Combinatorial beads

Recently we have employed the COPAS Biobead bead sorter (Union Biometrica, Somerville, MA) to isolate positive beads from combinatorial libraries. Combinatorial libraries are incubated as described above with a fluorescently tagged protein. The beads are washed to remove unbound proteins and then sorted based on the fluorescence intensity of the protein bound to individual beads. Combinatorial beads can be sorted at a rate of 50-100 thousand beads per hour and the positive beads dispensed into 96-well plates. If necessary, the combinatorial beads can be prescreened to remove beads with excessive fluorescence background. 

OBOC combinatorial bead-libraries can be considered as chemical microarrays that are spatially separable but non-addressable. The identity of the chemical compound on the positive beads can be determined directly with an automatic sequencer if it is an N-terminally unprotected peptide, by mass spectroscopy, or through chemical encoding. i A synthetic scheme for the OBOC library is shown in Figure 2. Using the highly efficient split-mix synthesis method,literally hundreds of thousands to millions of compounds can be prepared within a week. The recent... 

Combinatorial chemistry has significantly increased the nurnjjers of molecules that can be synthesised in a modern chemical laboratory. The classic approach to combinatorial synthesis involves the use of a solid support (e.g. polystyrene beads) together with a scheme called split-mix. Solid-phase chemistry is particularly appealing because it permits excess reagent to be used, so ensuring that the reaction proceeds to completion. The excess... 

Small-diameter packed columns offer (17) the substantial advantages of small volumetric flow rates (1-20 (p.L min )), which have environmental advantages, as well as permitting the use of exotic or expensive mobile phases. Peak volumes are reduced (see Table 1.1), driven by the necessity of analysing the very small (pico-mole) amounts of substance available, for example, in small volumes of body fluids, or in the products of single-bead combinatorial chemistry. 

By this time Polya s Theorem had become a familiar combinatorial tool, and it was no longer necessary to explain it whenever it was used. Despite that, expositions of the theorem have continued to proliferate, to the extent that it would be futile to attempt to trace them any further. I take space, however, to mention the unusual exposition by Merris [MerRSl], who analyzes in detail the 4-bead 3-color necklace problem, and interprets it in terms of symmetry classes of tensors — an interpretation that he has used to good effect elsewhere (see [MerRSO, 80a]). 

Two main approaches to combinatorial chemistry are used—parallel synthesis and split synthesis. In parallel synthesis, each compound is prepared independently. Typically, a reactant is first linked to the surface of polymer beads, which are then placed into small wells on a 96-well glass plate. Programmable robotic instruments add different sequences of building blocks to tfie different wells, thereby making 96 different products. When the reaction sequences are complete, the polymer beads are washed and their products are released. 

Introduced in the early 1990s, the split-and-recombine concept contributed much to the early success of combinatorial chemistry. Often, all combinatorial methods were identified with this concept. Split-and-recombine synthesis offered easy access to large number of individual compounds in few steps. If conducted on polymer beads, these are easily separated mechanically and can be identified subsequent to a screening step. 

Combinatorial Chemistry. Figure 2 Chemical libraries are prepared either by parallel synthesis or by the split-and-recombine method. In the latter case, coupling m building blocks in m separated reaction flasks through n synthetic cycles on a beaded polymer carrier generates a combinatorial library with nf individual compounds and one compound per bead. 

Because of their ease of synthesis and their structural similarity to peptides, many laboratories have used peptoids as the basis for combinatorial drug discovery. Peptoids were among the first non-natural compounds used to establish the basic principles and practical methods of combinatorial discovery [17]. Typically, diverse libraries of relatively short peptoids (< 10 residues) are synthesized by the mix-and-split method and then screened for biological activity. Individual active compounds can then be identified by iterative re-synthesis, sequencing of compounds on individual beads, or indirect deduction by the preparation of positional scanning libraries. 

Certain types of molecules, especially polypeptides and polynucleotides, lend themselves to synthesis on solid supports. In such syntheses, the starting material is attached to a small particle (bead) or a surface and the molecule remains attached during the course of the synthetic sequence. Solid phase synthesis also plays a key role in creation of combinatorial libraries, that is, collections of many molecules synthesized by a sequence of reactions in which the subunits are systematically varied to create a range of structures (molecular diversity). 

Park SI, Renil M, Vikstrom B et al (2002) The use of one-bead one-compound combinatorial library method to identify peptide ligands for cz4f31 integrin receptor in non-Hodgkin s lymphoma. Lett Pept Sci 8 171-178... 

Jacobsen proposed a related heterogeneous catalyst in which the complex is bound via an ester linkage to commercial hydroxymethylpolystyrene beads. Such catalysts are of value not only in large-scale processes, but also in the combinatorial automated synthesis of libraries of new compounds.180... 

Swali, V., Wells, N.J., Langley, G.J., and Bradley, M. (1997) Solid-phase dendrimer synthesis and the generation of super-high-loading resin beads for combinatorial chemistry. J. Org. Chem. 62, 4902-4903. 

Effective syntheses, convergent as well as batch-splitting, are possible using these methods, which can provide both combinatorial arrays and fragment sets of new molecules (Fig. 2). The act of immobilisation can be accomplished in a variety of ways such as on beads, active... 

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