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Purified Rat Anti-Mouse CD16/CD32
Purified Rat Anti-Mouse CD16/CD32

Flow cytometric analysis of CD16/32 expression on mouse splenic lymphocytes. BALB/c mouse splenic leucocytes were stained with either Purified Rat IgG2a, λ Isotype Control (Cat. No. 553996; dashed line histogram) or Purified Rat Anti-Mouse CD16/CD32 antibody (Cat. No. 567021; solid line histogram) at 1 μg/test. The cells were washed and then stained with APC Goat Anti-Rat Ig (Cat. No. 551019). A histogram showing the expression of CD16/CD32 (or Ig Isotype control staining) was derived from gated events with the forward and side light-scatter characteristics of viable lymphocytes. Flow cytometry and data analysis were performed using a BD LSRFortessa™ Cell Analyzer System and FlowJo™ software. Data shown on this Technical Data Sheet are not lot specific.

Flow cytometric analysis of CD16/32 expression on mouse splenic lymphocytes. BALB/c mouse splenic leucocytes were stained with either Purified Rat IgG2a, λ Isotype Control (Cat. No. 553996; dashed line histogram) or Purified Rat Anti-Mouse CD16/CD32 antibody (Cat. No. 567021; solid line histogram) at 1 μg/test. The cells were washed and then stained with APC Goat Anti-Rat Ig (Cat. No. 551019). A histogram showing the expression of CD16/CD32 (or Ig Isotype control staining) was derived from gated events with the forward and side light-scatter characteristics of viable lymphocytes. Flow cytometry and data analysis were performed using a BD LSRFortessa™ Cell Analyzer System and FlowJo™ software. Data shown on this Technical Data Sheet are not lot specific.

Product Details
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BD Pharmingen™
Fcgr3/Fcgr2b; Fc gamma RIII/Fc gamma RIIB; FcγRIII/FcγRIIB
Mouse (QC Testing)
Rat WI, also known as Wistar (outbred) IgG2a, λ
Mouse Pre-B cells
Flow cytometry (Routinely Tested)
0.5 mg/ml
AB_2870011
Aqueous buffered solution containing ≤0.09% sodium azide.
RUO


Preparation And Storage

Store undiluted at 4°C. The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography.

Recommended Assay Procedures

Fc Receptor Blocking for Flow Cytometric Analysis

To reduce Fc receptor-mediated binding by antibodies of interest to cells that express CD16 and/or CD32 for flow cytometric analysis:

1. Preincubate cell suspension with Purified Ab93 antibody (Cat. No. 567021) at ≤ 1 μg of Ab93/million cells in 100 μl) at 4˚C for 5 minutes.

2. Add antibody of interest directly to cells preincubated with Purified Ab93 antibody (ie, the Purified Ab93 antibody is not washed away before staining cells).

3. If an Anti-Ig second-step antibody is being used, the secondary reagent must be chosen so that it does not bind to (crossreact with) Purified Ab93 antibody which has a Rat IgG2a, λ isotype.

BD™ CompBeads can be used as surrogates to assess fluorescence spillover (Compensation). When fluorochrome conjugated antibodies are bound to BD CompBeads, they have spectral properties very similar to cells. However, for some fluorochromes there can be small differences in spectral emissions compared to cells, resulting in spillover values that differ when compared to biological controls. It is strongly recommended that when using a reagent for the first time, users compare the spillover on cells and BD CompBead to ensure that BD CompBeads are appropriate for your specific cellular application.

