Synthesis, integrity, and biocompatibility of scFvD2B coated gold nanoparticles in human blood

Abstract

In this study, gold nanoparticles (AuNPs) were synthesized in water and characterized using ultraviolet-visible spectroscopy (UV-Vis), dynamic light scattering (DLS) and Zeta potential analysis. AuNPs were coated with thiol functionalized polyethylene glycol (PEG-SH) and scFvD2B (antibody fragment of D2B mAb) well known to recognize the extracellular epitope of the human prostate specific membrane antigen (hPSMA). In fact, scFvD2B binds PSMA and induces its endocytosis, thereby marking PSMA as a docking site for the delivery of therapeutic agents. AuNPs successful coating was confirmed by DLS that show an increase in the citrateAuNPs size from 26 to ~29 nm for scFvD2B-AuNPs and from 29 to ~48 nm for PEG-scFvD2BAuNPs. Moreover, Zeta potential of citrate-AuNPs increased from -34 mV to -19 mV for scFvD2B-AuNPs and from -19 mV to -3 mV for mixed PEG-scFvD2B-AuNPs. UV-Visible spectroscopy also revealed a surface plasmon resonance (SPR) band shift of about 2 nm for scFvD2B-AuNPs and 4 nm for PEG-scFvD2B-AuNPs. Likewise, conjugated AuNPs stability in vivo was characterized post-incubation with human blood plasma using gel electrophoresis, zeta potential and DLS measurements. PEG-AuNPs and PEG-scFvD2B-AuNPs showed similar reduced variation in charge and binding affinity to plasma proteins. However, citrate-AuNPs and scFvD2B-AuNPs revealed a drastic change in size compared with their pre-plasma incubation state. Additionally, neutrophil function test and pyridine formazan extraction showed lower neutrophil activation level of 9 and 10% with PEG- and PEG-scFvD2B-AuNPs, respectively; compared with a 14% activation level with both citrate and scFvD2B-AuNPs. All AuNPs were blood compatible with less than 10% hemolysis. In conclusion, our data provide a novel scFvD2BAuNPs (± PEG) with proof that PEG-scFvD2B-AuNPs serve as promising vehicles for drug delivery with minimal protein adsorption affinity, insignificant charge and size variation, low immunorecognition, and reduced hemolytic activity.