Description
The MIH37 monoclonal antibody specifically recognizes Programmed cell death 1 ligand 2 (PD-L2) which is also known as CD273 and Butyrophilin B7-DC (B7-DC). This regulatory molecule is a 42-kDa type I membrane glycoprotein encoded by Pdcd1lg2 which belongs to the B7 family of the Ig superfamily. CD273 (PD-L2) is comprised of an extracellular region with an N-terminal IgV-like domain followed by an IgC2-type domain, a transmembrane sequence, and a cytoplasmic tail. Although not detected on resting leucocytes, its expression is upregulated upon activation of macrophages and dendritic cells (DC) by a variety of stimulatory factors including IL-4, IL-13, or GM-CSF. CD273 (PD-L2) serves as a ligand for the coinhibitory receptor, CD279 (PD-1), a receptor that likewise binds to the CD274 (PD-L1) ligand. CD273 (PD-L2) also binds to Repulsive guidance molecule b (RGMb) that is expressed by T cells, macrophages, neutrophils, and DC. RGMb may associate with other receptors that transduce costimulatory or coinhibitory signals. The MIH37 antibody reportedly blocks the binding of CD273 (PB-L2) to its receptors, CD279 (PD-1) or RGMb and can inhibit antigen-specific T cell responses and hapten-induced contact hypersensitivity reactions. The TY25 monoclonal antibody reportedly binds with lower affinity near or at the same mouse CD273 (PD-L2) epitope recognized by the MIH37 antibody.
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® CompBeads to ensure that BD® CompBeads are appropriate for your specific cellular application.
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 that minimize unconjugated dye and antibody.
1. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
2. Please refer to http://regdocs.bd.com to access safety data sheets (SDS).
3. For U.S. patents that may apply, see bd.com/patents.
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. Since applications vary, each investigator should titrate the reagent to obtain optimal results.
6. The production process underwent stringent testing and validation to assure that it generates a high-quality conjugate with consistent performance and specific binding activity. However, verification testing has not been performed on all conjugate lots.
7. When using high concentrations of antibody, background binding of this dye to erythroid fragments produced by ammonium chloride-based lysis, such as with BD Pharm Lyse™ Lysing Buffer (Cat. No. 555899), has been observed when the antibody conjugate was present during the lysis procedure. This may cause nonspecific staining of target cells, such as leukocytes, which have bound the resulting erythroid fragments. This background can be mitigated by any of the following: titrating the antibody conjugate to a lower concentration, fixing samples with formaldehyde, or removing erythrocytes before staining (eg, gradient centrifugation or pre-lysis with wash). This background has not been observed when cells were lysed with BD FACS™ Lysing Solution (Cat. No. 349202) after staining.
8. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
9. An isotype control should be used at the same concentration as the antibody of interest.
10. Cy is a trademark of Global Life Sciences Solutions Germany GmbH or an affiliate doing business as Cytiva.
11. Please observe the following precautions: We recommend that special precautions be taken (such as wrapping vials, tubes, or racks in aluminum foil) to protect exposure of conjugated reagents, including cells stained with those reagents, to any room illumination. Absorption of visible light can significantly affect the emission spectra and quantum yield of tandem fluorochrome conjugates.
12. Tandem fluorochromes contain both an energy donor and an energy acceptor. Although every effort is made to minimize the lot-to-lot variation in the efficiency of the fluorochrome energy transfer, differences in the residual emission from the donor may be observed. Additionally, multi-laser cytometers may directly excite both the donor and acceptor fluorochromes. Therefore, we recommend for every tandem conjugate, a matched individual single-stain control be acquired for generating a compensation or spectral unmixing matrix.
