Proceedings book download - 5Z.com

Table 1. Sequences and analytical data of synthesized peptidesESI-MS; [M+H]PeptideSequence+ (m/z)calculated foundPenta ERN(Glc)GH-NH 2 773.35 773.64Peg-Penta NH 2 -Peg-ERN(Glc)GH -NH 2 1091.35 1091.42Hepta VERN(Glc)GHS-NH 2 959.45 959.76Peg-Hepta NH 2 -Peg-VERN(Glc)GHS-NH 2 1278.05 1278.32was coated in a polystyrene microplate and the shortened penta- and heptapeptides weretested in a competition assay to evaluate their affinity to specific antibodies. All thesynthesized shortened peptides were able to inhibit antibodies presenting similar IC 50 .Therefore, the autoantibody recognition in ELISA is not affected by Peg modifications.Moreover, competition assay was also performed by SPR instrumentation.Immobilization of CSF114(Glc) was performed on the sensor chip surface following theamine coupling strategy. This sensor chip was used to test the inhibition of specificautoantibodies in Multiple Sclerosis patients’ sera with the synthesized shortened pentaandheptapeptide sequences. Results showed that the new modified peptides were able torecognize autoantibodies in patients’ sera by a SPR biosensor.The new shortened peptides containing Peg spacer were able to detect specific anti-CSF114(Glc) antibodies in both competition ELISA and SPR, then modifications have notmodified the epitope of recognition. Further experiments to facilitate shortened peptidesequences immobilization will be performed in ELISA. Moreover, SPR technology will beuseful for monitoring the peptide coating on the sensor surface. The possibility ofmonitoring immobilization of synthetic probes in real-time directly on the sensor chipsurface will enable to develop new sequences for antibody detection. Binding interactionswith autoantibodies in MS patients’ sera will be measured using all the synthetic shortenedglucosylated peptide sequences.AcknowledgmentsEnte Cassa Risparmio di Firenze, PRIN 2008, and ANR Chaire d’Excellence PepKit 2009-2013(France) are gratefully acknowledged.References1. (a) Lolli, F., et al. Proc. Natl. Acad. Sci. U.S.A. 102, 10273-10278 (2005); (b) Lolli, F., et al. J.Neuroimm. 167, 131-137 (2005); (c) Papini, A.M. Nat. Med. 11, 13 (2005); (d) Carotenuto, A., etal. J. Med. Chem. 49, 5072-5079 (2006); (e) Papini, A.M., Rovero, P., Chelli, M., Lolli, F. GrantedU.S.A. Patent & PCT Application WO 03/000733 A2; (f) Carotenuto. A., et al. J. Med. Chem. 51,5304-5309 (2008).2. Nuti, F., Mulinacci, B., Peroni, E., Alcaro, M.C., Paolini, I., Benedetti, F., Carotenuto, A., Ciolli, F.,Lolli, F., Chelli, M., Rovero, P., Papini, A.M. Adv. Exp. Med. Biol. 611, 431-432 (2009).499