Immobilization of high-affinity heparin oligosaccharides to radiofrequency plasma-modified polyethylene.
Oligosaccharides of heparin with high affinity for antithrombin III (ATIII) have been immobilized onto surface-modified NHLBI Primary Reference low density polyethylene (PE). PE was modified by radiofrequency plasma polymerized (< 150 nm thick) films derived from N-vinyl-2-pyrrolidone (PPNVP) or allyl alcohol (PPAA), and coupled by chemical derivatization to either 3-aminopropyltriethoxysilane or amino-terminated poly(ethylene oxide). High affinity heparin oligosaccharides (HA-heparin, anti-factor Xa activity of 592 +/- 120 IU/mg) prepared by partial deaminative cleavage of commercial crude heparin and fractionated by agarose-ATIII affinity chromatography, were immobilized to surface-modified PE by reductive amination. The anticoagulant activity, as determined by a chromogenic assay for the inhibition of factor Xa, was estimated to be 30-70 mIU/cm2, with binding estimated to be 56-119 ng/cm2. The highest activity was obtained for the HA-heparin immobilized to PE modified by PPNVP with a PEO spacer. Visual confirmation of ATIII binding to immobilized HA-heparin was demonstrated by a gold-labeled double antibody method with imaging by SEM.
Yuan
S
S
Department of Biomedical Engineering, Case Western Reserve University, Cleveland, Ohio 44106.
Szakalas-Gratzl
G
G
Ziats
N P
NP
Jacobsen
D W
DW
Kottke-Marchant
K
K
Marchant
R E
RE
D000433:1-Propanol / Q000737:chemistry; D000818:Animals; D000925:Anticoagulants / Q000494:pharmacology; D000990:Antithrombin III / Q000737:chemistry; D002417:Cattle; D002846:Chromatography, Affinity; D065427:Factor Xa Inhibitors; D006493:Heparin / Q000737:chemistry* / Q000494:pharmacology; D007150:Immunohistochemistry; D008855:Microscopy, Electron, Scanning; D008970:Molecular Weight; D009844:Oligosaccharides / Q000737:chemistry* / Q000494:pharmacology; D011095:Polyethylenes / Q000737:chemistry* / Q000528:radiation effects; D020005:Propanols; D011760:Pyrrolidinones / Q000737:chemistry; D011846:Radio Waves; D013055:Spectrophotometry, Infrared; D013462:Sulfur Radioisotopes
D000925:Anticoagulants; D065427:Factor Xa Inhibitors; D009844:Oligosaccharides; D011095:Polyethylenes; D020005:Propanols; D011760:Pyrrolidinones; D013462:Sulfur Radioisotopes; C006463:allyl alcohol; C042670:N-vinyl-2-pyrrolidinone; D000990:Antithrombin III; D006493:Heparin; D000433:1-Propanol