The cetuximab-IRDye800 immunoconjugate was produced under cGMP conditions at the University of Alabama (UAB) Vector Production Facility before shipment to Stanford University Hospital Pharmacy, which is nearly identical to that found in the literature.47 (link) In brief, cetuximab (ImClone LLC, Eli Lilly and Company) was provided as a 2 mg/mL solution and concentrated, and the pH was adjusted by buffer exchange to a 10 mg/mL solution in 50 mmol/L potassium phosphate, pH 8.5. IRDye 800CW NHS ester (LI-COR Biosciences, Lincoln, NE) underwent conjugation to cetuximab for 2 h at 20 °C in the dark, whereby an average molar ratio of 1:2.3 cetuximab:IRDye800 was attained. After column filtration to remove unconjugated dye and exchange buffer to phosphate-buffered saline (PBS), pH 7, final protein concentration adjusted to 2 mg/mL, the product was sterilized by filtration and placed into single-use vials. After conjugation, the agent was shipped to Stanford University Hospital Pharmacy. During shipping of the immunoconjugate, the temperature was stable at 4 °C. The immunoconjugate was stored at 4 °C until used upon its receipt. The same lot was used for the cohort of animals in the treatment group.