Molecular Probes™ pHrodo™ Red and Green Epidermal Growth Factor (EGF) and pHrodo™ Red Transferrin Conjugates

The pHrodo Red and pHrodo Green Epidermal Growth Factor (EGF) conjugates consist of pHrodo Red or pHrodo Green dye directly conjugated to a highly active human recombinant EGF molecule and are used for the detection of internalized EGF receptor in cells. Fluorescently labeled EGF has enabled scientists to investigate receptor-membrane interactions, study receptor distribution, calculate rate constants for the interaction of EGF with its receptor, and more. Fluorescent labels that respond to a pH change add another level of detection.

pHrodo Red Transferrin Conjugate consists of pHrodo Red dye directly conjugated to a highly active transferrin molecule and is used for the detection of internalized transferrin in cells. Transferrin is a monomeric serum glycoprotein (∼80,000 daltons) that binds to a specific receptor on the surface of vertebrate cells and delivers up to two Fe³⁺ atoms via receptor-mediated endocytosis. Once iron-carrying transferrin proteins are inside endosomes, the acidic environment favors dissociation of iron from the transferrin–receptor complex. Following the release of iron, the apotransferrin is recycled to the plasma membrane, where it is released from its receptor to scavenge more iron.

The highly photostable pHrodo Red and pHrodo Green dyes are weakly fluorescent outside of cells at neutral pH, but when internalized specifically through EGF or transferrin endocytosis, become brightly fluorescent in acidic endosomes. This allows the measurement of internalized EGF or transferrin, as well as the dynamic monitoring of EGF or transferrin internalization in live cells.

The pHrodo Red EGF Conjugate and the pHrodo Red Transferrin Conjugate have excitation/emission spectra of 560/585 nm and can be multiplexed with a wide variety of blue, red, and far-red dyes reporters such as Mitosox Red, CellEvent Caspase 3/7 Red, NucBlue, RFPs, and Mitotracker Deep Red, among many others. The pHrodo Green EGF Conjugate has excitation/emission spectra of 509/533 nm and can be multiplexed with a wide variety of blue, red, and far-red dyes reporters such as Mitosox Red, CellEvent Caspase 3/7 Red, NucBlue, RFPs, and Mitotracker Deep Red, among many others.

The pHrodo Red and pHrodo Green EGF conjugates and the pHrodo Red Transferrin Conjugate are compatible in applications including traditional fluorescence microscopy, high content screening (HCS), flow cytometry, and microplate-based fluorimetry or high throughput screening (HTS). The conjugates are also compatible with various widefield or confocal fluorescence microscopes, flow cytometers, fluorescence plate readers, and high content instruments.