DEVELOPMENT OF OLIGOBODIES (APTAMERS) AGAINST THE BOVINE PRP (PRION)

VALDIVIESO AG; TAMINELLI GL; TEIBER ML; DANKERT MA; SANTA COLOMA TA

Rosario, Argentina.

Congreso; Sociedad Argentina de Investigación en Bioquímica y Biología Molecular (SAIB), XLII Reunión Anual.; 2006

Sociedad Argentina de Investigación en Bioquímica y Biología Molecular

Oligobodies (Obs) are a new generation of aptamers, which possess higher specificity than their predecessors. The objective of this work was to develop polyclonal Obs against synthetic peptides corresponding to three different regions of bovine PrP (PrPc, non-infective), and to test them in their ability to recognize cellular PrP (PrPc) and recombinant PrP (rPrP), as a first approach in the development of Obs against the infective prion (PrPsc). Three synthetic peptides were designed as temporary targets for PrP. Selections were performed incubating membranes saturated with each target peptide and with the single strand library. The bound and eluted oligonucleotides were then amplified by PCR and the selection repeated until a clear binding to the target was obtained. Then, the peptide targets were changed for the entire PrP protein present in a complex mixture. This procedure adds a pressure of selection towards high specificity, instead of high affinity, which distinguishes our method from SELEX, the method to produce aptamers. The polyclonal Obs obtained were able to recognize rPrP (50 ng) in Western blot. Even thought the specificity was not optimal yet, they migth be used in a blood assay if a step previous treatment with proteinasa K is performed. We will now introduce random mutations on the monoclonal Obs to attempt to improve their specificity. Acknowledgments: CONICET and UBA.