High Throughput Screening

The sets of compounds produced by combinatorial chemistry are generally referred to as libraries, which depending on how the solid-phase is handled, may be either mixtures or individual compounds. There are a range of options for testing the libraries in a biological assay:

Test mixture in solution
All the compounds are cleaved from the beads and tested in solution. If the resin beads were intimately mixed, it is not possible to test the products separately, but rather as a mixture. If activity in a pharmacological screen is observed it is not possible to say which compound or compounds are active. In order to identify the most active component, it is necessary to resynthesize the compounds individually and thereby find the most potent. This iterative process of resynthesis and screening is one of the most simple and successful methods for identifying active compounds from libraries.

Test individual compounds in solution
A second method is to separate the beads manually into individual wells and cleave the compounds from the solid-phase. These compounds can now be tested as individual entities.

Test compounds on the beads
A third method for screening is testing on the beads, using a colorimetric or fluorescent assay technique. If there are active compounds, the appropriate beads can be selected by color or fluorescence, ‘picked’ out by micromanipulation and the product structure, if a peptide, determined by sequencing on the bead. Non-peptide structures would need to be identified by one of the tagging methods. Screening on the bead may be an inappropriate method for drug discovery, as the bead and linker present conformational restrictions that may prevent binding to the receptor. Furthermore for pharmaceutical applications compounds will be invariably need to act, and thus ideally need to be tested in solution.

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