Expression of Recombinant Human Programmed Cell Death Protein-1 and Development of Camel Polyclonal Antibody

Aims Programmed cell death protein-1 (PD-1) is a membrane receptor expressed on the surface of T and B lymphocytes, monocytes, natural killers, and dendritic cells. In cancer, the PD-1/PD-L1 system prevents the proliferation of T lymphocytes and causes the release of cytokines and cytotoxicity, which leads to the apoptosis of tumor-specific T cells, thereby preventing the immune response to cancer cells.  Methods & Materials In this study, the extracellular part of the humanized PD-1 protein was cloned and expressed, and the protein was injected as an antigen into a camel (Camelus dromedarius) to obtain a camel polyclonal antibody against PD-1 protein.  Findings The obtained results indicate the proper expression of the protein in the prokaryotic system. Also, using various tests, such as ELISA and western blot, it was confirmed that the polyclonal antibody obtained from camel can identify PD-1 protein.  Conclusion This study showed that because of the advantages, such as the ability to bind multiple epitopes, camel polyclonal antibodies can be used in antibody-based research for effective and strong molecular applications to detect PD-1 receptors

Characteristics and Lethality of a Novel Recombinant Dermonecrotic Venom Phospholipase D from Hemiscorpius lepturus

Hemoscorpius lepturus is the most medically important scorpion in Iran. The clinical signs of H. lepturus envenomation are remarkably similar to those reported for brown spiders, including dermonecrosis, hematuria, renal failure and even death. The lethality and toxicity of brown spiders’ venom have been attributed to its phospholipase D activity. This study aims to identify a phospholipase D with possible lethality and dermonecrotic activity in H. lepturus venom. In this study, a cDNA library of the venom glands was generated by Illumina RNA sequencing. Phospholipase D (PLD) from H. lepturus was characterized according to its significant similarity with PLDs from brown spiders. The main chain designated as Hl‐RecPLD1 (the first recombinant isoform of H. lepturus PLD) was cloned, expressed and purified. Sphingomyelinase, dermonecrotic and lethal activities were examined. Hl‐PLD1 showed remarkable sequence similarity and structural homology with PLDs of brown spiders. The conformation of Hl‐PLD1 was predicted as a “TIM beta/alpha‐barrel”. The lethal dose 50 (LD50) and dermonecrotic activities of Hl‐RecPLD1 were determined as 3.1 μg/mouse and 0.7 cm2 at 1 μg respectively. It is the first report indicating that a similar molecular evolutionary mechanism has occurred in both American brown spiders and this Iranian scorpion. In conclusion, Hl‐RecPLD1 is a highly active phospholipase D, which would be considered as the lethal dermonecrotic toxin in H. lepturus venom