Overview

  • Product name

    Human HLA-DR ELISA Kit
    See all HLA-DR kits
  • Detection method

    Colorimetric
  • Precision

    Intra-assay
    Sample n Mean SD CV%
    Cell extract 5 3.6%
    Inter-assay
    Sample n Mean SD CV%
    Cell extract 3 12.7%
  • Sample type

    Cell culture extracts, Tissue Homogenate
  • Assay type

    Sandwich (quantitative)
  • Sensitivity

    5 pg/ml
  • Range

    31.3 pg/ml - 2000 pg/ml
  • Recovery

    Sample specific recovery
    Sample type Average % Range
    Cell culture extracts 98 89% - 105%
    Tissue Homogenate 94 86% - 100%

  • Assay time

    1h 30m
  • Assay duration

    One step assay
  • Species reactivity

    Reacts with: Human
  • Product overview

    HLA-DR in vitro SimpleStep ELISA® (Enzyme-Linked Immunosorbent Assay) kit is designed for the quantitative measurement of HLA-DR protein in cell and tissue extract.


    The SimpleStep ELISA® employs an affinity tag labeled capture antibody and a reporter conjugated detector antibody which immunocapture the sample analyte in solution. This entire complex (capture antibody/analyte/detector antibody) is in turn immobilized via immunoaffinity of an anti-tag antibody coating the well. To perform the assay, samples or standards are added to the wells, followed by the antibody mix. After incubation, the wells are washed to remove unbound material. TMB substrate is added and during incubation is catalyzed by HRP, generating blue coloration. This reaction is then stopped by addition of Stop Solution completing any color change from blue to yellow. Signal is generated proportionally to the amount of bound analyte and the intensity is measured at 450 nm. Optionally, instead of the endpoint reading, development of TMB can be recorded kinetically at 600 nm.


    HLA class II histocompatibility antigen, DR alpha chain (HLA-DR) is the alpha chain of a cell surface receptor heterodimer that binds peptides for presentation to CD4 T cells.  The alpha chain is fairly constant between individuals and pairs with varying alleles of the beta chain.  HLA-DR has been implicated in multiple auto immune conditions, as well as graft vs host disease. 

  • Tested applications

    Suitable for: Sandwich ELISAmore details
  • Platform

    Pre-coated microplate (12 x 8 well strips)

Properties

  • Storage instructions

    Store at +4°C. Please refer to protocols.
  • Components 1 x 96 tests
    10X Wash Buffer PT (ab206977) 1 x 20ml
    50X Cell Extraction Enhancer Solution (ab193971) 1 x 1ml
    5X Cell Extraction Buffer PTR (ab193970) 1 x 10ml
    Antibody Diluent 4BI 1 x 6ml
    10X Human HLA-DR Detector Antibody 1 x 600µl
    10X Human HLA-DR Capture Antibody 1 x 600µl
    Human HLA-DR Lyophilized Recombinant Protein 1 x 600µl
    Plate Seals 1 unit
    Sample Diluent NS 1 x 50ml
    SimpleStep Pre-Coated 96-Well Microplate (ab206978) 1 unit
    Stop Solution 1 x 12ml
    TMB Development Solution 1 x 12ml
  • Research areas

  • Function

    Binds 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 similarities

    Belongs to the MHC class II family.
    Contains 1 Ig-like C1-type (immunoglobulin-like) domain.
  • Post-translational
    modifications

    Ubiquitinated 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 localization

    Cell 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
  • Alternative names

    • DASS-397D15.1
    • DR alpha chain
    • DR alpha chain precursor
    • DRA_HUMAN
    • DRB1
    • DRB4
    • FLJ51114
    • Histocompatibility antigen HLA DR alpha
    • Histocompatibility antigen HLA-DR alpha
    • HLA class II histocompatibility antigen
    • HLA class II histocompatibility antigen DR alpha chain
    • HLA DR1B
    • HLA DR3B
    • HLA DRA
    • HLA DRA1
    • HLA DRB1
    • HLA DRB3
    • HLA DRB4
    • HLA DRB5
    • HLA-DR histocompatibility type
    • HLA-DRA
    • HLADR4B
    • HLADRA1
    • HLADRB
    • Major histocompatibility complex class II DR alpha
    • Major histocompatibility complex class II DR beta 1
    • Major histocompatibility complex class II DR beta 3
    • Major histocompatibility complex class II DR beta 4
    • Major histocompatibility complex class II DR beta 5
    • MGC117330
    • MHC cell surface glycoprotein
    • MHC class II antigen DRA
    • MHC II
    • MLRW
    • OTTHUMP00000029406
    • OTTHUMP00000029407
    see all
  • Database links

Applications

Our Abpromise guarantee covers the use of ab223593 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Application Abreviews Notes
Sandwich ELISA Use at an assay dependent concentration.

Images

  • SimpleStep ELISA technology allows the formation of the antibody-antigen complex in one single step, reducing assay time to 90 minutes. Add samples or standards and antibody mix to wells all at once, incubate, wash, and add your final substrate. See protocol for a detailed step-by-step guide.

     

  • Background-subtracted data values (mean +/- SD) are graphed.  

  • The concentrations of HLA-DR were measured in duplicates, interpolated from the HLA-DR standard curves corrected for sample dilution and extract load. Undiluted samples are as follows: Daudi cell extract 32.3 µg/mL, Raji cell extract 490 µg/mL, and human skin extract 500 µg/mL. The interpolated dilution factor and corrected values are plotted (mean +/- SD, n=2). The mean HLA-DR concentration was determined to be 52.0 pg/µg in Daudi Extract, 1.93 pg/µg in Raji Extract and 0.85 pg/µg in human skin homogenate extract.

Protocols

References

ab223593 has not yet been referenced specifically in any publications.

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