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

MW 4,000	Approx. 14 Angstroms
MW 10,000	Approx. 23 Angstroms
MW 20,000	Approx. 33 Angstroms
MW 40,000	Approx. 45 Angstroms
MW 70,000	Approx. 60 Angstroms
MW 150,000	Approx. 85 Angstroms

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

References

1. Rankin JC, Jeanes A. 1954. Evaluation of the Periodate Oxidation Method for Structural Analysis of Dextrans. J. Am. Chem. Soc.. 76(17):4435-4441. https://doi.org/10.1021/ja01646a046

2. Dimler RJ, Wolff IA, Sloan JW, Rist CE. 1955. Interpretation of Periodate Oxidation Data on Degraded Dextran. J. Am. Chem. Soc.. 77(24):6568-6573. https://doi.org/10.1021/ja01629a044

3. Van Cleve JW, Schaefer WC, Rist CE. 1956. The Structure of NRRL B-512 Dextran. Methylation Studies2. J. Am. Chem. Soc.. 78(17):4435-4438. https://doi.org/10.1021/ja01598a064

4. Lindberg B, Svensson S, Sjövall J, Zaidi NA. 1968. Structural Studies on Dextran from Leuconostoc mesenteroides NRRL B-512.. Acta Chem. Scand.. 221907-1912. https://doi.org/10.3891/acta.chem.scand.22-1907

5. Larm O, Lindberg B, Svensson S. 1971. Studies on the length of the side chains of the dextran elaborated by Leuconostoc mesenteroides NRRL B-512. Carbohydrate Research. 20(1):39-48. https://doi.org/10.1016/s0008-6215(00)84947-2

6. Bovey FA. 1959. Enzymatic polymerization. I. Molecular weight and branching during the formation of dextran. J. Polym. Sci.. 35(128):167-182. https://doi.org/10.1002/pol.1959.1203512813

7. Senti FR, Hellman NN, Ludwig NH, Babcock GE, Tobin R, Glass CA, Lamberts BL. 1955. Viscosity, sedimentation, and light-scattering properties of fraction of an acid-hydrolyzed dextran. J. Polym. Sci.. 17(86):527-546. https://doi.org/10.1002/pol.1955.120178605

8. Arond LH, Frank HP. 1954. Molecular Weight Distribution and Molecular Size of a Native Dextran. J. Phys. Chem.. 58(11):953-957. https://doi.org/10.1021/j150521a006

9. Elias VH. 1959. Ultrazentrifugen- und Diffusionsmessungen an nicht-Newtonschen Lösungen nativer Dextrane. Über extrem große Makromoleküle. IV. Makromol. Chem.. 33(1):166-180. https://doi.org/10.1002/macp.1959.020330112

10. Antonini E, Bellelli L, Bruzzesi MR, Caputo A, Chiancone E, Rossi-Fanelli A. 1964. Studies on dextran and dextran derivatives. I. Properties of native dextran in different solvents. Biopolymers. 2(1):27-34. https://doi.org/10.1002/bip.1964.360020105

11. Supplier's data..

12. Granath KA. 1958. Solution properties of branched dextrans. Journal of Colloid Science. 13(4):308-328. https://doi.org/10.1016/0095-8522(58)90041-2

13. Basedow AM, Ebert KH. 1979. Production, characterization, and solution properties of dextran fractions of narrow molecular weight distributions. J. polym. sci., C Polym. symp.. 66(1):101-115. https://doi.org/10.1002/polc.5070660113

14. Allen, PW. 1959 . Techiques of Polymer Characterization, . p. 131 . Butterworths Scientific Publications .

15. Granath KA, Flodin P. 1961. Makromol. Chem.. 48(1):160-171. https://doi.org/10.1002/macp.1961.020480116

16. Isbell H, Snyder C, Holt N, Dryden M. 1953. Determination of molecular weights of dextrans by means of alkaline copper reagents. J. RES. NATL. BUR. STAN.. 50(2):81. https://doi.org/10.6028/jres.050.014

17. Jermyn M. 1975. Increasing the sensitivity of the anthrone method for carbohydrate. Analytical Biochemistry. 68(1):332-335. https://doi.org/10.1016/0003-2697(75)90713-7

18. Ingelman, B, Halling, M. 1949 . Ark. Kemi . 1 61.

19. de Belder A, Granath K. 1973. Preparation and properties of fluorescein-labelled dextrans. Carbohydrate Research. 30(2):375-378. https://doi.org/10.1016/s0008-6215(00)81824-8

20. Arfors, K, Hint, H. 1971 . Studies of Microcirculation using Fluorescent Dextran Microvascular Research . 3(4):440.

21. Arfors, K, Rutili, G. 1972 . Microvasc. Res . 4 466.

22. Arfors, K, Rutili, G. 1972 . The usefulness of fluorescein labelled dextran in the study of macromolecular passage across the capillary and blood-lymph barrier Abstracts. . VII Conference on Microcirculation Aberdeen p. p. 3.

23. Baron-Epel O, Gharyal PK, Schindler M. 1988. Pectins as mediators of wall porosity in soybean cells. Planta. 175(3):389-395. https://doi.org/10.1007/bf00396345

24. Rutili, G, Arfors, KE. 1973 . Abstracts. Nordic Microcirculation Group Meeting Ustaoset.

25. Rutili, G, Arfors, K. 1972 . Abstracts. VII Conference on Microcirculation Aberdeen . p. p. 101.