Clone:
REA528
Type of antibody:
Primary antibodies, Recombinant antibodies
Isotype:
recombinant human IgG1
Applications:
FC, MICS, IF, IHC
Alternative names:
MHC class II IAb

Extended validation for MHC Class II (I-Ab) Antibody, anti-mouse, REAfinity™

Specificity

Epitope competition
In order to compare the epitope specificity of an antibody, the clone being used is compared with other known clones recognizing the same antigen in a competition assay.
Other clonesOverlap in epitope recognition with REA528
25-9-17+
AF6-120.1++
KH74-
Cells were incubated with an excess of purified unconjugated MHC Class II (I-Ab) (REA528) antibody followed by staining with fluorochrome-conjugated antibodies of other known clones against the same marker. Based on the fluorescence signal obtained, the clones were identified as recognizing completely overlapping (++), partially overlapping (+), or completely different epitopes (-) of the marker.

Sensitivity

Performance comparison
Selected fluorochrome conjugated antibodies from Miltenyi Biotec were compared to commercially available hybridoma clones in flow cytometry analysis.
View details
Flow cytometric comparison of different clones for Anti-MHC Class II (I-Ab). Splenocytes from C57BL/6 mice were stained with Anti-MHC Class II (I-Ab) antibodies and with a suitable counterstaining. As a control, Anti-MHC Class II (I-Ab) antibody staining was omitted and cells were measured in the same channels. Flow cytometry was performed with the MACSQuant® Analyzer. Cell debris, dead cells, and cell doublets were excluded from the analysis based on scatter signals and 4',6-diamidino-2-phenylindole (DAPI) fluorescence. No FcR Blocking Reagent was used. The recommended titers of respective antibodies from different suppliers were used.
View details
Flow cytometric comparison of different clones for Anti-MHC Class II (I-Ab). Splenocytes from C57BL/6 mice were stained with Anti-MHC Class II (I-Ab) antibodies and with a suitable counterstaining. As a control, Anti-MHC Class II (I-Ab) antibody staining was omitted and cells were measured in the same channels. Flow cytometry was performed with the MACSQuant® Analyzer. Cell debris, dead cells, and cell doublets were excluded from the analysis based on scatter signals and 4',6-diamidino-2-phenylindole (DAPI) fluorescence. No FcR Blocking Reagent was used. The recommended titers of respective antibodies from different suppliers were used.
View details
Flow cytometric comparison of different clones for Anti-MHC Class II (I-Ab). Splenocytes from C57BL/6 mice were stained with Anti-MHC Class II (I-Ab) antibodies and with a suitable counterstaining. As a control, Anti-MHC Class II (I-Ab) antibody staining was omitted and cells were measured in the same channels. Flow cytometry was performed with the MACSQuant® Analyzer. Cell debris, dead cells, and cell doublets were excluded from the analysis based on scatter signals and 4',6-diamidino-2-phenylindole (DAPI) fluorescence. No FcR Blocking Reagent was used. The recommended titers of respective antibodies from different suppliers were used.
View details
Flow cytometric comparison of different clones for Anti-MHC Class II (I-Ab). Splenocytes from C57BL/6 mice were stained with Anti-MHC Class II (I-Ab) antibodies and with a suitable counterstaining. As a control, Anti-MHC Class II (I-Ab) antibody staining was omitted and cells were measured in the same channels. Flow cytometry was performed with the MACSQuant® Analyzer. Cell debris, dead cells, and cell doublets were excluded from the analysis based on scatter signals and 4',6-diamidino-2-phenylindole (DAPI) fluorescence. No FcR Blocking Reagent was used. The recommended titers of respective antibodies from different suppliers were used.
View details
Flow cytometric comparison of different clones for Anti-MHC Class II (I-Ab). Splenocytes from C57BL/6 mice were stained with Anti-MHC Class II (I-Ab) antibodies and with a suitable counterstaining. As a control, Anti-MHC Class II (I-Ab) antibody staining was omitted and cells were measured in the same channels. Flow cytometry was performed with the MACSQuant® Analyzer. Cell debris, dead cells, and cell doublets were excluded from the analysis based on scatter signals and 4',6-diamidino-2-phenylindole (DAPI) fluorescence. No FcR Blocking Reagent was used. The recommended titers of respective antibodies from different suppliers were used.
Flow cytometric comparison of different clones for Anti-MHC Class II (I-Ab). Splenocytes from C57BL/6 mice were stained with Anti-MHC Class II (I-Ab) antibodies and with a suitable counterstaining. As a control, Anti-MHC Class II (I-Ab) antibody staining was omitted and cells were measured in the same channels. Flow cytometry was performed with the MACSQuant® Analyzer. Cell debris, dead cells, and cell doublets were excluded from the analysis based on scatter signals and 4',6-diamidino-2-phenylindole (DAPI) fluorescence. No FcR Blocking Reagent was used. The recommended titers of respective antibodies from different suppliers were used.
Fixation data
To provide an indication on how an antibody performs after fixation of cells, in-house data on staining results before and after fixation with 3.7% formaldehyde using Miltenyi Biotec antibodies are provided. Different experimental settings may lead to different results.
No fixation
Post fixation
View details
Comparison of staining pattern on non-fixed and fixed cells using MHC Class II (I-Ab) (REA528). The performance of the antibody after fixation was tested by comparing the staining pattern on fresh (no fixation) versus fixed (post fixation) cells.
View details
Comparison of staining pattern on non-fixed and fixed cells using MHC Class II (I-Ab) (REA528). The performance of the antibody after fixation was tested by comparing the staining pattern on fresh (no fixation) versus fixed (post fixation) cells.
Comparison of staining pattern on non-fixed and fixed cells using MHC Class II (I-Ab) (REA528). The performance of the antibody after fixation was tested by comparing the staining pattern on fresh (no fixation) versus fixed (post fixation) cells.

