Fluorescent Labeling Reagents

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Fluorescent labeling reagents are compounds with a chemical structure of a conjugated double bond system. When irradiated by ultraviolet light or blue-violet light, they can be excited into an excited state, and when the excited state is restored to the ground state, it emits fluorescence. The advantages of fluorescent labeling reagents, such as no radioactive pollution and easy operation, make the application of fluorescent labeling reagents in many research fields increasingly widespread.

Introduction and application of FITC and TRITC

Fluorescein isothiocyanate (FITC) is an organic fluorescent dye. At 495/517nm, the dye produces excitation/emission peaks, and can bind to different antibodies by means of an isothiocyanate reactive group. It can be combined with amino, sulfhydryl, imidazole, tyrosyl, carbonyl and other groups on protein. Another dye that is often used at the same time as FITC is TRITC, tetramethylrhodamine-5(6)-isothiocyanate. TRITC is a derivative of the rhodamine family and also has a large conjugated aromatic electronic system, which triggers their fluorescence behavior. TRITC (479g/mol) is excited by light in the green spectrum with a maximum wavelength of 550nm, and its maximum emission wavelength is 573 nm. Binding to proteins (eg, antibodies) is also based on isothiocyanate reactive groups.

Introduction and application of cyan pigment

Such fluorescent dyes are relatively rare, derived from cyan pigments, including Cy2, Cy3, Cy5 and Cy7. All the above-mentioned cyan pigments can be linked to nucleic acids or proteins through their reactive groups. For example, a maleimide group labeled with a protein is used. Cy5 is very sensitive to its surrounding electronic environment, and this feature can be used for enzyme determination. The conformational change of the attached protein will cause a positive or negative change in fluorescence emission. In addition, Cy3 and Cy5 can also be used for FRET tests.

Definition and application of FAM dyes

Carboxyfluorescein (FAM) is a kind of fluorescein derivative. Compared with FITC, FAM reacts faster with amino groups and the product is more stable, but the amount of FITC binding protein is larger and the process is easier to control. FAM is also suitable for the 488 nm spectral line of Argon-ion Laser, Abs/Em=492/518nm (pH=9.0), which has the general characteristics of fluorescein derivatives and is stable in water. 5-FAM is mainly used in automatic DNA sequencing to label the d/ddCTP, and it is also often used in PCR product quantification, nucleic acid probes, etc.

Introduction and application of DNA staining

Sometimes, the precise location and number of cells must be determined by examining the nucleus. One of the most commonly used DNA stains is DAPI (4',6-diamidino-2-phenylindole), which can be combined with the A-T rich region of the DNA double helix. The dye is excited by ultraviolet light with a maximum wavelength of 358 nm, and its emission spectrum is very broad, reaching a peak at 461 nm. In addition, RNA binding detection can be performed on weak fluorescence. In addition, Hoechst dye and Propidium-Iodide (PI) are also used in DNA staining.

Creative Peptides is committed to providing customers with a rich variety of fluorescent labeling reagents.