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Fluorescein-5-isothiocyanate (FITC)

Synonyms: Fluorescein isothiocyanate isomer I, 5-FITC

Fluorescein-5-isothiocyanate (FITC, Fluorescein isothiocyanate isomer I, 5-FITC) is a fluorescent probe capable of being conjugated to tissue and proteins.

Fluorescein-5-isothiocyanate (FITC) Chemical Structure

Fluorescein-5-isothiocyanate (FITC) Chemical Structure

CAS: 3326-32-7

Selleck's Fluorescein-5-isothiocyanate (FITC) has been cited by 1 publication

Purity & Quality Control

Batch: S692801 DMSO] 78 mg/mL] false] Ethanol] 10 mg/mL] false] Water] Insoluble] false Purity: 98.88%
98.88

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Biological Activity

Description Fluorescein-5-isothiocyanate (FITC, Fluorescein isothiocyanate isomer I, 5-FITC) is a fluorescent probe capable of being conjugated to tissue and proteins.
In vitro
In vitro

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.
NCT Number Recruitment Conditions Sponsor/Collaborators Start Date Phases
NCT05312411 Recruiting Osteosarcoma Seattle Children''s Hospital|Umoja BioPharma Inc. May 20 2022 Phase 1
NCT02743468 Completed Polymorphism|Lung Inflammation National Institute of Environmental Health Sciences (NIEHS)|National Institutes of Health Clinical Center (CC) January 8 2019 --
NCT02878083 Terminated ULCERATIVE COLITIS Nantes University Hospital|Takeda|Mauna Kea Technologies|Institut National de la Santé Et de la Recherche Médicale France January 11 2017 Not Applicable

Chemical lnformation & Solubility

Molecular Weight 389.38 Formula

C21H11NO5S
CAS No. 3326-32-7 SDF --
Smiles OC1=CC2=C(C=C1)C3(OC(=O)C4=C3C=CC(=C4)N=C=S)C5=C(O2)C=C(O)C=C5
Storage (From the date of receipt) 3 years -20°C(in the dark) powder

In vitro
Batch:

DMSO : 78 mg/mL ( (200.31 mM)； Moisture-absorbing DMSO reduces solubility. Please use fresh DMSO.)

Ethanol : 10 mg/mL

Water : Insoluble


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In vivo
Batch:

Add solvents to the product individually and in order.


In vivo Formulation Calculator

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In vivo Formulation Calculator (Clear solution)

Step 1: Enter information below (Recommended: An additional animal making an allowance for loss during the experiment)

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Step 2: Enter the in vivo formulation (This is only the calculator, not formulation. Please contact us first if there is no in vivo formulation at the solubility Section.)

% DMSO % % Tween 80 % ddH2O
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Working concentration: mg/ml;

Method for preparing DMSO master liquid: mg drug pre-dissolved in μL DMSO ( Master liquid concentration mg/mL, Please contact us first if the concentration exceeds the DMSO solubility of the batch of drug. )

Method for preparing in vivo formulation: Take μL DMSO master liquid, next addμL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL ddH2O, mix and clarify.

Method for preparing in vivo formulation: Take μL DMSO master liquid, next add μL Corn oil, mix and clarify.

Note: 1. Please make sure the liquid is clear before adding the next solvent.
2. Be sure to add the solvent(s) in order. You must ensure that the solution obtained, in the previous addition, is a clear solution before proceeding to add the next solvent. Physical methods such
as vortex, ultrasound or hot water bath can be used to aid dissolving.

Tech Support

Answers to questions you may have can be found in the inhibitor handling instructions. Topics include how to prepare stock solutions, how to store inhibitors, and issues that need special attention for cell-based assays and animal experiments.

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