These advanced biotinylation reagents contain UV-traceable benzophenone moietiy with absorption maximum of 350 nm, surrounded by two long hydrophilic PEG4 spacer arms. The hydrophilic arm enhances water solubility while reducing the tendency of some proteins to aggregate/precipitate during biotinylation. Extended PEG spacers improve conjugate yield while reducing non-specific interactions for enhanced assay signal at higher biotin loads. After biotinylation, the conjugate's absorbance at 280 nm and 350 nm is used to rapidly and quantitatively measure the level of biotin incorporation. Use these UV Tracer reagents for improved labeling reproducibility and higher yields at higher biotin loads for maximum assay sensitivity and robustness.