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PgP(G268V) Trafficking Assay

P-glycoprotein (PgP), an ATP-binding cassette transporter, plays a significant role in drug absorption and disposition. The mutant PgP(G268V) has been used as a selectivity screen for CFTR drug discovery efforts.
A PgP(G268V) Trafficking Assay is now available for compound selectivity screening. The assay is offered as a contract screening services or as product for in-house use.
Technology Overview
PgP(G268V) has been engineered with a Fluorogen Activating Peptide (FAP) which, when bound to a fluorogenic dye, provides a signal that can be quantified by high-speed flow cytometry. Using a combination of cell-excluded and cell permeant dyes, cell surface and total protein labeling can be determined.
PgPFAP  PgPFAPimage1
Left Panel: a FAP-PgP(G268V) fusion is created. An N-terminal extension is used to place the FAP in the extracellular space. Cell excluded fluorogen (red triangles) labels only the PgP(G268V) at the cell surface. Unbound fluorogen (gray triangles) remains non-fluorescent when free in solution. Right Panel: Confocal image of PgP(G268V) surface labeling.
Corrector Compounds
The assay shows the effect of corrector compounds in increasing levels of PgP at the cell surface. VRT-325 is used as a positive control for cell surface rescue. The confocal images below show the selective labeling of surface protein and the large increase in surface expression upon VRT-325 treatment. The signal is homogeneous and total fluorescence is proportional to surface protein levels so imaging is not required for quantification. Results are acquired by high-speed flow cytometry for higher throughput and the sensitivity required for the low expression levels of untreated cells.
PgPFAPimage2  PgPFAPimage3
Data generated with the PgP(G268V) Trafficking Assay quantifying effects of trafficking rescue compounds VRT-325 and VX-809 and the general trafficking enhancer C4.
How To Order
For additional information or a price quote, please contact us at sales@mayflowerbio.com and one of our representatives will contact you directly.
Fluorogen Activating Peptide (FAP) technology is covered by US Patent 8,664,364 and other US and international pending patent applications.