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ABSTRACT

Immobilization of engineered antibodies on a screen printed surface

F.Dalla Riva*, M.De Francesco, L.Della Seta,
O.Bitti, A Masci, M.E. Villani, R.Franconi, R.Pilloton

Enea C.R.Casaccia, Via Anguillarse 301 - I00060 Rome - Italy
*University of Pavia, Chemistry Department, Via Taramelli 12 - I27100 Pavia – Italy

Recently, biosensor research was not only oriented to improve analytical performances but also focused on mass production, lower manufacturing costs and reproducibility. Screen-printing was chosen as one of the most promising technology, allowing biosensors to be largely on the market especially with disposable devices. Additionally, the development of new biosensors based on genetically engineered molecules may help to fulfill such requirements providing new reagents at high yields and low cost.
In this perspective, the availability of repertoires of recombinant antibodies displayed on phage may allow the quick isolation of new binding molecules with high specificity to virtually any analyte.

As a first step, several immobilization techniques were tried in order to couple a commercially available polyclonal antibody against atrazine on a PVC-based amperometric device with graphite (doped with ferrocene) and Ag/Pd inks as electrode materials. A direct ELISA format, with an HRP labeled antigen, was used to check both immobilization and electrochemical detection.
Subsequently, a single-chain antibody fragment (scFv), against the plant virus cucumber mosaic (CMV), was isolated from a synthetic phage display library. Oriented immobilization on thick film graphite layers was obtained exploiting the ability of the six histidine tag, at the C-terminus of the scFv, to give a stable complex with nickel ions when added to a suitable chelating agent.
Coupling the high sensitivity and specificity of scFvs molecules as sensing elements to an electrochemical transduction system, represents an interesting breakthrough in the development of new generic biosensors.