Anti-PD-1 [5C4.B8 (Nivolumab)]

Description:

Recombinant monoclonal antibody to PD-1. Manufactured using AbAb’s Recombinant Platform with variable regions (i.e. specificity) from the therapeutic antibody 5C4.B8 (Nivolumab).

Clone Name:

5C4.B8 (Nivolumab)

Published Application(s):

SPR, Block, FC, IHC

Published Species Reactivity:

Cynomolgus, Human

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Products based on this clone:

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Ab00791-13.12 Anti-PD-1 [5C4.B8 (Nivolumab)]
Cynomolgus, Human
Human IgG4-S228P
Purified
In Stock

Ab00791-23.0 Anti-PD-1 [5C4.B8 (Nivolumab)]
Cynomolgus, Human
Rabbit IgG
Purified
In Stock

Ab00791-15.0 Anti-PD-1 [5C4.B8 (Nivolumab)]
Cynomolgus, Human
Human IgM
Purified
In Stock

Ab00791-1.1 Anti-PD-1 [5C4.B8 (Nivolumab)]
Cynomolgus, Human
Mouse IgG1
Purified
In Stock

Ab00791-21.0 Anti-PD-1 [5C4.B8 (Nivolumab)]
Cynomolgus, Human
Mouse IgM
Purified
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UniProt Accession Number of Target Protein: Q15116

Alternative Name(s) of Target: 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
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).

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. PD-1 is an inhibitory receptor expressed on the surface of T cells. It is able to bind to its ligands PDL-1 and PDL-2 which results in an inhibitory signal leading to decreased T cell proliferation, cytokine protuction and cytotoxic activtiy. PDL-1 is often expressed in human tumors such as melanoma, lung and kidney where it is able to overactivate PD-1 and plays a role in the evasion of cancer cells from the immune system.

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. Nivolumab at very low concentrations (∼1.5 ng/mL) is able to enhance T-cell reactivity in the presence of a T-cell receptor stimulus - nivolumab has no stimulatory effect in the absence of antigen or T-cell receptor stimulus. In the therapeutically used human IgG4 (S228P) format, this antibody is unable to mediate ADCC (antibody-dependent cell-mediated cytotoxicity) or CDC (complement-dependent cytotoxicity).

Antibody first published in: 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.

Binding Curve

Flow-cytometry on rhesus monkey lymphocytes using Ab00791. Click on image for more details.

Binding Curve

Flow-cytometry using anti-PD-1 antibody Nivolumab (Ab00791) Rhesus monkey lymphocytes were stained with an isotype control (Ab00102-23.0, red) or the rabbit-chimeric version of Nivolumab (Ab00791-23.0, blue) at a concentration of 1 µg/ml for 30 mins at RT. After washing, bound antibody was detected using a AF488 conjugated donkey anti-rabbit antibody (ab150073) and cells analysed on a FlowJo single-cell flow cytometer.

Binding Curve

Flow-cytometry on cynomolgus monkey lymphocytes using Ab00791. Click on image for more details.

Binding Curve

Flow-cytometry using anti-PD-1 antibody Nivolumab (Ab00791) Cynomolgus monkey lymphocytes were stained with an isotype control (Ab00102-23.0, red) or the rabbit-chimeric version of Nivolumab (Ab00791-23.0, blue) at a concentration of 1 µg/ml for 30 mins at RT. After washing, bound antibody was detected using a AF488 conjugated donkey anti-rabbit antibody (ab150073) and cells analysed on a FlowJo single-cell flow cytometer.