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.

Development of a specific fluorescent phage endolysin for in situ detection of Clostridium species associated with cheese spoilage

“…As an alternative to antibody ligands, Lavilla et al . () produced superparamagnetic beads coated with specific peptides to specifically bind C. tyrobutyricum spores for concentration of spores prior to detection.…”

Revisiting 30 years of biofunctionalization and surface chemistry of inorganic nanoparticles for nanomedicine

“…may be attached to the nanoparticle surface without prior modification (see Figure 8) (Pandey et al, 2007; Susumu et al, 2007; Rostro-Kohanloo et al, 2009; Conde et al, 2012a; Lavilla et al, 2012). For instance, using the EDC chemistry, Weissleder and coworkers created a library of MNPs decorated with different synthetic small molecules for the development of magnetofluorescent reporters (Weissleder et al, 2005).…”

Evaluation of methods for DNA extraction from Clostridium tyrobutyricum spores and its detection by qPCR

“…In that study, the detection limit was established in 3 spores/100 mL. The use of magnetic nanoparticles with specific ligands, such as antibodies or affine peptides, were also proposed by Lavilla et al (2012) as novel methods for the detection of C. tyrobutyricum spores. Although those methods are faster and highly specific, their introduction into the routine analysis is not easy due to the high cost of the materials and equipment required.…”