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Mix-and-Read, Bead- and Cell-Based Immunoassays Using the FMAT 8100 HTS System

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The increasing demands for higher throughput in HTS assays has resulted in greater exploration of the ability of homogeneous, or mix-and-read assays to eliminate time-consuming or hard-to-automate wash steps but still provide quality leads. The FMAT 8100 HTS System addresses these needs by enabling both fluorescent live-cell and bead-based assays in a mix-and-read format. Thanks to the FMAT systems simple format, miniaturization in 384-well plates is now easy. Higher throughput can be achieved by the instruments color- and size-based multiplexing capabilities.

A wide variety of HTS assays have already been adapted for the FMAT system, including live-cell receptor/ ligand binding4, apoptosis5, immunofluorescence5, and bead-based fluorescent immunoassays (FLISAs)3.

In the experiments discussed here, immunofluorescence and FLISA assays were combined in a single well, and the secreted cytokines and cell surface expression of an inducible marker were quantitated simultaneously. The generation of standard curves in buffers that match likely test samples, demonstrates that quantitative FLISA measurements can be obtained using tissue culture media with cells in the well. The ability to perform mix-and-read, beadand cell-based assays simultaneously greatly simplifies HTS while increasing yield and data quality.

References

1 Pober, J.S. and Cotran, R.S. (1990) Physiol. Rev. 70, 42751.

2 Chen, C.C. and Manning, A.M. (1996) Cytokine 8, 5865.

3 Swartzman, E. E., Miraglia, S. J., Mellentin-Michelotti, J., Evangelista, L., and Yuan, P. M. (1999) Anal. Biochem. 271, 14351.

4 Mellentin-Michelotti, J., Evangelista, L.T., Swartzman, E.E., Miraglia, S.J., Werner, W.E., and Yuan, P.M. (1999) Anal. Biochem. 272, 182190.

5 Miraglia, S., Swartzman, E. E., Mellentin-Michelotti, J., Evangelista, L., Smith, C.,
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