Specific peptides as alternative to antibody ligands for biomagnetic separation of Clostridium tyrobutyricum spores |
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Authors: | Lavilla Maria Moros Maria Puertas Sara Grazú Valeria Pérez María Dolores Calvo Miguel de la Fuente Jesus M Sánchez Lourdes |
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Institution: | (1) Tecnolog?a de los Alimentos, Facultad de Veterinaria, Universidad de Zaragoza, C/Miguel Servet, 177, 50013 Zaragoza, Spain;(2) Instituto de Nanociencia de Arag?n (INA), Universidad de Zaragoza, C/Mariano Esquillor s/n, 50018 Zaragoza, Spain; |
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Abstract: | Nowadays, the reference method for the detection of Clostridium tyrobutyricum in milk is the most-probable-number method, a very time-consuming and non-specific method. In this work, the suitability
of the use of superparamagnetic beads coated with specific antibodies and peptides for bioseparation and concentration of
spores of C. tyrobutyricum has been assessed. Peptide or antibody functionalized nanoparticles were able to specifically bind C. tyrobutyricum spores and concentrate them up to detectable levels. Moreover, several factors, such as particle size (200 nm and 1 μm),
particle derivatization (aminated and carboxylated beads), coating method, and type of ligand have been studied in order to
establish the most appropriate conditions for spore separation. Results show that concentration of spore is favored by a smaller
bead size due to the wider surface of interaction in relation to particle volume. Antibody orientation, related to the binding
method, is also critical in spore recovery. However, specific peptides seem to be a better ligand than antibodies, not only
due to the higher recovery ratio of spores obtained but also due to the prolonged stability over time, allowing an optimal
recovery of spores up to 3 weeks after bead coating. These results demonstrate that specific peptides bound to magnetic nanoparticles
can be used instead of traditional antibodies to specifically bind C. tyrobutyricum spores being a potential basis for a rapid method to detect this bacterial target. |
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