CD32 Recombinant Rabbit Monoclonal Antibody [JM10-70]

Overview

Product Name

CD32 Recombinant Rabbit Monoclonal Antibody [JM10-70] - BSA and Azide free

Antibody Type

Recombinant Rabbit monoclonal Antibody

Immunogen

Recombinant protein within Human CD32 aa 23-274 / 310.

Species Reactivity

Human

Validated Applications

WB, IHC-P, IF-Cell, FC, IP

Target Molecular Weight

Predicted band size: 35 kDa

Positive Control

THP-1 cell lysate, K-562 cell lysate, human spleen tissue, K-562.

Conjugation

unconjugated

Clone Number

JM10-70

Product Features

Form

Liquid

Concentration

1mg/ml

Storage Instructions

Store at +4℃ after thawing. Aliquot store at -20℃ or -80℃. Avoid repeated freeze / thaw cycles.

Storage Buffer

1*PBS (pH7.4).

Isotype

IgG

Purification Method

Protein A affinity purified.

Application Dilution

WB
1:1,000-1:5,000

IHC-P
1:400

IF-Cell
1:100

FC
1:1,000

IP
1-2μg/sample

Target

Function

CD32 (also designated Fc gamma RII) is a low affinity receptor for the Fc fragment of aggregated IgG. CD32 is responsible for the clearance of immunocomplexes by macrophages and also plays an important role in the regulation of antibody production by B cells. IgG can noncooperatively bind either one or two highly glycosylated CD32 molecules, and this binding delivers a negative signal for B cells . CD32 exists as several isoforms that are produced by alternative splicing of three distinct genes, A, B, and C . These isoforms are designated FcgRIIA, FcgRIIB1, FcgRIIB3, and FcgRIIC . All isoforms are present on monocytes, placental trophoblasts and endothelial cells . In addition, the FcgRIIB forms are present on B lymphocytes, and the FcgRIIA and FcgRIIC forms are found on neutrophils.

Background References

1. Sips M et al. Fc receptor-mediated phagocytosis in tissues as a potent mechanism for preventive and therapeutic HIV vaccine strategies. Mucosal Immunol 9:1584-1595 (2016).

2. Karagiannis P et al. IgG4 subclass antibodies impair antitumor immunity in melanoma. J Clin Invest 123:1457-74 (2013).

Tissue Specificity

Found on monocytes, neutrophils and eosinophil platelets.

Post-translational Modification

Phosphorylated by SRC-type Tyr-kinases such as LYN, BLK, FYN, HCK and SYK.

Subcellular Location

Cell membrane.

UNIPROT

SwissProt: P12318 Human

Synonyms

CD32 antibody

CD32A antibody

CD32B antibody

CD32C antibody

CDw32 antibody

Fc fragment of IgG low affinity IIa receptor antibody

Fc fragment of IgG low affinity IIa receptor for (CD32) antibody

Fc fragment of IgG low affinity IIb receptor antibody

Fc fragment of IgG low affinity IIb receptor for (CD32) antibody

Fc fragment of IgG low affinity IIc receptor antibody

Expand

Images

☑ Cell treatment (CT)

Western blot analysis of CD32 on different lysates with Rabbit anti-CD32 antibody (HA750382) at 1/2,000 dilution.

Lane 1: THP-1 cell lysate
Lane 2: THP-1 cell lysate treated with deglycosylation
Lane 3: K-562 cell lysate

Lysates/proteins at 20 µg/Lane.

Predicted band size: 35 kDa
Observed band size: 40/31 kDa

Exposure time: 3 minutes; ECL: K1801;
4-20% SDS-PAGE gel.

Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (HA750382) at 1/2,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.
Immunohistochemical analysis of paraffin-embedded human spleen tissue with Rabbit anti-CD32 antibody (HA750382) at 1/400 dilution.

The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH₂O and PBS, and then probed with the primary antibody (HA750382) at 1/400 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.
Immunocytochemistry analysis of K-562 cells labeling CD32 with Rabbit anti-CD32 antibody (HA750382) at 1/100 dilution.

Cells were fixed in 80% precooled methanol for 5 minutes at room temperature, then blocked with 1% BSA in 10% negative goat serum for 1 hour at room temperature. Cells were then incubated with Rabbit anti-CD32 antibody (HA750382) at 1/100 dilution in 1% BSA in PBST overnight at 4 ℃. Goat Anti-Rabbit IgG H&L (iFluor™ 488, HA1121) was used as the secondary antibody at 1/1,000 dilution. PBS instead of the primary antibody was used as the secondary antibody only control. Nuclear DNA was labelled in blue with DAPI.
Flow cytometric analysis of K-562 cells labeling CD32.

Cells were fixed and permeabilized. Then stained with the primary antibody (HA750382, 1/1,000) (red) compared with Rabbit IgG Isotype Control (green). After incubation of the primary antibody at +4℃ for an hour, the cells were stained with a iFluor™ 488 conjugate-Goat anti-Rabbit IgG Secondary antibody (HA1121) at 1/1,000 dilution for 30 minutes at +4℃. Unlabelled sample was used as a control (cells without incubation with primary antibody; black).
CD32 was immunoprecipitated from 0.2 mg THP-1 cell lysate with HA750382 at 2 µg/10 µl beads. Western blot was performed from the immunoprecipitate using HA750382 at 1/2,000 dilution. HRP Conjugated Anti-Rabbit IgG for IP Nano-secondary antibody at 1/5,000 dilution was used for 1 hour at room temperature.

Lane 1: THP-1 cell lysate (input)
Lane 2: HA750382 IP in THP-1 cell lysate
Lane 3: Rabbit IgG instead of HA750382 in THP-1 cell lysate

Blocking/Dilution buffer: 5% NFDM/TBST
Exposure time: 1 minute 7 seconds; ECL: K1801

Please note: All products are "FOR RESEARCH USE ONLY AND ARE NOT INTENDED FOR DIAGNOSTIC OR THERAPEUTIC USE"