glut4 primary antibody  (Proteintech)

Journal: The Journal of Biological Chemistry

Article Title: Secreted EMC10 inhibits muscle GLUT4 activity and glucose uptake in mice

doi: 10.1016/j.jbc.2025.110296

Figure Lengend Snippet: scEMC10 inhibits glucose uptake and GLUT4 expression in skeletal muscle cells . A , L6-GLUT4myc myotubes were treated with 1 μg/ml recombinant scEMC10 protein for various times as indicated in the presence of 100 μmol/l 2-NBDG. 2-NBDG uptake into the myotubes was measured at the FITC channel by a flow cytometer. Meanwhile, 2-NBDG uptake stimulated by 100 nM insulin for 30 min was used as a positive control. B , L6-GLUT4myc myotubes were treated with 100 nM insulin in combination with or without 1 μg/ml recombinant scEMC10 protein for the times as indicated. 2-NBDG uptake into the myotubes was measured. C–F , C2C12 myotubes were treated with recombinant scEMC10 protein at different concentrations as indicated for 12 h, and then the levels of GLUT4 mRNA ( C ) and protein ( E ) were determined by quantitative PCR and Western blot, respectively. C2C12 myotubes were treated with 1 μg/ml recombinant scEMC10 protein for various times as indicated, and then the levels of GLUT4 mRNA ( D ) and protein ( F ) were determined by quantitative PCR and Western blot, respectively. All cell culture experiments were repeated three to four times. All data are presented with means ± SD. In experiments where samples were prepared in several batches, the control conditions were set at 100% in individual experiments and therefore had no error estimates. Statistical analyses were performed using unpaired two-tailed Student's t test, and significant differences were indicated with p values. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, and ∗∗∗∗ p < 0.0001. 2-NBDG, 2-[ N -(7-nitrobenz-2-oxa-1,3-diazol-4-yl) amino]-2-deoxy- d -glucose; scEMC10, secreted isoform of endoplasmic reticulum membrane protein complex subunit 10.

Article Snippet: Then the cells were fixed, nonpermeabilized, or permeabilized with 0.1% (v/v) Triton X-100, blocked with 10% goat serum (Sango Biotech), and incubated with GLUT4 primary antibody (66846-1-Ig; Proteintech) overnight at 4 °C.

Techniques: Expressing, Recombinant, Flow Cytometry, Positive Control, Real-time Polymerase Chain Reaction, Western Blot, Cell Culture, Control, Two Tailed Test, Membrane

Journal: The Journal of Biological Chemistry

Article Title: Secreted EMC10 inhibits muscle GLUT4 activity and glucose uptake in mice

doi: 10.1016/j.jbc.2025.110296

Figure Lengend Snippet: scEMC10 suppresses GLUT4 translocation in L6-GLUT4myc myoblasts . A and B , L6-GLUT4myc myoblasts grown on slides were treated with 1 μg/ml recombinant scEMC10 protein, 100 nmol/l insulin, or scEMC10 in combination with insulin for 30 min, respectively, and then GLUT4 distribution was assessed using immunofluorescent staining in either permeabilized ( A ) or nonpermeabilized ( B ) cells. In B , white arrows indicate ring-like staining of GLUT4. Scale bar represents 50 μm ( A ) and 20 μm ( B ). All cell culture experiments were repeated three to four times. scEMC10, secreted isoform of endoplasmic reticulum membrane protein complex subunit 10.

Article Snippet: Then the cells were fixed, nonpermeabilized, or permeabilized with 0.1% (v/v) Triton X-100, blocked with 10% goat serum (Sango Biotech), and incubated with GLUT4 primary antibody (66846-1-Ig; Proteintech) overnight at 4 °C.

Techniques: Translocation Assay, Recombinant, Staining, Cell Culture, Membrane

Journal: The Journal of Biological Chemistry

Article Title: Secreted EMC10 inhibits muscle GLUT4 activity and glucose uptake in mice

doi: 10.1016/j.jbc.2025.110296

Figure Lengend Snippet: scEMC10 inhibits muscle glucose uptake and GLUT4 expression in mice. A and B , representative images ( A ) and quantifications ( B ) of 18 F-FDG uptake into gastrocnemius determined using PET–CT scan in Emc10 KO (n = 3) or WT mice (n = 4) fed an HFD. C and D , levels of GLUT4 mRNA ( C ) and protein ( D ) were determined in gastrocnemius tissue of the mice (n = 5–6) indicated by quantitative PCR and Western blot, respectively. E and F , representative images ( E ) and quantifications ( F ) of 18 F-FDG uptake into gastrocnemius determined using PET–CT scan in mice intraperitoneally injected with recombinant mouse scEMC10 protein (6 mg per kg body weight) (n = 5) or vehicle (human IgG1 Fc) (n = 6). G , GLUT4 protein levels were measured in gastrocnemius tissue of mice intraperitoneally injected with recombinant mouse scEMC10 protein (n = 5) or vehicle (n = 5) by Western blot. All data are presented with means ± SD. Statistical analyses were performed using unpaired two-tailed Student's t test, and significant differences were indicated with p values. ∗ p < 0.05, ∗∗∗ p < 0.001. 18 F-FDG, 18 F-fluorodeoxyglucose; CT, computed tomography; HFD, high-fat diet; scEMC10, secreted isoform of endoplasmic reticulum membrane protein complex subunit 10.

