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  4. FITC

FITC  (Synonyms: Fluorescein 5-isothiocyanate)

Cat. No.: HY-66019 Purity: 98.77%
COA Handling Instructions

FITC (Fluorescein Isothiocyanate), is one of the green fluorescein derivatives widely used in biology. FITC has the characteristics of high absorptivity, excellent fluorescence quantum yield and good water solubility. The isothiocyanate group of FITC can be combined with amino, sulfhydryl, imidazole, tyrosyl, carbonyl and other groups on the protein, so as to achieve protein labeling including antibodies and lectins. In addition to its use as a protein marker, FITC can also be used as a fluorescent protein tracer to rapidly identify pathogens by labeling antibodies, or for microsequencing of proteins and peptides (HPLC). The maximum excitation wavelength of FITC is 494 nm. Once excited, it fluoresces yellow-green at a maximum emission wavelength of 520 nm.

For research use only. We do not sell to patients.

FITC Chemical Structure

FITC Chemical Structure

CAS No. : 3326-32-7

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10 mL/1 mL in DMSO USD 55 In-stock
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Customer Review

Based on 32 publication(s) in Google Scholar

FITC Related Antibodies

Top Publications Citing Use of Products

    FITC purchased from MCE. Usage Cited in: Adv Sci (Weinh). 2022 Jan 20;e2104344.  [Abstract]

    For detection of localization of WBC100 in cells, FITC labeled WBC100 (WBC100‐FITC) is used. HEK293T cells are transfected with the indicated plasmids for 48 h. Cells are treated with 1 μM WBC100‐FITC or FITC for 24 h.

    FITC purchased from MCE. Usage Cited in: J Nanobiotechnology. 2022 Feb 2;20(1):61.  [Abstract]

    To exam the ability of anti-PD-L1 antibodies (aPD-L1) nanovesicles to binding to tumor cells, B16F10 cells are cultured for 24 h. FITC-labeled aPD-L1 nanovesicles or nanovesicles are incubated with the cancer cells for 3 h.

    FITC purchased from MCE. Usage Cited in: Nat Commun. 2021 Nov 5;12(1):6426.  [Abstract]

    Cinnamate acetate (CIA) is labeled with FITC. Distribution of CIA/FITC in root cells of wild-type, ala7 mutant and complemented Arabidopsis seedlings. The seedlings are treated with CIA/FITC (5 μg/mL) and FM4-64 (8 μM) for 1 h, 2 h, and 8 h.

    FITC purchased from MCE. Usage Cited in: ACS Nano. 2020 Nov 24;14(11):14907-14918.  [Abstract]

    FITC-labeled MnO2/BPD are added at a concentration of 50 μg/mL and incubated for another 4 h to allow the nanoparticles to enter the cells. By conjugating FITC to MnO2/BPD, FL could be monitored at 488 nm, and the vessels could be observed at 660 nm by Evans Blue staining.

    FITC purchased from MCE. Usage Cited in: Chem Eng J. 2020, 127870.

    FITC (1 mg/mL) solution is prepared with dimethyl sulfoxide. Then FITC solution is added dropwise into laccase (5 mg/mL) solution, and the mixed solution is then stirred at 4 °C for 4 h. Laccase is labelled by FITC and exhibited green emissions.

    • Biological Activity

    • Protocol

    • Purity & Documentation

    • References

    • Customer Review

    Description

    FITC (Fluorescein Isothiocyanate), is one of the green fluorescein derivatives widely used in biology. FITC has the characteristics of high absorptivity, excellent fluorescence quantum yield and good water solubility. The isothiocyanate group of FITC can be combined with amino, sulfhydryl, imidazole, tyrosyl, carbonyl and other groups on the protein, so as to achieve protein labeling including antibodies and lectins. In addition to its use as a protein marker, FITC can also be used as a fluorescent protein tracer to rapidly identify pathogens by labeling antibodies, or for microsequencing of proteins and peptides (HPLC). The maximum excitation wavelength of FITC is 494 nm. Once excited, it fluoresces yellow-green at a maximum emission wavelength of 520 nm.

    In Vitro

    Protocol

    1.Protein Preparetion
    1) In order to obtain the best labeling effect, please prepare the protein (antibody) concentration as 2 mg/mL.
    2) The pH value of protein solution shall be 8.5±0.5. If the pH is lower than 8.0, 1m sodium bicarbonate shall be used for adjustment.
    3) If the protein concentration is lower than 2mg/ml, the labeling efficiency will be greatly reduced. In order to obtain the best labeling efficiency, it is recommended that the final protein concentration range is 2-10 mg/mL.
    4) The protein must be in the buffer without primary amine (such as Tris or glycine) and ammonium ion, otherwise the labeling efficiency will be affected.
    2.Dye Preparation
    Add anhydrous DMSO into the vial of FITC to make a 10 mM stock solution. Mix well by pipetting or vortex.
    3.Calculation of dye dosage
    The amount of FITC required for reaction depends on the amount of protein to be labeled, and the optimal molar ratio of FITC to protein is about 10.
    Example: assuming the required marker protein is 1 mL 2 mg/mL IgG (MW=150,000), use 1 mL DMSO dissolve 1 mg FITC, the required FITC volume is 40 μL.
    4.Run conjugation reaction
    1) A good volume of freshly prepared 10 mg/mL FITC is slowly added to 0.5 mL protein sample. In solution, gently shake to mix, then centrifuge briefly to collect the sample at the bottom of the reaction tube. Don't mix well to prevent protein samples from denaturation and inactivation.
    2) The reaction tubules were placed in a dark place and incubated gently at room temperature for 60 minutes at intervals.For 10-15 minutes, gently reverse the reaction tubules several times to fully mix the two reactants and raise the bar efficiency.
    5.Purify the conjugation
    The following protocol is an example of dye-protein conjugate purification by using a Sephadex G-25 column.
    1) Prepare Sephadex G-25 column according to the manufacture instruction.
    2) Load the reaction mixture (From "Run conjugation reaction") to the top of the Sephadex G-25 column.
    3) Add PBS (pH 7.2-7.4) as soon as the sample runs just below the top resin surface.
    4) Add more PBS (pH 7.2-7.4) to the desired sample to complete the column purification.
    Combine the fractions that contain the desired dye-protein conjugate.

