Cellulose based bio-interfaces for immunodiagnostic applications

Aalto University, P.O. Box 11000, FI-00076 Aalto www.aalto.fi Author Hannes Orelma Name of the doctoral dissertation Cellulose based bio-interfaces for immunodiagnostic applications Publisher School of Chemical Technology Unit Department of Forest Products Technology Series Aalto University publication series DOCTORAL DISSERTATIONS 138/2012 Field of research Forest Products Chemistry Manuscript submitted 5 June 2012 Date of the defence 9 November 2012 Permission to publish granted (date) 25 September 2012 Language English Monograph Article dissertation (summary + original articles) Abstract In this work, the interactions between various proteins and modified cellulose surfaces were investigated. The work focused on the development of immobilization methods for the covalent attachment of specific immunological antibodies (proteins) onto cellulose substrate. The immobilization methods were explored using cellulose model surfaces and surface sensitive techniques, such as quartz crystal microbalance with dissipation monitoring (QCM-D) and surface plasmon resonance (SPR). The highest adsorption of globular proteins on unmodified cellulose surfaces occurred at their respective isoelectric points, suggesting a non-electrostatic adsorption mechanism. An increased surface charge at the cellulose substrate was found to enhance the adsorption of all the proteins investigated. This indicated the presence of attractive electrostatic interactions and the adsorption was found to be mainly irreversible. In addition, the effect of oligosaccharide regions of proteins on their adsorption on cellulose was examined with one glycoprotein, avidin. The adsorption of avidin on cellulose was driven by a combination of electrostatic and non-electrostatic forces, and the adsorption was mainly irreversible. Moreover, the oligosaccharide regions of avidin decreased its adsorption strength to cellulose. In this work, several strategies for covalent immobilization of antibodies onto functionalized cellulose matrices were developed. The novel biointerfaces were capable of sensing antigens both selectively and quantitatively. The use of traditional conjugation chemistries typically leads to a random conformation of immobilized antibodies on the surfaces which in turn may decrease the ability of immobilized antibodies to bind antigens due to the sterical hindrances. Therefore, in this work, the antibodies were immobilized onto cellulose in more oriented manner using avidin-biotin linkage. This approach resulted in over two-fold higher antigen response when compared to those of the traditional conjugation chemistry. In the last part of this work, a biointerface was prepared on a water-resistant nanofibrillar cellulose (NFC) film. The NFC film was made amine reactive by using sequential TEMPO-mediated oxidation and EDC/NHS activation. Activated NFC-films were observed to bind antibodies covalently, and the antibodies could be deposited using standard inkjet printing techniques. The developed NFC-based biointerfaces are expected to open new venues for using cellulose in immunodiagnostic applications.In this work, the interactions between various proteins and modified cellulose surfaces were investigated. The work focused on the development of immobilization methods for the covalent attachment of specific immunological antibodies (proteins) onto cellulose substrate. The immobilization methods were explored using cellulose model surfaces and surface sensitive techniques, such as quartz crystal microbalance with dissipation monitoring (QCM-D) and surface plasmon resonance (SPR). The highest adsorption of globular proteins on unmodified cellulose surfaces occurred at their respective isoelectric points, suggesting a non-electrostatic adsorption mechanism. An increased surface charge at the cellulose substrate was found to enhance the adsorption of all the proteins investigated. This indicated the presence of attractive electrostatic interactions and the adsorption was found to be mainly irreversible. In addition, the effect of oligosaccharide regions of proteins on their adsorption on cellulose was examined with one glycoprotein, avidin. The adsorption of avidin on cellulose was driven by a combination of electrostatic and non-electrostatic forces, and the adsorption was mainly irreversible. Moreover, the oligosaccharide regions of avidin decreased its adsorption strength to cellulose. In this work, several strategies for covalent immobilization of antibodies onto functionalized cellulose matrices were developed. The novel biointerfaces were capable of sensing antigens both selectively and quantitatively. The use of traditional conjugation chemistries typically leads to a random conformation of immobilized antibodies on the surfaces which in turn may decrease the ability of immobilized antibodies to bind antigens due to the sterical hindrances. Therefore, in this work, the antibodies were immobilized onto cellulose in more oriented manner using avidin-biotin linkage. This approach resulted in over two-fold higher antigen response when compared to those of the traditional conjugation chemistry. In the last part of this work, a biointerface was prepared on a water-resistant nanofibrillar cellulose (NFC) film. The NFC film was made amine reactive by using sequential TEMPO-mediated oxidation and EDC/NHS activation. Activated NFC-films were observed to bind antibodies covalently, and the antibodies could be deposited using standard inkjet printing techniques. The developed NFC-based biointerfaces are expected to open new venues for using cellulose in immunodiagnostic applications.