Anti-TLR4 antibody

• Flow Cyt

Supplier Data

Flow Cytometry - Anti-TLR4 antibody (AB13556)

Flow cytometry analysis of THP-1 (Human monocytic leukemia cell line). Cells were fixed with 2% formaldehyde for 10 minutes at room temperature. ab13556 was used at 2 μg/10⁶ cells for 60 minutes at 37°C (green). Goat Anti- Rabbit Dylight 488 was used as a secondary antibody at 1/200 dilution for 40 minutes at 37°C. Isotype control was Rabbit IgG under the same conditions (blue).

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-TLR4 antibody (AB13556)

Immunohistochemistry analysis using Anti-TLR4 antibody (ab13556). Positive staining on mouse spleen tissue. Primary antibody ab13556 was used at 1 : 100.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-TLR4 antibody (AB13556)

Immunohistochemistry analysis using Anti-TLR4 antibody. Positive staining on mouse colon colitis. Primary antibody was used at a dilution of 1 : 100000 for 12 hours at 4°C. Secondary Antibody was biotin Goat Anti-Rabbit at 1 : 2000 for 1 hour at RT. Counterstain : Methyl Green at 200uL for 2 min at RT. Localization : Inflammatory cells.

• WB

Supplier Data

Western blot - Anti-TLR4 antibody (AB13556)

Blocking buffer : 5% Skim Milk powder in TBST.

Lane 1:

No primary antibody

Lane 2:

Western blot - Anti-TLR4 antibody (ab13556) at 1/1000 dilution

All lanes:

HeLa cell lysate at 10 µg

Secondary

All lanes:

HRP Goat Anti-Rabbit IgG at 1/4000 dilution

Predicted band size: 75 kDa

Observed band size: 75 kDa

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• WB

Lab

Western blot - Anti-TLR4 antibody (AB13556)

Western blot : Anti-TLR4 antibody (ab13556) staining at 1/1000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in magenta. In Western blot, ab13556 was shown to bind specifically to TLR4. First, unboiled samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in fluorescent western blot (TBS-based) blocking solution before incubation with primary antibodies overnight at 4°C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L 800CW and Goat anti-Mouse IgG H&L 680RD at 1/20000 dilution.

All lanes:

Western blot - Anti-TLR4 antibody (ab13556) at 1/1000 dilution

Lane 1:

HEK-293T overexpressing TLR4 (unboiled) cell lysate at 20 µg

Lane 2:

HEK-293T untransfected (unboiled) cell lysate at 20 µg

Observed band size: 70 kDa

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• WB

CiteAb

Western blot - Anti-TLR4 antibody (AB13556)

TLR4 western blotting using Anti-TLR4 antibody ab13556. Publication image and figure legend from Hu, X., Fatima, S., et al., 2021, Cell Death Dis, Pubmed 34385421.

Palmitic acid increases CRC cell proliferation in a TLR4-dependent manner.Protein expression of TLR4, MyD88, and TIRAP in A SW480 and B HCT116 cells after palmitic acid (PA, 50 µm) treatments. C Kinetics analysis of palmitic acid binding to TLR4 protein based on SPR platform Biacore X100. Representative sensorgrams were obtained from injections of palmitic acid at different concentrations as indicated. Expression of p-NFκB in D SW480 and E HCT116 cells after palmitic acid (PA, 50 µm) treatments. Proliferation of F SW480 and G HCT116 cells after palmitic acid (PA, 50 µm) treatments in the presence or absence of C34. H Knockout of TLR4 in HCT116 cells (HCT116TLR4-KO). I Proliferation of HCT116 cells and HCT116TLR4-KO cells after palmitic acid (PA, 50 µm) treatments in the presence or absence of C34. Data are shown as means ± SEM. n = 3 independent experiments. *p < < 0.05, **p < < 0.01 as indicated. Fatty acid treatments were done in the presence of 1% fatty acid-free BSA, 1% fatty acid-free BSA alone served as control (Ctrl).

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• WB

CiteAb

Western blot - Anti-TLR4 antibody (AB13556)

TLR4 western blotting using Anti-TLR4 antibody ab13556. Publication image and figure legend from Lai, C. H., Wang, K. C., et al., 2016, PLoS One, Pubmed 26741694.

TLR4 mediates cytokine production in HASMCs.(A) Basal levels of IL-6 and MCP-1 were assessed after being cultured in fresh medium without stimulation for 24 hours (n = 4 per group). (B, C, D) The protein expression of TLR4 in cell lysates (B) and levels of IL-6 (C) and MCP-1 (D) in supernatants were assessed after HMGB1 treatment for 24 hours (n = 4 per group). (E) Levels of IL-6 and MCP-1 were assessed after HMGB1 treatment (100 ng/dl) for 24 hours (n = 4 per group). (*P<0.05, **P<0.01 compared with untreated or control siRNA group.)

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• WB

CiteAb

Western blot - Anti-TLR4 antibody (AB13556)

TLR4 western blotting using Anti-TLR4 antibody ab13556. Publication image and figure legend from Dhlamini, Q., Wang, W., et al., 2022, Mol Med, Pubmed 35764933.

The effects of FGF1 pretreatment of LPS-induced cell death. A Representative images of TUNEL assay staining to detect apoptotic cells in the lung sections of mice at 12- and 24 h. Scale bar = 100 μm. Apoptotic cells were observed under fluorescence microscopy and the percentage of TUNEL positive cells per 40x magnification field was calculated. B WB analysis and quantification of TLR4 and the downstream apoptosis-related protein cleaved-caspase 3 in murine lungs. WB assays were performed at 12 h post LPS treatment. Data are presented as mean ± SD (n = 3 per group). **p < < 0.01, ***p < < 0.001, ****p < < 0.0001; One-way ANOVA with Tukey's post-hoc test

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