Gelatin-Immobilised Poly(hydroxyethyl methacrylate) Cryogel for Affinity Purification of Fibronectin

Abstract

In this work, fibronectin purification from human plasma with the gelatin-immobilised poly(hydroxyethyl methacrylate) (PHEMA) cryogel has been evaluated. The PHEMA cryogel was prepared by cryo-polymerisation which proceeds in an aqueous solution of monomer frozen inside a plastic syringe. The PHEMA cryogel contained interconnected macrochannels of 10-200 μm in diameter. Gelatin molecules were covalently immobilised onto the PHEMA cryogel via carbodiimide activation. The gelatin-immobilised PHEMA cryogel was used to purify fibronectin from human plasma. Fibronectin adsorption from human plasma on the PHEMA cryogel was 0.30 mg/ml, while much higher adsorption values, up to 38 mg/ml, was obtained with the gelatin-immobilised PHEMA cryogel. The fibronectin adsorption capacity of the gelatin-immobilised PHEMA cryogel did not change with an increase in the flow rate of plasma. Up to 92 % of the adsorbed fibronectin was eluted using 2 M urea containing 1 M NaCl as elution agent. The adsorption-elution cycle was repeated ten times using the same PHEMA cryogel. No remarkable decrease was detected in the adsorption capacity of the gelatin-immobilised PHEMA cryogel.

The usage of composite nanomaterials in biomedical engineering applications

“…These methacrylated based nanosized materials can be shaped in different forms such as cryogels (Avcibaşi et al, 2010;Perçin et al, 2013;Savina et al, 2007), membranes (Arica & Hasirci, 1993;Demir et al, 2018;Kermis et al, 2003) or beads (Akgöl et al, 2007;Jayakrishnan & Thanoo, 1990;Vianna-Soares et al, 2004). They can be integrated to nanosized materials to seperate and isolate biomolecules with using different affinity chromatography techniques such as metal chelate (Karakoc et al, 2009;Odabasi et al, 2003), dye (Mislovičová et al, 1995;Unsal et al, 2005;Yavuz et al, 2006), boronate (Kuru et al, 2014;Srivastava et al, 2012;Toprak et al, 2015), lectin (Bereli et al, 2005;M.-X.…”

Adsorption of cellulase on poly(2-hydroxyethyl methacrylate) cryogels containing phenylalanine

“…A schematic illustration of p(HEMA-MAPA) cryogel with cellulase enzyme is given in the Scheme. 17 SEM analysis revealed that scaffolds possess large and interconnected macropores generated by the process of cryogelation. The interconnected macropores allow cellulase enzyme molecules to flow easily from the cryogel.…”

“…As seen in the figures, cryogel has a sponge-like structure and is supermacroporous (10 to 200 µ m in diameter), providing a size that can result in less fouling of pores with bioparticles and easy cleaning of the cryogel. 17 SEM analysis revealed that scaffolds possess large and interconnected macropores generated by the process of cryogelation. The interconnected macropores allow cellulase enzyme molecules to flow easily from the cryogel.…”