anti-CD36 & anti-CAV1 antibodies. LipidTOX was used to label lipid droplets. Images were taken under a Zeiss LSM-780 microscope in a 3D Z-stack mode & reconstructed using Imaris 9.2.0. b The other set of cells was subjected to surface biotinylation assay & blotted with indicated antibodies. c, d On day 4 of differentiation, 3T3-L1 cells were infected with lentivirus encoding scrambled shRNA or shRNAs against CD36 or CAV1. On day 5, cells were selected with 5 μg/ml puromycin. On day 8, cells were pretreated as in (a) & treated with oleate (100 μM) for 4 h, followed by immunostaining with anti-CD36 & anti-CAV1 antibodies (c), or surface biotinylation assay (d). e, f 3T3-L1 adipocytes were pretreated as in (a) & treated with BSA-conjugated FAs with different chain lengths or saturation (100 μM) for 4 h. Cells were subjected to immunostaining with anti-CD36 antibody (e), or surface biotinylation assay (f). After oleate treatment for 4 h, 3T3-L1 adipocytes were switched to serum-free medium for indicated time & harvested for immunostaining (g) & surface biotinylation (h). The scale bars were as indicated in each figure. These experiments were repeated at least three times. Source data are provided as a Source Data file. Image collected & cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/32958780), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Western Blot: CD36 Antibody (1283D) - BSA Free [NBP2-54790] -

Western Blot: CD36 Antibody (1283D) - BSA Free [NBP2-54790] - FAs trigger internalization of CD36.a, b On day 8 of differentiation, 3T3-L1 adipocytes were pretreated with serum-free medium for 4 h, & then treated with BSA-conjugated oleate (100 μM) for indicated time. a One set of cells was subjected to immunostaining with anti-CD36 & anti-CAV1 antibodies. LipidTOX was used to label lipid droplets. Images were taken under a Zeiss LSM-780 microscope in a 3D Z-stack mode & reconstructed using Imaris 9.2.0. b The other set of cells was subjected to surface biotinylation assay & blotted with indicated antibodies. c, d On day 4 of differentiation, 3T3-L1 cells were infected with lentivirus encoding scrambled shRNA or shRNAs against CD36 or CAV1. On day 5, cells were selected with 5 μg/ml puromycin. On day 8, cells were pretreated as in (a) & treated with oleate (100 μM) for 4 h, followed by immunostaining with anti-CD36 & anti-CAV1 antibodies (c), or surface biotinylation assay (d). e, f 3T3-L1 adipocytes were pretreated as in (a) & treated with BSA-conjugated FAs with different chain lengths or saturation (100 μM) for 4 h. Cells were subjected to immunostaining with anti-CD36 antibody (e), or surface biotinylation assay (f). After oleate treatment for 4 h, 3T3-L1 adipocytes were switched to serum-free medium for indicated time & harvested for immunostaining (g) & surface biotinylation (h). The scale bars were as indicated in each figure. These experiments were repeated at least three times. Source data are provided as a Source Data file. Image collected & cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/32958780), licensed under a CC-BY license. Not internally tested by Novus Biologicals.

Western Blot: CD36 Antibody (1283D) - BSA Free [NBP2-54790] -

Western Blot: CD36 Antibody (1283D) - BSA Free [NBP2-54790] - CD36-mediated caveolar endocytosis transports FAs into cells.a–c 3T3-L1 adipocytes were pretreated with serum-free medium for 4 h, & then treated with BSA-conjugated PacFA (50 μM) for 20 min, followed by UV crosslinking on ice for 30 min. a, b Cells were subjected to click chemistry using an N3-Alexa Fluro 488, & immunostained with anti-CD36 antibody. Colocalization of PacFA & CD36 was quantified from 24 cells over three independent experiments & plotted in (b). The value represents mean ± SEM. c Cells were lysed & subjected to click chemistry assay using N3-biotin. PacFA-labeled proteins were captured with streptavidin beads & subjected to western blot using anti-CD36 & anti-FABP4 antibodies. d WT, Cav1−/−, Cd36−/−, & Cav1−/−;Cd36−/− SVFs were isolated & differentiated into adipocytes & treated with 100 μM 3H-oleate (specific activity, 2268 dpm/nmol) for 1 h. Lipid fractions were extracted from the cells & subjected to scintillation counting. The radioactive counting was normalized to protein content. Each value represents mean ± SEM obtained from three samples. Two-sided Student’s t test was performed between WT & each of the knockout cells. These experiments were repeated twice. Source data are provided as a Source Data file. Image collected & cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/32958780), licensed under a CC-BY license. Not internally tested by Novus Biologicals.