Orientation and conformation of anti-CD34 antibody immobilised on untreated and plasma treated polycarbonate.
The conformation and orientation of proteins immobilised on synthetic materials determine their ability to bind their antigens and thereby the sensitivity of the microarrays and biosensors employing them. Plasma immersion ion implantation (PIII) of polymers significantly increases both their wettability and protein binding capacity. This paper addresses the hypothesis that a PIII treated polymer surface modifies the native protein conformation less significantly than a more hydrophobic untreated surface and that the differences in surface properties also affect the protein orientation. To prove this, the orientation and conformation of rat anti-mouse CD34 antibody immobilized on untreated and PIII treated polycarbonate (PC) were investigated using ToF-SIMS and FTIR-ATR spectroscopy. Analysis of the primary structure of anti-CD34 antibody and principal component analysis of ToF-SIMS data were applied to detect the difference in the orientation of the antibody attached to untreated and PIII treated PC. The difference in the antibody conformation was analysed using deconvolution of the Amide I peak (in FTIR-ATR spectra) and curve-fitting. It was found that compared to the PIII treated sample, the antibody immobilized on the untreated PC sample has a secondary structure with a lower fraction of β-sheets and a higher fraction of α-helices and disordered fragments. Also, it was found that anti-CD34 antibody has a higher tendency to occur in the inactive 'tail-up' orientation when immobilized on an untreated PC surface than on a PIII treated surface. These findings confirm the above hypothesis.
The School of Physics, University of Sydney, NSW 2006, Australia. Electronic address: elenak@physics.usyd.edu.au.
E
E
Kosobrodova
Department of Physics and Centre for Materials and Surface Science, La Trobe University, Melbourne, VIC 3086, Australia.
R T
RT
Jones
The School of Physics, University of Sydney, NSW 2006, Australia.
A
A
Kondyurin
Faculty of Pharmacy, University of Sydney, NSW 2006, Australia.
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W
Chrzanowski
Department of Physics and Centre for Materials and Surface Science, La Trobe University, Melbourne, VIC 3086, Australia.
P J
PJ
Pigram
The School of Physics, University of Sydney, NSW 2006, Australia.
D R
DR
McKenzie
The School of Physics, University of Sydney, NSW 2006, Australia.
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MM
Bilek
D000327:Adsorption; D000911:Antibodies, Monoclonal / Q000737:chemistry* / Q000276:immunology*; D018952:Antigens, CD34 / Q000276:immunology*; D020099:Coated Materials, Biocompatible / Q000138:chemical synthesis; D008422:Materials Testing; D058626:Plasma Gases / Q000737:chemistry*; D011076:Polycarboxylate Cement / Q000737:chemistry*; D011487:Protein Conformation; D013499:Surface Properties
D000911:Antibodies, Monoclonal; D018952:Antigens, CD34; D020099:Coated Materials, Biocompatible; D058626:Plasma Gases; D011076:Polycarboxylate Cement; C005506:polycarbonate
AFM; Antibody orientation; Conformation; Plasma ion implantation; ToF-SIMS