Synthetic peptide corresponding to Human TLR4 aa 100-200 conjugated to Keyhole Limpet Haemocyanin (KLH).
Database link: O00206
Our Abpromise guarantee covers the use of ab22048 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|ELISA||Use at an assay dependent concentration. PubMed: 24952384|
|IHC-P||1/100. Perform heat mediated antigen retrieval before commencing with IHC staining protocol. Antigen retrieval with citrate buffer pH 6.|
|Flow Cyt||Use 1µg for 106 cells.
Methanol or paraformaldehyde fixed cells.
ab170192 - Mouse monoclonal IgG2b, is suitable for use as an isotype control with this antibody.
|ICC||Use at an assay dependent concentration.|
|ICC/IF||Use at an assay dependent concentration.|
ab22048 staining TLR4 in Human stomach tissue sections by Immunohistochemistry (IHC-P - formaldehyde-fixed, paraffin-embedded sections). Tissue was fixed with formaldehyde and blocked with 5% serum for 1 hour; antigen retrieval was by heat mediation in citrate buffer (10mM, pH 6) (ab64236). Samples were incubated with primary antibody (1/100) for 1 hour at 23°C. An undiluted HRP-conjugated goat anti-mouse IgG polyclonal was used as the secondary antibody.
ab22048 staining TLR4 in Mouse spleen tissue sections by Immunohistochemistry (IHC-Fr - frozen sections). Tissue was fixed with acetone and blocked with 10% serum for 30 minutes at 20°C. Samples were incubated with primary antibody (1/100 in PBS-BSA) for 1 hour at 20°C. An Alexa Fluor®488-conjugated Rat anti mouse IgG polyclonal (1/200) was used as the secondary antibody.
TLR4 labelling (yellow) co-localises with CD35 labelling (red) on follicular dendritic cells in the spleen. TLR 4 alone (green) can also be seen on other cell types in the spleen- most probably macrophages.
Overlay histogram showing Jurkat cells (ab7899) stained with ab22048 (red line). The cells were fixed with methanol (5 min) and incubated in 1x PBS / 10% normal goat serum (ab7481) / 0.3M glycine to block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab22048, 1µg/1x106 cells) for 30 min at 22°C. The secondary antibody used was DyLight® 488 goat anti-mouse IgG (H+L) (ab96879) at 1/500 dilution for 30 min at 22°C. Isotype control antibody (black line) was mouse IgG2b [PLPV219] (ab91366, 2µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a positive signal in Jurkat cells fixed with 4% paraformaldehyde (10 min) used under the same conditions.
Please note that Abcam do not have data for use of this antibody on non-fixed cells. We welcome any customer feedback.
Immunofluorescence analysis of murine macrophages, staining TLR4 with ab22048.
Cells were fixed with formaldehyde, permeabilized with 0.1% Triton X-100 and blocked with 1% BSA for 10 minutes at 25°C. Samples were incubated with primary antibody (1/100 in 1% BSA) for 12 hours at 4°C. A TexasRed®-conjugated donkey anti-mouse (ab6818) polyclonal IgG (1/100) was used as the secondary antibody.
ab22048 (5 ug/ml) staining TLR4 in Human skin tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Membrane-cytoplasmic immunopositivity of TLR4 was primarily observed in the pigmented basel cells and the adjacent keratinocytes in the epidermal layer.
ab22048 at 5 µg/ml staining TLR4 in Human colon tissue sections by Immunohistochemistry (Formalin/ PFA-fixed paraffin-embedded tissue sections). Left hand image shows staining with an isotype control antibody whilst right one shows staining with ab22048.
ab22048 staining TLR4 in Rat's salivary gland tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). ab22048 at a dilution of 1:100 generated a membrane-cytoplasmic staining in the tissue with stronger signal in ductal epithelial cells.