The first high-risk step determined by FMEA was the optimization of the bioink formulation. Without covalent attachment, and only relying on physical biomolecule transfer, the immobilization was not strong enough to withstand washing stages necessary after lubrication (Figure ). Compared to conditions where no crosslinking agent was present or only EDC was included in the bioink, we observed enhanced SNRs when both EDC and NHS were incorporated. As opposed to the O-acylisourea intermediate formed by EDC, the NHS ester offers greater stability, which facilitates more efficient binding to the primary amines of biomolecules (Figure ). The ratio of EDC to NHS necessary for effective conjugation into a crosslinking complex showed optimal values of 5mg EDC to 3mg NHS (Figure and Figure ). Furthermore, the concentration of BSA-FITC, as the biomolecule of interest for printing, was optimized to 200μg/mL resulting in the highest SNR (Figure ).