Dye Series 5-FITC 3326-32-7
Not For Human Use, Lab Use Only.
FITC can bind to various antibody proteins, and the bound antibody does not lose the specificity of binding to a certain antigen, and has strong yellow-green fluorescence in alkaline solution.
5-FITC Cas No:3326-32-7| 95%+ |Kirkpeptide,.Ltd
| Product Name | 5-FITC | |
|---|---|---|
| Synonyms | FITC Fluorescein isothiocyanate isomer I |
|
| Cas No. | 3326-32-7 | |
| Purity | 95.00%+ | |
| Properties State | yellow powder | |
| Molecular Weight | 389.38 | |
| Molecular Formula | C21H11NO5S | |
| Density | 1.542 g/mL | |
| Description | Fluorescein isothiocyanate (FITC) is a yellow-orange molecule with an absorption maximum at 495nm. Upon excitation, it emits a yellow-green color with an emission maximum at 525nm. Fluorescein isothiocyanate is widely used to attach a fluorescent label to proteins via the amine group. The isothiocyanate group reacts with nucleophiles such as amino or thiol terminals. It has been used for the labeling of proteins including antibodies and lectins. Reagent grade, for research purpose. | |
| IUPAC Name | 3′,6′-dihydroxy-6-isothiocyanatospiro[2-benzofuran-3,9′-xanthene]-1-one | |
| InChI | InChI=1S/C21H11NO5S/c23-12-2-5-16-18(8-12)26-19-9-13(24)3-6-17(19)21(16)15-4-1-11(22-10-28)7-14(15)20(25)27-21/h1-9,23-24H | |
| InChIKey | MHMNJMPURVTYEJ-UHFFFAOYSA-N | |
| Canonical SMILES | C1=CC2=C(C=C1N=C=S)C(=O)OC23C4=C(C=C(C=C4)O)OC5=C3C=CC(=C5)O | |
| CBNumber | CB0173162 | |
| Ex/Em (nm) | 488/525 | |
| Storage Condition | store at under-20 ℃ | |
Product description
FITC can bind to various antibody proteins, and the bound antibody does not lose the specificity of binding to a certain antigen, and has strong yellow-green fluorescence in alkaline solution. The corresponding antigen can be qualitatively, localized or quantitatively detected by observation under a fluorescence microscope or flow cytometry analysis. Used in medicine, agriculture and animal husbandry, it can quickly diagnose diseases caused by bacteria, viruses and parasites.
FITC labeled antibody process The protein to be crosslinked (concentration ≥1mg/mL) was dialyzed three times against the crosslinking reaction solution at 4°C to pH=9.0. The preparation method of the crosslinking reaction solution: 7.56g NaHCO3, 1.06g Na2CO3, 7.36g NaCl, add water to make the volume to 1 L. Dissolve FITC in DMSO at a concentration of 1 mg/mL. FITC used for each cross-linking should be freshly prepared and protected from light. Slowly add FITC to the antibody solution at the ratio of P:F (Protein: FITC) = 1mg: 150μg, and gently shake to mix it with the antibody while adding, and react at 4°C in the dark for 8 hours. Add 5mol/L NH4Cl to a final concentration of 50mmol/L, and terminate the reaction at 4°C for 2h. Dialyze the cross-linked substance in PBS for more than four times until the dialysate is clear. Identification of cross-linked products Protein concentration (mg/mL) = [A280– 0.31×A495] / 1.4 F/P ratio: 3.1×A495/ [A280 – 0.31×A495], the value should be between 2.5 and 6.5. FITC cross-linked protein should be placed in a pH 7.4 phosphate buffer, added with 0.1% NaN3, 1% BSA, and stored at 4°C in the dark.
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. It is typically available as a mixture of isomers, fluorescein 5-isothiocyanate (5-FITC) and fluorescein 6-isothiocyanate (6-FITC). FITC is reactive towards nucleophiles including amine and sulfhydryl groups on proteins. It was synthesized by Robert Seiwald and Joseph Burckhalter in 1958.
A succinimidyl-ester functional group attached to the fluorescein core, creating “NHS-fluorescein”, forms another common amine reactive derivative that has much greater specificity toward primary amines in the presence of other nucleophiles.
FITC has excitation and emission spectrum peak wavelengths of approximately 495 nm and 519 nm, giving it a green color. Like most fluorochromes, it is prone to photobleaching. Due to the problem of photobleaching, derivatives of fluorescein such as Alexa 488 and DyLight 488 have been tailored for various chemical and biological applications where greater photostability, higher fluorescence intensity, or different attachment groups are needed. In addition, some experiments make use of FITC’s propensity for photobleaching in order to measure proteins’ lateral mobility in membranes, through the technique of fluorescence recovery after photobleaching.