Cat. No.
MABL-3717
Application
SPR, Block, FC, IHC
Isotype
Engineer antibody
Species Reactivity
Cynomolgus, Human
Clone No.
5C4.B8 (Nivolumab)
From
Recombinant Antibody
Specificity
Nivolumab binds to the extracellular portion of human PD-1 (2.6 nM - Scatchard analysis and SPR) - the antibody also binds to cynomolgus PD-1 with a similar affinity (3.9 nM - SPR). The antibody does not bind to other immunoglobulin superfamily proteins such as CD28, CTLA-4, ICOS and BTLA. The epitope of Nivolumab for both human and cynomolgus PD-1 includes the sequences SFVLNWYR- MSPSNQTDKLAAFPEDR (aa 29-53) and SGTYLCGAISLAPKAQIKE (aa 85-103), as shown by mass spectrometry of protease-treated fragments of PD-1 - these residues are thought to additionally be important for liand binding to PD-1.
Alternative Names
CD279; programmed death-1; programmed death 1 ; PD1; PD 1; Programmed cell death protein 1; hPD-1; hPD1; hPD 1; BMS-936558; MDX-1106;ONO-4538
UniProt
Q15116
Immunogen
mAb PD1.5 was prepared by immunizing IgH and IgK knock-out transgenic mice possessing a human immunoglobulin (heavy chain) minilocus with recombinant human PD-1-Fc protein consisting of the extracellular domain of PD-1 (amino acids 1–167) and the Fc portion of human IgG1, and Chinese hamster ovary (CHO) cells expressing human PD-1. Nivolumab was generated by grafting the variable regions of PD1.5 onto human kappa and IgG4 constant regions containing an S228P mutation (prevents Fab arm exchange with endogenous IgG4 antibodies).
Application Notes
Nivolumab has been shown to bind to PD-1-expressing CHO cells (EC50 ~1.66nM). Nivolumab binds CD4+ T cells (EC50 ~0.64 nM) and stains only memory and effector, and not naiive CD4+ or CD8+ T cells from human peripheral blood by FC. The antibody is able to block the interaction between PD-1 and its ligands PDL-1 and PDL-2 (IC50 ~2.52 nM and ~2.59 nM, respectively - determined by SPR) -these IC50 values are also similar to that measured by FACS to evaluate ligand binding to PD-1 expressed on CHO cells. In an allogenic T-cell/DC MLR, Nivolumab-mediated inhibition of PD-1 results in enhancement of IFNγ release, and also enhances IL-2 secretion (97-139% over an isotype control) in response to the superantigen SEB using human peripheral blood mononuclear cells. The same is also observed in a CMV- restimulation assay.
Antibody First Published
Wang et al. In Vitro Characterization of the Anti-PD-1 Antibody Nivolumab, BMS-936558, and In VivoToxicology in Non-Human Primates Cancer Immunol Res. 2014 Sep;2(9):846-56. PMID:24872026
Note on publication
Describes the generation and characterization of the anti-PD-1 antibody nivolumab, including its specicity, binding affinity and its in vivo and in vitro properties.
Size
100 μg Purified antibody.
Concentration
1 mg/ml.
Purification
Protein A affinity purified
Buffer
PBS with 0.02% Proclin 300.
Storage Recommendation
Store at 4⁰C for up to 3 months. For longer storage, aliquot and store at - 20⁰C.
