Gravimetric antigen detection utilizing antibody-modified lipid bilayers |
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Authors: | Larsson Charlotte Bramfeldt Hanna Wingren Christer Borrebaeck Carl Höök Fredrik |
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Institution: | a Department of Applied Physics, Chalmers University of Technology and Göteborg University, 41296 Göteborg, Sweden b Department of Immunotechnology, Lund University, 22007 Lund, Sweden |
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Abstract: | Lipid bilayers containing 5% nitrilotriacetic acid (NTA) lipids supported on SiO2 have been used as a template for immobilization of oligohistidine-tagged single-chained antibody fragments (scFvs) directed against cholera toxin. It was demonstrated that histidine-tagged scFvs could be equally efficiently coupled to an NTA-Ni2+-containing lipid bilayer from a purified sample as from an expression supernatant, thereby providing a coupling method that eliminates time-consuming protein prepurification steps. Irrespective of whether the coupling was made from the unpurified or purified antibody preparation, the template proved to be efficient for antigen (cholera toxin) detection, verified using quartz crystal microbalance with dissipation monitoring. In addition, via a secondary amplification step using lipid vesicles containing GM1 (the natural membrane receptor for cholera toxin), the detection limit of cholera toxin was less than 750 pM. To further strengthen the coupling of scFvs to the lipid bilayer, scFvs containing two histidine tags, instead of just one tag, were also evaluated. The increased coupling strength provided via the bivalent anchoring significantly reduced scFv displacement in complex solutions containing large amounts of histidine-containing proteins, verified via cholera toxin detection in serum. |
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Keywords: | QCM NTA Antibody scFv Cholera toxin GM1 Supported lipid bilayer |
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