Chromium-Loaded CPMV Nanoparticle Purification

CPMV nanoparticles were purified from Vigna unguiculata plants infected by mechanical inoculation using a 0.1 mg/mL solution of CPMV in 0.1 M potassium phosphate buffer. Yields of 1 mg/g of infected leaf material were obtained using established procedures.^{87 (link)} For chromium loading, 5 mg/mL CPMV was incubated with 50 mM CrCl₃ (Sigma Aldrich) and 30 mM ethylenediaminetetraacetic acid (EDTA) in 50 mM HEPES buffer, pH 7 overnight (^~60,000-fold molar excess). The sample was then dialyzed against 10 mM HEPES buffer with 10 mM EDTA using 12–14 kDa molecular weight cut-off tubing (Spectrum Laboratories) over 3 days. Some precipitation from the presence of NaOH in the buffer was observed over time and removed by a clearing spin at 10,000 rpm for 10 min. The release profile was studied by taking aliquots of the sample over a period of 7 days; the amount of drug in the sample was then determined using ICP-OES (see below).