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RB780 Rat Anti-Mouse CD274
RB780 Rat Anti-Mouse CD274
Multiparameter flow cytometric analysis using BD OptiBuild™ RB780 Rat Anti-Mouse CD274 (B7-H1) antibody (Cat. No. 755394; Right Plot) on viable C57BL/6 mouse splenocytes activated with plate-bound Anti-Mouse CD3 for 3 days, with Isotype Control (Cat. No. 568698; Left Plot). Flow cytometry was performed using a BD LSRFortessa™ X-20 Flow Cytometer System.
Multiparameter flow cytometric analysis using BD OptiBuild™ RB780 Rat Anti-Mouse CD274 (B7-H1) antibody (Cat. No. 755394; Right Plot) on viable C57BL/6 mouse splenocytes activated with plate-bound Anti-Mouse CD3 for 3 days, with Isotype Control (Cat. No. 568698; Left Plot). Flow cytometry was performed using a BD LSRFortessa™ X-20 Flow Cytometer System.
Product Details
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BD OptiBuild™
B7-H1, PD-L1; PD1L1; Programmed death ligand 1
Mouse (Tested in Development)
Rat SD, also known as Sprague-Dawley (outbred) IgG2a, λ
DBA/2 mouse lymphoma L5178Y transfected with Pdcd1lg1 cDNA
Flow cytometry (Qualified)
0.2 mg/ml
Aqueous buffered solution containing ≤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 that minimize unconjugated dye and antibody.

Recommended Assay Procedures

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.

Product Notices

  1. 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.
  2. Researchers should determine the optimal concentration of this reagent for their individual applications.
  3. 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.
  4. Please refer to www.bdbiosciences.com/us/s/resources for technical protocols.
  5. Please observe the following precautions: Absorption of visible light can significantly alter the energy transfer occurring in any tandem fluorochrome conjugate; therefore, we recommend that special precautions be taken (such as wrapping vials, tubes, or racks in aluminum foil) to prevent exposure of conjugated reagents, including cells stained with those reagents, to room illumination.
  6. An isotype control should be used at the same concentration as the antibody of interest.
  7. 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.
  8. Please refer to http://regdocs.bd.com to access safety data sheets (SDS).
  9. For fluorochrome spectra and suitable instrument settings, please refer to our Multicolor Flow Cytometry web page at www.bdbiosciences.com/colors.
  10. Since applications vary, each investigator should titrate the reagent to obtain optimal results.
  11. Cy is a trademark of Global Life Sciences Solutions Germany GmbH or an affiliate doing business as Cytiva.
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Antibody Details
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MIH5

The MIH5 monoclonal antibody specifically binds to CD274, also known as B7-H1 or PDL1, a 43-kDa glycoprotein encoded by the Pdcd1lg1 gene of the B7 family of the Ig superfamily. Pdcd1lg1 mRNA is expressed in more tissues than other members of the B7 family; transcripts are found in lymphoid tissues and many, but not all, non-lymphoid tissues. The protein has been detected at low levels on resting peripheral T and B lymphocytes, macrophages, and dendritic cells. B7-H1 mRNA and protein expression are upregulated upon activation of T and B cells, macrophages, dendritic cells, and epidermal keratinocytes by a variety of stimulatory factors. B7-H1's receptor, PD-1, contains an ITIM (Immunoreceptor Tyrosine-based Inhibitory Motif) on its intracytoplasmic region and is expressed on activated B and T lymphocytes, suggesting that B7-H1-PD-1 interaction may be involved in the negative regulation of immune responses. The second PD-1 ligand, B7-DC (PD-L2), is also a member of the B7 family of the Ig superfamily. Furthermore, B7-H1 may participate in positive immunoregulation, or costimulation of T cells, through an additional receptor, which is not PD-1 and distinct from the alternate receptor for B7-DC. The MIH5 antibody blocks the binding of PD-1-Ig to B7-H1 transfectants.

755394 Rev. 1
Format Details
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RB780
The BD Horizon RealBlue™ 780 (RB780) Dye is part of the BD family of blue dyes. It is a tandem fluorochrome with an excitation maximum (Ex Max) at 498-nm and an emission maximum (Em Max) at 781-nm. Driven by BD innovation, RB780 can be used on both spectral and conventional cytometers and is designed to be excited by the Blue laser (488-nm) with minimal excitation by the 561-nm Yellow-Green laser. For conventional instruments equipped with a Blue laser (488-nm), RB780 can be used as an alternative to PE-Cy7 and we recommend using an optical filter centered near 780-nm (eg, a 780/60-nm bandpass filter). For spectral instruments equipped with a Blue laser (488-nm), it can be used in conjunction with PE-Cy7. RB780 is on average brighter than PE-Cy7 and has minimal spillover into Yellow-Green detectors.
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RB780
Blue 488 nm
498 nm
781 nm
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Citations & References
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Development References (9)

  1. Ansari MJ, Salama AD, Chitnis T, et al. The programmed death-1 (PD-1) pathway regulates autoimmune diabetes in nonobese diabetic (NOD) mice. J Exp Med. 2003 July; 198(1):63-69. (Biology). View Reference
  2. Carreno BM, Collins M. The B7 family of ligands and its receptors: New pathways for costimulation and inhibition of immune responses. Annu Rev Immunol. 2002; 20:29-53. (Biology). View Reference
  3. Dong H, Chen L. B7-H1 pathway and its role in the evasion of tumor immunity. J Mol Med. 2003 May; 81(5):281-287. (Biology). View Reference
  4. Hessel EM, Chu M, Lizcano JO, et al. Immunostimulatory oligonucleotides block allergic airway inflammation by inhibiting Th2 cell activation and IgE-mediated cytokine induction. J Exp Med. 2005; 202(11):1563-1573. (Clone-specific: Flow cytometry). View Reference
  5. Liu X, Gao JX, Wen J, et al. B7DC/PDL2 promotes tumor immunity by a PD-1-independent mechanism. J Exp Med. 2003; 197:1721-1730. (Biology). View Reference
  6. Tamura H, Dong H, Zhu G, et al. B7-H1 costimulation preferentially enhances CD28-independent T-helper cell function. Blood. 2001; 97(6):1809-1816. (Biology). View Reference
  7. Tsushima F, Iwai H, Otsuki N, et al. Preferential contribution of B7-H1 to programmed death-1-mediated regulation of hapten-specific allergic inflammatory responses. Eur J Immunol. 2003; 33(10):2773-2782. (Immunogen: Blocking, Enhancement, Functional assay, Immunofluorescence, Immunohistochemistry, Inhibition, In vivo exacerbation). View Reference
  8. Wang S, Bajorath J, Flies DB, Dong H, Honjo T, Chen L. Molecular modeling and functional mapping of B7-H1 and B7-DC uncouple costimulatory function from PD-1 interaction. J Exp Med. 2003; 197:1083-1091. (Biology). View Reference
  9. Yamazaki T, Akiba H, Iwai H, et al. Expression of programmed death 1 ligands by murine T cells and APC. J Immunol. 2002; 169(10):5538-5545. (Biology). View Reference
View All (9) View Less
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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.