Heparin (Hep) is a sulfated glycosaminoglycan composed of repeating disaccharide units of D-iduronic acid (IdoUA) or D-glucuronic acid (GlcUA) and N-acetyl-D-glucosamine or N-sulfo-glucosamine. Hep is abundant in mast cells of intestine or lung exists as unbranched polysaccharide chains. This product is prepared by the fluorescent labeling of Hep derived from porcine intestine according to the method of Ogamo et al.1). Fluoresceinamine molecules are chemically attached to carboxyl groups of the GlcUA or IdoUA of Hep. This solution is dissolved in PBS (-) and sterilized by filtration. The endotoxin content is in accordance with the product specifications. The excitation wavelength is 490~500 nm and the emission wavelength is 515~525 nm. The enclosed Certification of Analysis lists actual values for product specifications.
Fluoresce inamine labeled Sodium Hyaluronate flyer download [PDF]
Typical disaccharide formula of Sodium Heparin
Analysis of binding activities with bFGF
Basic fibroblast growth factor (bFGF, 10μg/mL in PBS, 100 μL/well) was added into 96well plate (Nunc, 437111) and incubated at 37°C for 1h. The plate was washed with PBS, and then fluorescein-labeled GAG solution with various concentrations in PBS was added to each well (100μL/well ) and incubated at 37°C for 1h. After washing the plate, 0.1N NaOH was added (100μL/well), and the fluorescence intensity was measured using a fluoro-plate reader with an excitation wavelength of 485 nm and an emission wavelength of 535 nm. It was observed that the fluorescence intensity was increased with the concentration of FACS-E1, FAHS-P1 FAHep-N1 and FAHS-P1 whereas the intensity was little increased in the case of FAHA-M1. The result suggests an interaction between sulfated GAG and bFGF.
Analysis of binding activities with BMP-4 by fluorescence correlation spectroscopy (FCS)
Fluorescence correlation spectroscopy (FCS) is an analytical method of molecular- association or dissociation with one molecular level. The interaction of molecules can be detected as a prolongation of average diffusion time (DT) derived from a fluorescent molecule. The figure shows the effect of recombinant human bone morphogenetic protein-4 (rhBMP-4) on the DT of various fluoresceinamine-labeled GAGs. The result indicates relatively strong interactions between rhBMP-4 and FACS-E1, FACS-P1 or FAHep-N1. (J Cell Physiol, 2008, 217(3):769-777)