uated independently of the amount of murine IgG secreted into the media. Indeed, reliable data can be obtained for wells containing as high as 22 g/mL IgG to as low as 0.16 g/mL IgG.

Conclusion

The hybridoma screening assay developed for the FMAT system was simple to develop, set-up, and execute. Cells for the fluorometric microvolume assay were plated just minutes prior to the screen, unlike the cell ELISA where cells had to be plated 48 hours prior to the screen. This eliminates any edge effects due to uneven cell growth across the plate and reduces cell culture burden. Because of the mix-and-read format of the assay, all cell types (suspension or adherent) can be used without further manipulations. With a cell ELISA, suspension or weakly adherent cells must be fixed by some means to the well surface and thereby introduces potentially detrimental variables to the screen. Besides cell-surface antigens, the FMAT system can be used to screen for monoclonal antibodies directed against soluble antigens by using polystyrene beads coated against the antigen (Figure 5). With both the cell- and bead-based assays, no wash steps and only one incubation step are required. Moreover, only 5 L of the hybridoma supernatant is required for the assay, enabling parallel multi-specificity screening at the fusion plate stage. In contrast, the cell ELISA typically requires 50 to 100 L of hybridoma supernatant. In addition, the FMAT system allows for multiplexed assays based on bead size or dye color, enabling more than one assay to be performed in a single well. With its ease-of-use, flexibility and sensitivity, the FMAT system is ideally suited for hybridoma screening.
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