    Note
    1. FITC is sensitive to light and humidity. Immediately add FITC solution and discard the unused part.
    2. Low concentrations of sodium azide (≤3 mM or 0.02%) or thiomersal (≤0.02 mM or 0.01%) did not significantly interfere with protein labeling; However, 20-50% glycerol will reduce labeling efficiency.
    3. Avoid buffering with primary amines (e.g., Tris, glycine) or ammonium ions,It compete with labeled proteins.
    4. This product is only for scientific research by professionals, and shall not be used in clinical diagnosis or treatment, food or medicine.
    5. For your safety and health, please wear lab coat and disposable gloves.

    MCE has not independently confirmed the accuracy of these methods. They are for reference only.

    FITC Related Antibodies
    Molecular Weight

    389.38

    Appearance

    Solid

    Formula

    C21H11NO5S
    CAS No.
    Emission (Em)

    525
    Excitation (Ex)

    488
    SMILES

    O=C1OC2(C3=C(OC4=C2C=CC(O)=C4)C=C(O)C=C3)C5=C1C=C(N=C=S)C=C5
    Shipping

    Room temperature in continental US; may vary elsewhere.
    Storage

    -20°C, protect from light, stored under nitrogen

    *In solvent : -80°C, 6 months; -20°C, 1 month (protect from light, stored under nitrogen)

    Solvent & Solubility
    In Vitro: 

    DMSO : 50 mg/mL (128.41 mM; Need ultrasonic)

    H2O : < 0.1 mg/mL (insoluble)

    Preparing
    Stock Solutions
    Concentration Solvent Mass 1 mg 5 mg 10 mg
    1 mM 2.5682 mL 12.8409 mL 25.6819 mL
    5 mM 0.5136 mL 2.5682 mL 5.1364 mL
    10 mM 0.2568 mL 1.2841 mL 2.5682 mL
    *Please refer to the solubility information to select the appropriate solvent.
    In Vivo:
    • 1.

      Add each solvent one by one:  10% DMSO    40% PEG300    5% Tween-80    45% saline

      Solubility: ≥ 2.08 mg/mL (5.34 mM); Clear solution

    • 2.

      Add each solvent one by one:  10% DMSO    90% (20% SBE-β-CD in saline)

      Solubility: 2.08 mg/mL (5.34 mM); Suspended solution; Need ultrasonic

    *All of the co-solvents are available by MCE.
    Purity & Documentation

    Purity: 98.77%

    COA (256 KB) HNMR (280 KB) LCMS (270 KB)

    Handling Instructions (2659 KB)
    Dyeing Example
    References
    Cell Assay
    [1]

    The frequency of chondrocyte apoptosis is measured by flow cytometry with Annexin V-fluorescein isothiocyanate (FITC) and propidium iodide. A total of 1×104 treated chondrocytes are collected from each group, washed in cold PBS and incubated with Annexin V-FITC and PI at room temperature for 15 min in the dark on ice. These samples are then analyzed using a fluorescence-activated cell sorter. Cell Quest software is used to analyze the percentage of apoptosis. All tests are repeated in triplicate.

    MCE has not independently confirmed the accuracy of these methods. They are for reference only.
    References
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    FITC Related Classifications

    Help & FAQs
    • Do most proteins show cross-species activity?

      Species cross-reactivity must be investigated individually for each product. Many human cytokines will produce a nice response in mouse cell lines, and many mouse proteins will show activity on human cells. Other proteins may have a lower specific activity when used in the opposite species.

    • Molarity Calculator

    • Dilution Calculator

    The molarity calculator equation

    Mass (g) = Concentration (mol/L) × Volume (L) × Molecular Weight (g/mol)

    Mass   Concentration   Volume   Molecular Weight *
    = × ×

    The dilution calculator equation

    Concentration (start) × Volume (start) = Concentration (final) × Volume (final)

    This equation is commonly abbreviated as: C1V1 = C2V2

    Concentration (start) × Volume (start) = Concentration (final) × Volume (final)
    × = ×
    C1   V1   C2   V2

    Keywords:

    FITC3326-32-7Fluorescein 5-isothiocyanateFluorescent DyeInhibitorinhibitorinhibit

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