Article Snippet: Then the cells were fixed, nonpermeabilized, or permeabilized with 0.1% (v/v) Triton X-100, blocked with 10% goat serum (Sango Biotech), and incubated with GLUT4 primary antibody (66846-1-Ig; Proteintech) overnight at 4 °C.

Techniques: Expressing, Positron Emission Tomography-Computed Tomography, Real-time Polymerase Chain Reaction, Western Blot, Injection, Recombinant, Two Tailed Test, Computed Tomography, Membrane

Journal: The Journal of Biological Chemistry

Article Title: Secreted EMC10 inhibits muscle GLUT4 activity and glucose uptake in mice

doi: 10.1016/j.jbc.2025.110296

Figure Lengend Snippet: Antibody neutralization of scEMC10 enhances GLUT4 expression and glucose uptake in mouse skeletal muscle . A , representative images and quantifications of 18 F-FDG uptake into gastrocnemius determined using PET–CT scan in HFD-fed mice intraperitoneally injected with scEMC10-neutralizing antibody 4C2 (3 mg per kg body weight) (n = 5) or control IgG (n = 5). B and C , levels of GLUT4 mRNA ( B ) and protein ( C ) were determined in gastrocnemius tissue of the mice (n = 3–4) indicated by quantitative PCR and Western blot, respectively. D , immunofluorescent staining of GLUT4 and dystrophin in gastrocnemius tissue of the mice. E , quantitative analysis of membraneous GLUT4 immunofluorescence normalized to dystrophin in gastrocnemius muscle tissue of mice using ImageJ (n = 3–4). All data are presented with means ± SD. Statistical analyses were performed using unpaired two-tailed Student's t test, and significant differences were indicated with p values. ∗ p < 0.05. CT, computed tomography; 18 F-FDG, 18 F-fluorodeoxyglucose; HFD, high-fat diet; scEMC10, secreted isoform of endoplasmic reticulum membrane protein complex subunit 10.

Article Snippet: Then the cells were fixed, nonpermeabilized, or permeabilized with 0.1% (v/v) Triton X-100, blocked with 10% goat serum (Sango Biotech), and incubated with GLUT4 primary antibody (66846-1-Ig; Proteintech) overnight at 4 °C.

Techniques: Neutralization, Expressing, Positron Emission Tomography-Computed Tomography, Injection, Control, Real-time Polymerase Chain Reaction, Western Blot, Staining, Immunofluorescence, Two Tailed Test, Computed Tomography, Membrane

Journal: The Journal of Biological Chemistry

Article Title: Secreted EMC10 inhibits muscle GLUT4 activity and glucose uptake in mice

doi: 10.1016/j.jbc.2025.110296

Figure Lengend Snippet: Inhibition of scEMC10 reverses its inhibitory effect on GLUT4 translocation in L6-GLUT4myc myoblasts . A and B , L6-GLUT4myc myoblasts grown on slides were treated with 100 nmol/l insulin, 1 μg/ml recombinant scEMC10 protein, and 4 μg/ml scEMC10-neutralizing antibody 4C2 as indicated for 30 min, respectively, and then GLUT4 distribution was assessed using immunofluorescent staining in either permeabilized ( A ) or nonpermeabilized ( B ) cells. In B , white arrows indicate ring-like staining of GLUT4. Scale bar represents 50 μm ( A ) and 20 μm ( B ). All cell culture experiments were repeated three to four times. scEMC10, secreted isoform of endoplasmic reticulum membrane protein complex subunit 10.

Article Snippet: Then the cells were fixed, nonpermeabilized, or permeabilized with 0.1% (v/v) Triton X-100, blocked with 10% goat serum (Sango Biotech), and incubated with GLUT4 primary antibody (66846-1-Ig; Proteintech) overnight at 4 °C.

Techniques: Inhibition, Translocation Assay, Recombinant, Staining, Cell Culture, Membrane

Journal: The Journal of Biological Chemistry

Article Title: Secreted EMC10 inhibits muscle GLUT4 activity and glucose uptake in mice

doi: 10.1016/j.jbc.2025.110296

Figure Lengend Snippet: scEMC10 regulates expression of transcriptional factors of GLUT4 in C2C12 myotubes . A–E , C2C12 myotubes were treated with 1 μg/ml recombinant scEMC10 protein for various times as indicated, and then mRNA levels of MEF2D ( A ), MyoD ( B ), PGC-1α ( C ), KLF15 ( D ), and HDAC5 ( E ) were measured using quantitative PCR. F , MEF2D mRNA levels were assessed in gastrocnemius tissue of mice intraperitoneally injected with scEMC10-neutralizing antibody 4C2 (3 mg per kg body weight) (n = 4) or control IgG (n = 4). All cell culture experiments were repeated three to four times. All data are presented with means ± SD. In experiments where samples were prepared in several batches, the control conditions were set at 100% in individual experiments and therefore had no error estimates. Statistical analyses were performed using unpaired two-tailed Student's t test, and significant differences were indicated with p values. ∗ p < 0.05, ∗∗ p < 0.01, and ∗∗∗ p < 0.001. scEMC10, secreted isoform of endoplasmic reticulum membrane protein complex subunit 10.

Article Snippet: Then the cells were fixed, nonpermeabilized, or permeabilized with 0.1% (v/v) Triton X-100, blocked with 10% goat serum (Sango Biotech), and incubated with GLUT4 primary antibody (66846-1-Ig; Proteintech) overnight at 4 °C.

Techniques: Expressing, Recombinant, Real-time Polymerase Chain Reaction, Injection, Control, Cell Culture, Two Tailed Test, Membrane