Product nameAnti-HLA-DR antibody [TAL 1B5]
See all HLA-DR primary antibodies
DescriptionMouse monoclonal [TAL 1B5] to HLA-DR
Tested applicationsSuitable for: WB, IHC-P, Flow Cytmore details
Species reactivityReacts with: Human
Tissue, cells or virus corresponding to HLA-DR. Bristol 8 separated alpha chain preparation
- Ab20181 gave a positive signal in Raji, Ramos and Daudi whole cell lysates, and in the following human tissue lysates: spleen; tonsil; liver. This antibody gave a positive result in IHC in the following FFPE tissue: Human skin melanoma.
Storage instructionsShipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C or -80°C. Avoid freeze / thaw cycle.
Storage bufferpH: 7.40
Preservative: 0.02% Sodium azide
Constituents: PBS, 6.97% L-Arginine
Concentration information loading...
PurityProtein G purified
Clone numberTAL 1B5
Light chain typekappa
Our Abpromise guarantee covers the use of ab20181 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|WB||Use a concentration of 2 - 5 µg/ml. Detects a band of approximately 35 kDa (predicted molecular weight: 29 kDa).
We recommend using 3% milk as the blocking agent in Western Blot.
|IHC-P||Use a concentration of 0.1 µg/ml. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.|
|Flow Cyt||Use 0.005-0.01µg for 106 cells.
ab170190 - Mouse monoclonal IgG1, is suitable for use as an isotype control with this antibody.
FunctionBinds peptides derived from antigens that access the endocytic route of antigen presenting cells (APC) and presents them on the cell surface for recognition by the CD4 T-cells. The peptide binding cleft accommodates peptides of 10-30 residues. The peptides presented by MHC class II molecules are generated mostly by degradation of proteins that access the endocytic route, where they are processed by lysosomal proteases and other hydrolases. Exogenous antigens that have been endocytosed by the APC are thus readily available for presentation via MHC II molecules, and for this reason this antigen presentation pathway is usually referred to as exogenous. As membrane proteins on their way to degradation in lysosomes as part of their normal turn-over are also contained in the endosomal/lysosomal compartments, exogenous antigens must compete with those derived from endogenous components. Autophagy is also a source of endogenous peptides, autophagosomes constitutively fuse with MHC class II loading compartments. In addition to APCs, other cells of the gastrointestinal tract, such as epithelial cells, express MHC class II molecules and CD74 and act as APCs, which is an unusual trait of the GI tract. To produce a MHC class II molecule that presents an antigen, three MHC class II molecules (heterodimers of an alpha and a beta chain) associate with a CD74 trimer in the ER to form an heterononamer. Soon after the entry of this complex into the endosomal/lysosomal system where antigen processing occurs, CD74 undergoes a sequential degradation by various proteases, including CTSS and CTSL, leaving a small fragment termed CLIP (class-II-associated invariant chain peptide). The removal of CLIP is facilitated by HLA-DM via direct binding to the alpha-beta-CLIP complex so that CLIP is released. HLA-DM stabilizes MHC class II molecules until primary high affinity antigenic peptides are bound. The MHC II molecule bound to a peptide is then transported to the cell membrane surface. In B-cells, the interaction between HLA-DM and MHC class II molecules is regulated by HLA-DO. Primary dendritic cells (DCs) also to express HLA-DO. Lysosomal miroenvironment has been implicated in the regulation of antigen loading into MHC II molecules, increased acidification produces increased proteolysis and efficient peptide loading.
Sequence similaritiesBelongs to the MHC class II family.
Contains 1 Ig-like C1-type (immunoglobulin-like) domain.
modificationsUbiquitinated by MARCH1 or MARCH8 at Lys-244 leading to down-regulation of MHC class II. When associated with ubiquitination of the beta subunit of HLA-DR: HLA-DRB4 'Lys-254', the down-regulation of MHC class II may be highly effective.
Cellular localizationCell membrane. Endoplasmic reticulum membrane. Golgi apparatus > trans-Golgi network membrane. Endosome membrane. Lysosome membrane. Late endosome membrane. The MHC class II complex transits through a number of intracellular compartments in the endocytic pathway until it reaches the cell membrane for antigen presentation.
- Information by UniProt
- DR alpha chain antibody
- DR alpha chain precursor antibody
- DRA_HUMAN antibody
All lanes : Anti-HLA-DR antibody [TAL 1B5] (ab20181) at 5 µg/ml
Lane 1 : Raji (Human Burkitt's lymphoma cell line) Whole Cell Lysate
Lane 2 :
Daudi whole cell lysate (ab3951)
Lane 3 : Ramos (Human Burkitt's lymphoma cell line) Whole Cell Lysate
Lane 4 : Human spleen tissue lysate - total protein (ab29699)
Lane 5 : Tonsil (Human) Tissue Lysate
Lane 6 : Human liver tissue lysate - total protein (ab29889)
Lysates/proteins at 20 µg per lane.
