High purity, high bioactivity immune checkpoint proteins

Immune checkpoint proteins are considered to be the main regulators of the immune response against threats and maintain self-tolerance. The classic immune checkpoint includes co-inhibitory receptors (CTLA-4, PD-1, TIGIT, LAG-3) that suppress T-cell function and co-stimulatory receptors (GITR, 4-1BB, OX40, CD40) that tend to trigger T cell activation.

Analysis on the co-inhibitory receptor CTLA-4 (cytotoxic T-lymphocyte antigen 4) and PD-1 (programmed cell death protein 1) demonstrated considerably enhanced survival rates of patients with metastatic solid tumors following the suppression of such receptor pathways. Therefore, antibody drugs against immune checkpoints for tumors, cancer and other disease therapeutics are a primary mode in cancer therapy.

Pathways of immune checkpoint

Image Credit: ACROBiosystems

Product features

  • High purity verified by HPLC or MALS
  • High bioactivity verified by ELISA/SPR/FACS, etc.
  • High batch-to-batch consistency ensured by stringent QC testing

Main applications

  • Antibody binding assay
  • Antibody affinity assay
  • Neutralizing antibody binding assay
  • Cell-based assay

TNFSF proteins. Image Credit: ACROBiosystems

CD47 proteins. Image Credit: ACROBiosystems

CTLA-4 homodimer proteins. Image Credit: ACROBiosystems

B7-CD28 family proteins. Image Credit: ACROBiosystems

Key features

Authentic structure verified by SEC-MALS

The purity of Human CD27 Ligand, His, Flag Tag (active trimer) (MALS verified) (Cat. No. CDL-H52Da) was more than 95% in HP-SEC, and around 55-70 kDa verified by SEC-MALS. Image Credit: ACROBiosystems

High batch-to-batch consistency

The uniformity in purity and bioactivity is vital for recombinant proteins. All the proteins are in need to pass a range of purity and bioactivity assessments.

Biotinylated Human PD-1 / PDCD1 Protein, Avitag™, His Tag (recommended for biopanning) (MALS verified)Cat. No. PD1-H82E4. Image Credit: ACROBiosystems

Application 1: Development and screening of antibody drugs

Antibody binding assay

Immobilized Human PD-1, His Tag (Cat. No. PD1-H5221) at 2 μg/mL (100 μL/well) can bind Nivolumab with a linear range of 0.1-3 ng/mL (Routinely tested). Image Credit: ACROBiosystems

Serial dilutions of nivolumab were added into Human PD-1, Fc Tag, low endotoxin (Cat. No. PD1-H5257): Biotinylated Human PD-L1, Fc, Avitag, His Tag (Cat. No. PD1-H82F3) binding reactions. The half maximal inhibitory concentration (IC50) is 0.5381 μg/mL (Routinely tested). Image Credit: ACROBiosystems

Moreover, ACROBiosystems have also developed PD1: PD-L1 (Cat. No. EP-101) and CD47: SIRP alpha (Cat. No. EP-102) inhibitor screening assay kits that can perform high-throughput screening of neutralizing antibodies or small molecules.

Antibody drug affinity assay

Anti-LAG-3 MAb (Human IgG1) captured on CM5 chip via Anti-Human IgG Fc antibodies surface, can bind Human LAG-3, His Tag (Cat. No. LA3-H5222) with an affinity constant of 4.57 nM as determined in a SPR assay (Biacore T200) (Routinely tested). Image Credit: ACROBiosystems

Loaded Anti-LAG-3 MAb (Human IgG1) on AHC Biosensor, can bind Human LAG-3, His Tag (Cat. No. LA3-H5222) with an affinity constant of 7.47 nM as determined in BLI assay (ForteBio Octet Red96e) (Routinely tested). Image Credit: ACROBiosystems

Neutralizing antibody binding assay

Serial dilutions of Human TIGIT Neutralizing antibody were added into Human TIGIT, Fc Tag (Cat. No. TIT-H5254): Biotinylated Human CD155, Fc, Avitag (Cat. No. CD5-H82F6) binding reactions. The half maximal inhibitory concentration (IC50) is 0.55834 μg/mL (Routinely tested). Image Credit: ACROBiosystems

FACS analysis shows that the binding of Human CD47 to ACHN expressing SIRP-a was inhibited by increasing concentration of neutralizing SIRP-a antibody. The concentration of Human CD47 used is 20 μg/mL. IC50 = 9.334 μg/mL. Image Credit: ACROBiosystems

Cell-based assay

Human 4-1BB Ligand (71-254), Fc Tag, active trimer (Cat. No. 41L-H5265) induce IL-8 secretion in HT1080 human CD137 cell line. The EC50 for this effect is 0.079 μg/mL (Routinely tested). Image Credit: ACROBiosystems

Pharmacokinetics (PK) analysis

Immobilized Human CTLA-4, His Tag, active dimer (Cat. No. CT4-H52H9) at 2 μg/mL, add increasing concentrations of CTLA-4 x OX40 Bispecific Antibody in 50% Human serum and then add Biotinylated Human OX40, Avitag, His Tag(Cat. No. TN4-H82E4) at 1 μg/mL. Detection was performed using HRP-conjugated streptavidin with sensitivity of 4 ng/mL (Intact assay, Routinely tested). Image Credit: ACROBiosystems