How do you prepare a protein fluorescein conjugate from FITC?
Dissolve the FITC in anhydrous DMSO at 1 mg/ml. Note: This should be prepared fresh for each labeling reaction. 3. For each 1 ml of protein solution, add 50 µl of FITC solution, very slowly in 5 µl aliquots while gently and continuously stirring the protein solution.
Why is FITC used?
FITC is used for labeling of different biomolecules, e.g. immunoglobulins, lectins and other proteins, peptides, nucleic acids, nucleotides; oligo-and polysaccha- rides.
Is FITC the same as fluorescein?
FITC (Fluorescein isothiocyanate) is the form of fluorescein used for conjugation to all JIR antibodies and purified proteins, with the exception of streptavidin. Fluorescein conjugates absorb light maximally at 492 nm and fluoresce maximally at 520 nm.
What is meant by FITC?
Fluorescein isothiocyanate (FITC) is a derivative of fluorescein used in wide-ranging applications including flow cytometry. First described in 1942, FITC is the original fluorescein molecule functionalized with an isothiocyanate reactive group (−N=C=S), replacing a hydrogen atom on the bottom ring of the structure.
How do you label a protein with FITC?
3-In routine, you can want to label between 0.5 and 2mg protein. Calculate the quantity of FITC required, by using a molar ratio FITC/ prot of 100. For example, if you want to graft 1mg of protein of 20kDa, the FITC quantity required is 1.9mg.
What wavelength is FITC?
FITC emits fluorescence from 475 to 650 nm, peaking at 525 nm, which falls in the green spectrum.
How do you use FITC?
Covalent conjugation. FITC is covalently coupled to primary amines (lysines) of the immunoglobulin. Dissolve 10 mgs (the entire contents of 1 vial; no need to weigh) of FITC in 1 mL anhydrous DMSO immediately before use. Add FITC to give a ratio of 40-80 µg per mg of antibody; mix immediately.
Is FITC blue or green?
Is FITC a GFP?
Available Options. Green Fluorescent Protein (GFP) Antibody (FITC) is a Rabbit Polyclonal antibody conjugated to FITC against GFP.
How does FITC react with proteins?
FITC reacts with a primary amine on the protein to form a covalent amide bond. FITC-labelled protein substrates/peptides, antibodies, peptide hormones are used as specific probes in enzyme kinetics, immunocytochemistry as well as flow cytometry and identification of receptors on target cells, respectively.
What is covalent conjugation of FITC?
Covalent conjugation. FITC is covalently coupled to primary amines (lysines) of the immunoglobulin. Dissolve 10 mgs (the entire contents of 1 vial; no need to weigh) of FITC in 1 mL anhydrous DMSO immediately before use.
What antibodies can be conjugated with FITC?
FITC can be easily conjugated with antibodies against tags such as FLAG, His, c-Myc, and with several others. In 2011, SJ Fleishman used FITC-conjugated c-Myc antibody in his flow cytometry computational experiment for the treatment of influenza.
What is the use of ab102884 FITC conjugation kit?
FITC Conjugation Kit / FITC Labeling Kit ab102884 uses a simple and quick process for FITC labeling / conjugation of antibodies. It can also be used to conjugate other proteins or peptides.
What is the difference between TCP and FITC reactions?
Different methods for reacting a TCP (target cellular protein) with a fluorophore like fluorescein. Whereas FITC can directly react with amine residues on the target protein, other methods utilize multiple steps; this is why FITC is such a commonly utilized fluorophore. Image source: Acta Naturae.