How to Choose a Suitable Fluorescent Reagent for Your Peptide
It is based on your experimental needs.
Molecular imaging techniques play an important role in modern diagnostics for their high specificities. They can provide biological information at the molecular level in living systems, enable visualization some of the specific molecular events in disease process and monitor therapeutic responses. As one of important molecular imaging technology, fluorescent imaging has its specific advantages over others, such as high sensitivity, using nonradioactive materials, safe detection by instruments with moderate cost.
Biologically active peptides can be labeled with a fluorophore and applied in the fluorescent imaging fields. The fluorophore molecule has the ability to absorb photons of energy at one wavelength and emit the energy at a longer wavelength immediately. There are several criteria for measuring its dye ability and efficiency (absorbing photons and emitting photons, and repeating the cycles). Among them, molar extinction coefficientεand quantum yield (QY) are two important factors which determine the overall fluorescent intensity of a fluorophore and correlate with absorption and fluorescence. Normally, 5000~250000 cm-1 M-1 forεand 0.05-1.0 for QY will meet the requirement.
Choose a suitable fluorophore for research is based on your experimental needs. The selection can be simply divided into two categories: in vivo and in vitro applications. For in vivo research, fluorophores with emisstion wavelength in the near-infrared region (650-900 nm) are normally recommended, such as ICG, Cy5.5 analogs, Nile Blue. They have better tissue penetration and interfere less with the back ground generated from water, hemoglobin and deoxyhemoglobin (absorbance 560nm). For in vitro research, fluorophores with emisstion wavelength between 400 and 600 nm are overwhelmingly used, such as AMC, FITC and TAMARA.
The fluororescent peptides have been widely applied in modern pharmaceutical research. For example, Cy5.5 conjugated c-Met binding peptide (cMBP) was designed for investigate the possibility of c-Met receptor targeting. cMBP with sequence KSLSRHDHIHHH containing GGGSC as a linker at C terminal. It was labeled with Cy5.5-Maleimide through Cys side chain. The binding affinity of Cy5.5-cMBP was in the nanomolar range to U87MG cells and the probe displayed high tumor uptake at 24 hours postinjection in a U87MG tumor model.