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The details of the immunogen for this antibody are not available.
Our Abpromise guarantee covers the use of ab25333 in the following tested applications.
|Flow Cyt||Use 1µl for 106 cells.
ab18536 - Rat monoclonal IgG2b, is suitable for use as an isotype control with this antibody.
|ICC/IF||Use at an assay dependent concentration.|
|IHC-P||1/150. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.|
|Functional Studies||Use at an assay dependent concentration.
Complement-dependent cytotoxicity assays.
Flow Cytometry analysis of BALB/c mouse splenocytes labeling MHC Class II with ab25333 at 1 μg/106 cells (purple). A Mouse Anti-Rat IgG2b-AF488 was used as the secondary antibody. Grey - Isotype Control, Rat IgG2b-UNLB, followed by Mouse Anti-Rat IgG2b-AF488.
ab25333 staining MHC Class II in mouse spleen, white pulp tissue sections by Immunohistochemistry (IHC-P - paraformaldehyde-fixed, paraffin-embedded sections). Tissue was fixed with paraformaldehyde, permeabilized with 0.1% Tween-20 in PBS and blocked with 1% BSA for 40 minutes at room temperature; antigen retrieval was by heat mediation in citrate buffer pH 6. Samples were incubated with primary antibody (1/150 in TBS + 1% BSA) for 18 hours at 4°C. A Biotin-conjugated goat anti-rat IgG polyclonal (1/100) was used as the secondary antibody.
ICC/IF image of ab25333 stained Raw264.7 cells. The cells were 4% Formaldehyde fixed (10 min) and then incubated in 1%BSA / 10% normal Goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab25333, 10µg/ml) overnight at +4°C. The secondary antibody (green) was ab98420, DyLight® 488 Goat anti-Rat IgG (H+L) used at a 1/250 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.
ab25333 has not yet been referenced specifically in any publications.