title: Preparation and characterization of nickel nanoparticles for binding to his-tag proteins and antigens.
abstract: OBJECTIVE
  The purpose of these studies was to prepare nanoparticles (NPs) with a small amount of surface-chelated nickel for obtaining enhanced binding of histidine-tagged (his-tag) proteins compared to non-histidine-tagged protein binding to charged nanoparticles.
  
  
  METHODS
  NPs were prepared from oil-in-water microemulsion precursors using emulsifying wax, 3 mM Brij 78 and 0.1 mM DOGS-NTA-Ni lipid (referred to as Ni-NPs). The amount of lipid entrapped in the NPs was quantitated by atomic emission spectroscopy (AES). The Ni-NPs were investigated for binding to two his-tag proteins, green fluorescent protein (GFP) and his-tag HIV-1 Gag p24. In vivo studies in mice were carried out to evaluate the immune responses obtained to his-tag Gag p24 bound to Ni-NPs.
  
  
  RESULTS
  AES studies demonstrated that approximately 5% of the DOGS-NTA-Ni lipid used was entrapped in the NPs. The optimal binding ratio his-tag GFP and his-tag Gag p24 to Ni-NPs was found to be 1:33.7 and 1:35.4 w/w, respectively. This interaction was stable at 37 degrees C in PBS, pH 7.4 over 4 h and the interaction of his-tag GFP with the Ni-NPs was enhanced compared to control NPs prepared with no Ni on the surface (NTA-NPs). The in vivo studies demonstrated enhanced serum IgG and IgG2a responses to his-tag Gag p24 bound to Ni-NPs compared to protein adjuvanted with Alum or adsorbed on the surface of control NTA-NPs.
  
  
  CONCLUSIONS
  Ni-NPs can be used to bind strongly to his-tag proteins. This system was demonstrated to have potential applications in vaccine delivery for enhancing immune responses to protein-based vaccines.
authors.affiliation: Department of Pharmaceutical Sciences, College of Pharmacy, University of Kentucky, Lexington, KY 40536-0082, USA.
authors.forename: Jigna D
authors.initials: JD
authors.lastname: Patel
authors.forename: Ronan
authors.initials: R
authors.lastname: O'Carra
authors.forename: Julia
authors.initials: J
authors.lastname: Jones
authors.forename: Jerold G
authors.initials: JG
authors.lastname: Woodward
authors.forename: Russell J
authors.initials: RJ
authors.lastname: Mumper
mesh_terms: D000941:Antigens / Q000737:chemistry*; D015153:Blotting, Western; D018791:CD4 Lymphocyte Count; D049109:Cell Proliferation / Q000187:drug effects; D002626:Chemistry, Pharmaceutical; D004655:Emulsions; D004797:Enzyme-Linked Immunosorbent Assay; D049452:Green Fluorescent Proteins / Q000737:chemistry; D016655:HIV Core Protein p24 / Q000737:chemistry; D006639:Histidine / Q000737:chemistry*; D006863:Hydrogen-Ion Concentration; D007074:Immunoglobulin G / Q000096:biosynthesis; D007371:Interferon-gamma / Q000378:metabolism; D053758:Nanoparticles; D009532:Nickel / Q000737:chemistry*; D011092:Polyethylene Glycols; D011485:Protein Binding; D011506:Proteins / Q000737:chemistry*; D013054:Spectrophotometry, Atomic; D013154:Spleen / Q000166:cytology; D014779:Virus Replication / Q000187:drug effects; D014885:Waxes
chemical_list: D000941:Antigens; D004655:Emulsions; D016655:HIV Core Protein p24; D007074:Immunoglobulin G; D011506:Proteins; D014885:Waxes; D049452:Green Fluorescent Proteins; D011092:Polyethylene Glycols; D006639:Histidine; D009532:Nickel; D007371:Interferon-gamma; C043444:octadecyl polyoxyethylene ether