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BV421 Rat Anti-Mouse CD16/CD32
BV421 Rat Anti-Mouse CD16/CD32

Two-color flow cytometric analysis of CD16/32 expression on mouse splenic lymphocytes. BALB/C mouse splenic leucocytes were stained with APC Armenian Hamster Anti-Mouse CD3e antibody (Cat. No. 553066/561826) and either BD Horizon™ BV421 Rat IgG2a, λ Isotype Control (Cat. No. 557076; Left Plot) or BD Horizon™ BV421 Rat Anti-Mouse CD16/CD32 antibody (Cat. No. 567020; Right Plot) at 0.5 μg/test. A pseudocolor density plot showing the correlated expression of CD16/CD32 (or Ig Isotype control staining) versus CD3e 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.

Two-color flow cytometric analysis of CD16/32 expression on mouse splenic lymphocytes. BALB/C mouse splenic leucocytes were stained with APC Armenian Hamster Anti-Mouse CD3e antibody (Cat. No. 553066/561826) and either BD Horizon™ BV421 Rat IgG2a, λ Isotype Control (Cat. No. 557076; Left Plot) or BD Horizon™ BV421 Rat Anti-Mouse CD16/CD32 antibody (Cat. No. 567020; Right Plot) at 0.5 μg/test. A pseudocolor density plot showing the correlated expression of CD16/CD32 (or Ig Isotype control staining) versus CD3e 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 Horizon™
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)
0.2 mg/ml
AB_2870012
Aqueous buffered solution containing BSA and ≤0.09% sodium azide.
RUO


Preparation And Storage

Store undiluted at 4°C and protected from prolonged exposure to light. Do not freeze. The monoclonal antibody was purified from tissue culture supernatant or ascites by affinity chromatography. The antibody was conjugated to the dye under optimum conditions and unconjugated antibody and free dye were removed.

Recommended Assay Procedures

       BD™ CompBeads can be used as surrogates to assess fluorescence spillover (Compensation).  When fluorochrome conjugated antibodies are bound to 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 CompBead to ensure that BD Comp beads are appropriate for your specific cellular application.

       For optimal and reproducible results, BD Horizon Brilliant Stain Buffer should be used anytime two or more BD Horizon Brilliant dyes are used in the same experiment.  Fluorescent dye interactions may cause staining artifacts which may affect data interpretation.  The BD Horizon Brilliant Stain Buffer was designed to minimize these interactions.  More information can be found in the Technical Data Sheet of the BD Horizon Brilliant Stain Buffer (Cat. No. 563794/566349) or the BD Horizon Brilliant Stain Buffer Plus (Cat. No. 566385).

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. Source of all serum proteins is from USDA inspected abattoirs located in the United States.
  4. 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.
  5. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
  6. Pacific Blue™ is a trademark of Molecular Probes, Inc., Eugene, OR.
  7. BD Horizon Brilliant Violet 421 is covered by one or more of the following US patents: 8,158,444; 8,362,193; 8,575,303; 8,354,239.
  8. BD Horizon Brilliant Stain Buffer is covered by one or more of the following US patents: 8,110,673; 8,158,444; 8,575,303; 8,354,239.
  9. Please refer to http://regdocs.bd.com to access safety data sheets (SDS).
  10. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
567022 Rev. 2
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.

The antibody was conjugated to BD Horizon BV421 which is part of the BD Horizon Brilliant™ Violet family of dyes. With an Ex Max near 407 nm and Em Max near 421 nm, BD Horizon BV421 can be excited by the violet laser (405 nm) and detected with a 450/50 nm filter. BD Horizon BV421 conjugates are very bright, often exhibiting a 10 fold improvement in brightness compared to Pacific BlueTM conjugates. Due to nearly identical excitation and emission properties but different spillover characteristics, BD Horizon BV421, Pacific Blue, and BD Horizon V450 cannot be used simultaneously.

567022 Rev. 2
Format Details
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BV421
The BD Horizon Brilliant Violet™ 421 (BV421) Dye is part of the BD Horizon Brilliant Violet™ family of dyes. This polymer-technology based dye has an excitation maximum (Ex Max) of 407-nm and an emission maximum (Em Max) at 423-nm. Driven by BD innovation, BV421 is designed to be excited by the violet laser (405-nm) and detected using an optical filter centered near 420-nm (e.g., a 431/28-nm or 450/50-nm bandpass filter). BV421 is an ideal alternative for V450 as it is approximately ten times brighter with less spillover into the BV510/V500 detector. Please ensure that your instrument’s configurations (lasers and optical filters) are appropriate for this dye.
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BV421
Violet 405 nm
407 nm
423 nm
567022 Rev.2
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
567022 Rev. 2

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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.