All lanes : Goat Anti-Mouse IgG H&L (HRP) preadsorbed (ab97040) at 1/10000 dilution
Performed under reducing conditions.
Predicted band size: 29 kDa
Observed band size: 29,35 kDa why is the actual band size different from the predicted?
Exposure time: 16 minutes
Ab20181 staining human normal thymus tissue. Staining is localised to cellular membranes.
Left panel: with primary antibody at 1 ug/ml. Right panel: isotype control.
Sections were stained using an automated system DAKO Autostainer Plus , at room temperature. Sections were rehydrated and antigen retrieved with the Dako 3-in-1 antigen retrieval buffer EDTA pH 9.0 in a DAKO PT Link. Slides were peroxidase blocked in 3% H2O2 in methanol for 10 minutes. They were then blocked with Dako Protein block for 10 minutes (containing casein 0.25% in PBS) then incubated with primary antibody for 20 minutes and detected with Dako Envision Flex amplification kit for 30 minutes. Colorimetric detection was completed with diaminobenzidine for 5 minutes. Slides were counterstained with Haematoxylin and coverslipped under DePeX. Please note that for manual staining we recommend to optimize the primary antibody concentration and incubation time (overnight incubation), and amplification may be required.
Human peripheral blood lymphocytes stained with ab20181 (red line). Human whole blood was processed using a modified protocol based on Chow et al, 2005 (PMID: 16080188). In brief, human whole blood was fixed in 4% formaldehyde (methanol-free) for 10 min at 22°C. Red blood cells were then lyzed by the addition of Triton X-100 (final concentration - 0.1%) for 15 min at 37°C. For experimentation, cells were treated with 50% methanol (-20°C) for 15 min at 4°C. Cells were then incubated with the antibody (ab20181, 0.01μg/1x106 cells) for 30 min at 4°C. The secondary antibody used was Alexa Fluor® 488 goat anti-mouse IgG (H&L) (ab150113) at 1/2000 dilution for 30 min at 4°C. Isotype control antibody (black line) was mouse IgG1 [ICIGG1] (ab91353, 1μg/1x106 cells) used under the same conditions. Unlabelled sample (blue line) was also used as a control. Acquisition of >30,000 total events were collected using a 20mW Argon ion laser (488nm) and 525/30 bandpass filter. Gating strategy – peripheral blood lymphocytes.
Anti-HLA-DR antibody [TAL 1B5] (ab20181) at 1 µg/ml + whole tissue lysate prepared from human colon at 50 µg
HRP conjugated goat anti-mouse polyclonal at 1/3000 dilution
Developed using the ECL technique.
Predicted band size: 29 kDa
Observed band size: 33 kDa why is the actual band size different from the predicted?
Exposure time: 5 seconds
IHC image of HLA-DR [TAL 1B5] staining in Human skin melanoma formalin fixed paraffin embedded tissue section, performed on a Leica Bond™ system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab20181, 0.1µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.
For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.
Human peripheral blood mononuclear cells isolated from whole blood were treated with BD Cytofix/Cytoperm™ kit for intracellular staining. Cells were then incubated in 10% normal goat serum to block non-specific protein-protein interactions followed by the antibody (ab20181, 0.005μg/1x106 cells) for 30 min at 4°C. The secondary antibody used was Alexa Fluor® 488 goat anti-mouse IgG (H&L) (ab150117) at 1/2000 dilution for 30 min at 4°C.
Isotype control antibody (black line) was mouse IgG1 [15-6E10A7] (ab170190, 0.005μg/1x106 cells) used under the same conditions. Unlabelled sample (blue line) was also used as a control.
Acquisition of 10,000 total events were collected using a 50mW Blue laser (488nm) and 530/30 bandpass filter. Gating strategy – lymphocytes.
This product has been referenced in:
- Zhao SJ et al. SPARCL1 suppresses osteosarcoma metastasis and recruits macrophages by activation of canonical WNT/ß-catenin signaling through stabilization of the WNT-receptor complex. Oncogene 37:1049-1061 (2018). Read more (PubMed: 29084211) »
- Álvaro-Benito M et al. Quantification of HLA-DM-Dependent Major Histocompatibility Complex of Class II Immunopeptidomes by the Peptide Landscape Antigenic Epitope Alignment Utility. Front Immunol 9:872 (2018). Read more (PubMed: 29774024) »