Product Notices

  1. Since applications vary, each investigator should titrate the reagent to obtain optimal results.
  2. An isotype control should be used at the same concentration as the antibody of interest.
  3. Caution: Sodium azide yields highly toxic hydrazoic acid under acidic conditions. Dilute azide compounds in running water before discarding to avoid accumulation of potentially explosive deposits in plumbing.
  4. Sodium azide is a reversible inhibitor of oxidative metabolism; therefore, antibody preparations containing this preservative agent must not be used in cell cultures nor injected into animals. Sodium azide may be removed by washing stained cells or plate-bound antibody or dialyzing soluble antibody in sodium azide-free buffer. Since endotoxin may also affect the results of functional studies, we recommend the NA/LE (No Azide/Low Endotoxin) antibody format, if available, for in vitro and in vivo use.
  5. Please refer to http://regdocs.bd.com to access safety data sheets (SDS).
  6. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
567021 Rev. 1
Antibody Details
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Ab93

The Ab93 monoclonal antibody (aka, Antibody93) specifically recognizes a common epitope on the extracellular domains of mouse CD16 (Fc gamma RIII/FcγRIII encoded by Fcgr3) and CD32 (Fc gamma RIIB/FcγRIIB encoded by Fcgr2b). Therefore, Ab93 is referred to as an Anti-CD16/32 or Anti-FcgRII/III antibody. CD16 is variably expressed on neutrophils, macrophages, and natural killer (NK) cells whereas CD32 is expressed on B cells, monocytes, granulocytes, platelets and endothelial cells. CD16 and CD32 serve as low affinity receptors for IgG Fc constant regions and are involved in regulating various cellular functions including antibody-dependent cellular toxicity (ADCC), phagocytosis, effector cell degranulation, and B cell proliferation. In addition to identifying CD16- or CD32-positive cells, the Ab93 antibody is useful in phenotyping studies for blocking nonspecific staining due to Fc receptor-mediated binding of other antibodies. Ab93 is also useful in functional studies due to its Fc receptor blocking capability or by its capacity to crosslink Fc receptors leading to signal transduction that triggers cellular responses. Ab93 (Rat IgG2a, λ) and clone 2.4G2 (Rat IgG2b, κ), another mouse CD16/32-specific antibody, reportedly have similar specificities. The differences in the Ig heavy chain or Ig light chain isotypes of these CD16/32-specific antibodies afford flexibility in the design of experimental model systems involving other antibodies.

        

        

567021 Rev. 1
Format Details
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Purified
Tissue culture supernatant is purified by either protein A/G or affinity purification methods. Both methods yield antibody in solution that is free of most other soluble proteins, lipids, etc. This format provides pure antibody that is suitable for a number of downstream applications including: secondary labeling for flow cytometry or microscopy, ELISA, Western blot, etc.
Purified
567021 Rev.1
Citations & References
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Development References (5)

  1. Hirohashi T, Uehara S, Chase CM, et al. Complement independent antibody-mediated endarteritis and transplant arteriopathy in mice.. Am J Transplant. 2010; 10(3):510-7. (Clone-specific: Immunohistochemistry). View Reference
  2. Honjo K, Kubagawa Y, Kubagawa H. Is Toso/IgM Fc receptor (FcμR) expressed by innate immune cells?. Proc Natl Acad Sci USA. 2013; 110(28):E2540-1. (Clone-specific: Blocking, Flow cytometry). View Reference
  3. Nimmerjahn F, Ravetch JV. FcγRs in health and disease. Curr Top Microbiol Immunol. 2011; 350:105-125. (Biology). View Reference
  4. Oliver AM, Grimaldi JC, Howard MC, Kearney JF. Independently ligating CD38 and Fc gammaRIIB relays a dominant negative signal to B cells.. Hybridoma. 1999; 18(2):113-9. (Immunogen: Blocking, Flow cytometry, Fluorescence microscopy, Functional assay, Immunofluorescence, Inhibition). View Reference
  5. Torii I, Oka S, Hotomi M, et al. PIR-B-deficient mice are susceptible to Salmonella infection.. J Immunol. 2008; 181(6):4229-39. (Clone-specific: Blocking, Flow cytometry). View Reference
View All (5) View Less
567021 Rev. 1

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Global - Refer to manufacturer's instructions for use and related User Manuals and Technical data sheets before using this products as described


Comparisons, where applicable, are made against older BD Technology, manual methods or are general performance claims.  Comparisons are not made against non-BD technologies, unless otherwise noted.

For Research Use Only. Not for use in diagnostic or therapeutic procedures.