Specifications for MHC Class II (I-Ab) Antibody, anti-mouse, REAfinity™

Overview

Clone REA528 recognizes the mouse MHC class II alloantigen I-Ab of H-2b bearing mouse strains. It also cross-reacts with the H-2k and H-2u haplotypes, but reactivity to other haplotypes like, for example, d, f, g7, p, q, r, and s has not been observed. MHC class II is expressed on antigen-presenting cells, such as dendritic cells, monocytes, macrophages, B cells in lymphoid and non-lymphoid tissue, thymic epithelial cells, and on subsets of hematopoietic progenitor cells in the bone marrow.
Additional information: Clone REA528 displays negligible binding to Fc receptors.

Alternative names

MHC class II IAb

Detailed product information

Technical specifications

CloneREA528
Clonalitymonoclonal
Isotyperecombinant human IgG1
Isotype controlREA Control Antibody, human IgG1
Hostcell line
Type of antibodyPrimary antibodies, Recombinant antibodies
Speciesmouse
AntigenMHC Class II (I-Ab)
Alternative names of antigenMHC class II IAb
Molecular mass of antigen [kDa]27
Distribution of antigenB cells, dendritic cells, macrophages, monocytes, epithelial cells, hematopoietic stem and progenitor cells
Entrez Gene ID14961
RRIDAB_2733747, AB_2784433, AB_2857711, AB_2652862, AB_2652863, AB_2652868, AB_2652869, AB_2733746

Resources for MHC Class II (I-Ab) Antibody, anti-mouse, REAfinity™

Certificates

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References for MHC Class II (I-Ab) Antibody, anti-mouse, REAfinity™

Publications

  1. Cuenca, M. et al. (2016) Targeting of Ly9 (CD229) disrupts marginal zone and B1 B cell homeostasis and antibody responses. J. Immunol. 196: 726-737
  2. Gradehandt, G. et al. (1993) The I-Ab-restricted alloresponse of D10.G4.1 T cells is based on the recognition of an endogenous peptide. Immunology 78(4): 592-599
  3. Tobita, T. et al. (2003) A role for the P1 anchor residue in the thermal stability of MHC class II molecule I-Ab. Immunol. Lett. 85(1): 47-52
  4. Zhu, Y. et al. (2003) Crystal structure of MHC class II I-Ab in complex with a human CLIP peptide: prediction of an I-Ab peptide-binding motif. J. Mol. Biol. 326(4): 1157-1174

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