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The aim of this review is to give a compendium of colorimetric assays and spectrophotometric‐based quantification methods applicable to solid‐phase organic chemistry. Comprehensive experimental details for commonly employed color tests performed on solid support will be documented. © 2001 John Wiley & Sons, Inc. Biotechnol Bioeng (Comb Chem) 71:110–118, 2000/2001.

Identification of peptide substrates for proteases can be a major undertaking. To overcome issues such as feasibility and deconvolution, associated with large peptide libraries, a 'small but smart' generic fluorescence resonance energy transfer rapid endopeptidase profiling library (REPLi) was synthesised as a tool for rapidly identifying protease substrates. Within a tripeptide core, flanked by Gly residues, similar amino acids were paired giving rise to a relatively small library of 3375 peptides divided into 512 distinct pools each containing only 8 peptides. The REPLi was validated with trypsin, pepsin, the matrix metalloprotease (MMP)-12 and MMP-13 and calpains-1 and -2. In the case of calpain-2, a single iteration step involving LC-MS, provided the definitive residue specificity from which a highly sensitive fluorogenic substrate, (FAM)-Gly-Gly-Gly-Gln-Leu-Tyr-Gly-Gly-DPA-Arg-Arg-Lys-(TAMRA), was then designed. The thorough validation of this 'small but smart' peptide library with representatives from each of the four mechanistic protease classes indicates that the REPLi will be useful for the rapid identification of substrates for multiple proteases.

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Detection and quantification of material synthesized on a single resin bead is now possible using standard UV spectroscopy and mass spectrometry by the introduction of an analytical construct (A) before the linker (L2; see scheme). Cleavage of the analytical fragment is achieved by the introduction of another, orthogonal, linker (L1) that releases the labeled substrate into solution.