Anti-Human CD32 (FcγRIIA) [Clone IV.3] — Purified in vivo GOLD™ Functional Grade

Human K562 leukemia cell line

This monoclonal antibody is aseptically packaged and formulated in 0.01 M phosphate buffered saline (150 mM NaCl) PBS pH 7.2 - 7.4 with no carrier protein, potassium, calcium or preservatives added. Due to inherent biochemical properties of antibodies, certain products may be prone to precipitation over time. Precipitation may be removed by aseptic centrifugation and/or filtration.

Functional grade preclinical antibodies are manufactured in an animal free facility using protein free cell culture techniques and are purified by a multi-step process including the use of protein A or G to assure extremely low levels of endotoxins, leachable protein A or aggregates.

Functional grade preclinical antibodies may be stored sterile as received at 2-8°C for up to one month. For longer term storage, aseptically aliquot in working volumes without diluting and store at ≤ -70°C. Avoid Repeated Freeze Thaw Cycles.

2-8°C wet ice

Applications in Research
Clone IV.3 is a gold-standard reagent for both in vitro and in vivo applications.

Fc Blocking: It is frequently used in flow cytometry and cell sorting protocols as an Fc blocking reagent to improve specific antibody staining and reduce background noise.

Functional Assays: The IV.3 antibody is capable of blocking the biological downstream effects of CD32 in in vitro functional assays. Furthermore, IV.3 F(ab')2 fragments have been successfully utilized to block CD32 in vivo in humanized transgenic mouse models to study immune pathway activation, therapeutic antibody efficacy, and viral trogocytosis (such as HIV-1 pathogenesis).

Human FcγRII - binds most strongly to the RIIA isoform

Biological Significance of CD32 (FcγRII)

CD32, also known as FcγRII, is a polymorphic, 40 kDa type I transmembrane glycoprotein belonging to the immunoglobulin (Ig) gene superfamily. It serves as a low-affinity receptor for the Fc region of monomeric IgG antibodies, but exhibits a high affinity for complexed or aggregated IgG. CD32 plays a critical role in cellular communication and the regulation of both activating and inhibitory immune responses.

The CD32 receptor is encoded by three distinct genes (A, B, and C), which undergo alternative mRNA splicing to generate at least six different isoforms (IIa1, IIa2, IIb1, IIb2, IIb3, and IIc). These isoforms are widely expressed across a variety of immune cells, including monocytes, macrophages, dendritic cells, neutrophils, granulocytes, platelets, and B cells. Functionally, CD32 enables interactions between FcγRII-expressing cells and opsonized antigens or immune complexes. This interaction drives essential immune mechanisms, including phagocytosis, antibody-dependent cellular cytotoxicity (ADCC), degranulation, cytokine release, and modulation of B cell proliferation.

Specificity and Mechanism of Clone IV.3
The Clone IV.3 monoclonal antibody specifically targets and binds to the FcγRIIA (CD32a) isoform, interacting with an epitope mapped to domain 2 within the ligand-binding site (amino acids 132-137: FSHLDP listed in the following paper: DOI: 10.4049/jimmunol.1101467).

Clone IV.3 is highly valued in immunological research because it acts as a potent blocking antibody. Non-specific binding of IgG antibodies to Fc receptors on macrophages, monocytes, and other immune cells is a common cause of false positives and high background noise in flow cytometry, cell separation, immunohistochemistry (IHC), and immunoprecipitation (IP). By selectively binding to the ligand-binding site of human CD32a, Clone IV.3 neutralizes the receptor, effectively preventing the non-specific uptake of immune complexes.

Fc region of monomeric IgG (weakly), aggregated IgG (strongly)

P12318

1.) Ramsland, P. A., et al. (2011). Structural basis for FcγRIIa recognition of human IgG and formation of inflammatory signaling complexes. The Journal of Immunology, 187(6), 3208-3217. DOI: 10.4049/jimmunol.1101467
2.) Walsh TG, Harper MT, Poole AW. SDF-1α is a novel autocrine activator of platelets operating through its receptor CXCR4. Cell Signal. 2015 Jan;27(1):37-46. doi: 10.1016/j.cellsig.2014.09.021. Epub 2014 Oct 5. PMID: 25283599; PMCID: PMC4265729.
3.). Ben Mkaddem S, Hayem G, Jönsson F, Rossato E, Boedec E, Boussetta T, El Benna J, Launay P, Goujon JM, Benhamou M, Bruhns P, Monteiro RC. Shifting FcγRIIA-ITAM from activation to inhibitory configuration ameliorates arthritis. J Clin Invest. 2014 Sep;124(9):3945-59. doi: 10.1172/JCI74572. Epub 2014 Jul 25. PMID: 25061875; PMCID: PMC4151227.
4.). Flinsenberg TWH, Janssen WJ, Herczenik E, Boross P, Nederend M, Jongeneel LH, Scholman RC, Boelens JJ, Maas C, van Gijn ME, van Montfrans JM, Leusen JH, Boes M. A novel FcγRIIa Q27W gene variant is associated with common variable immune deficiency through defective FcγRIIa downstream signaling. Clin Immunol. 2014 Nov;155(1):108-117. doi: 10.1016/j.clim.2014.09.006. Epub 2014 Sep 19. PMID: 25242138.
5.) Trist HM, Tan PS, Wines BD, Ramsland PA, Orlowski E, Stubbs J, Gardiner EE, Pietersz GA, Kent SJ, Stratov I, Burton DR, Hogarth PM. Polymorphisms and interspecies differences of the activating and inhibitory FcγRII of Macaca nemestrina influence the binding of human IgG subclasses. J Immunol. 2014 Jan 15;192(2):792-803. doi: 10.4049/jimmunol.1301554. Epub 2013 Dec 16. PMID: 24342805; PMCID: PMC4080885.