Fluorescein Isothiocyanate-Dextran

SYNONYM: FITC-Dextran

Structure

Dextran is a polymer of anhydroglucose. It is composed of approximately 95% alpha-D-(166) linkages. The remaining (163) linkages account for the branching of dextran.^1,2,3 Conflicting data on the branch lengths implies that the average branch length is less than three glucose units.^4,5 However, other methods indicate branches of greater than 50 glucose units exist.^6,7 Native dextran has been found to have a molecular weight (MW) in the range of 9 million to 500 million.^8,9,10 Lower MW dextrans will exhibit slightly less branching⁴ and have a more narrow range of MW distribution.¹¹ Dextrans with MW greater than 10,000 behave as if they are highly branched. As the MW increases, dextran molecules attain greater symmetry.^7,12,13 Dextrans with MW of 2,000 to 10,000, exhibit the properties of an expandable coil.¹² At MW below 2,000, dextran is more rod-like.¹⁴ The MW of dextran is measured by one or more of the following methods: low angle laser light scattering,¹⁵ size exclusion chromatography,¹⁶ copper-complexation¹⁷ and anthrone reagent¹⁸ colorometric reducing-end sugar determination and viscosity.¹²

Physical Properties

The approximate Stokes' radii for FITC-dextrans are as follows;¹¹

FITC is conjugated randomly to hydroxyl groups of dextran at a frequency of 0.003 to 0.02 moles of FITC per mole of glucose.

Fluorescent Properties

The excitation maximum of FITC-dextran is 490 nm. The emission maximum is 520 nm.¹¹ The fluorescence increases with pH and is optimal at pH 8 and above.¹¹

Method of Preparation

Our dextrans are derived from Leuconostoc mesenteroides, strain B 512. Various MW are produced by limited hydrolysis and fractionation. Our supplier's exact methods are held proprietary. Fractionation can be accomplished by size exclusion chromatography¹⁶ or ethanol fractionation, where the largest MW dextrans precipitate first.¹⁹ The FITC conjugation with hydroxyl groups of dextran is carried out in DMSO.²⁰

Storage/Stability

Stored properly at 2-8 °C and protected from light FITC-dextran powders should be stable for a minimum of two to three years.

Solubility/Solution Stability

We typically test the solubility of FITC dextrans in water at concentrations at or above 25 mg/ml. Solutions should be protected from light. In vivo, FITC-dextran is stable for more than 24 hours.²¹

Applications

FITC-dextran is used extensively in microcirculation and cell permeability research utilizing microfluorimetry.^22,23 FITC-dextran has been used to study plant cell wall porosity²⁴ and capillary permeability.^25,26 Plasma proteins have been shown not to bind to FITC-dextran.²⁶