title: Specific peptides as alternative to antibody ligands for biomagnetic separation of Clostridium tyrobutyricum spores.
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.
authors.affiliation: Tecnología de los Alimentos, Facultad de Veterinaria, Universidad de Zaragoza, C/Miguel Servet, 177, 50013 Zaragoza, Spain.
authors.forename: Maria
authors.initials: M
authors.lastname: Lavilla
authors.forename: Maria
authors.initials: M
authors.lastname: Moros
authors.forename: Sara
authors.initials: S
authors.lastname: Puertas
authors.forename: Valeria
authors.initials: V
authors.lastname: Grazú
authors.forename: María Dolores
authors.initials: MD
authors.lastname: Pérez
authors.forename: Miguel
authors.initials: M
authors.lastname: Calvo
authors.forename: Jesus M
authors.initials: JM
authors.lastname: de la Fuente
authors.forename: Lourdes
authors.initials: L
authors.lastname: Sánchez
mesh_terms: D000818:Animals; D000906:Antibodies / Q000737:chemistry; D002417:Cattle; D048014:Clostridium tyrobutyricum / Q000737:chemistry / Q000302:isolation & purification*; D005506:Food Contamination / Q000032:analysis; D018189:Immunomagnetic Separation / Q000295:instrumentation / Q000379:methods*; D008024:Ligands; D008892:Milk / Q000382:microbiology; D010455:Peptides / Q000737:chemistry*; D013171:Spores, Bacterial / Q000737:chemistry / Q000302:isolation & purification*
chemical_list: D000906:Antibodies; D008024:Ligands; D010455:Peptides