Fluorescein-methotrexate

N., Horie, T., Characteristics of transport of fluoresceinated methotrexate in rat small intestine, Life Sci. 2001, 69, 739-747. 

More recently Michnick and co-workers have introduced a dihydrofolate reductase complementation system, which seems to be particularly robust [61 - 65], They attribute the success of this system to the fact that the N-terminal (1 - 105) and C-terminal (106 - 186) DHFR fragments do not fold until they are dimerized. In addition to the obvious selection for essential metabolites dependent on the reduction of dihydrofolate to tetrahydrofolate, protein-protein interactions are detected based on the retention of a fluorescein-methotrexate conjugate. Several other enzymes have been employed for the design of complementation assays, including green fluorescent protein, which allows screens based on fluorescence or FRET [66 - 68]. As with the bacterial transcription assays, these complementation systems are new. It will be interesting to see if, as the selections are optimized, these systems prove competitive with the Y2H assay. 

Figure 43 Folate targeted PAMAM dendrimers. Dose-dependent binding of G5-FI-FA-MTX in KB cells. The cells were maintained in FA-free medium and incubated with different concentrations of the indicated dendrimers for 1 h. The cells were then rinsed and resuspended in PBS buffer, and the fluorescence was measured in a flow cytometer (a). In panel b, the mean cell fluorescence, after being normalized for the fluorescence of standard solutions of the dendrimers, is presented. FA, folic acid MTX, methotrexate. FI, fluorescein isothiocyanate. Reproduced with permission from Thomas, T. P. Majoros, I. J. Kotlyar, A. etal. J. Med. Chem. 2005, 48, 3729 310 Copyright 2005 American Chemical Society.

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