Specific and selective probes for Pseudomonas aeruginosa from phage-displayed random peptide libraries.
The design of novel biosensors for the detection of biological threats, such as Pseudomonas aeruginosa, requires probes that specifically bind biological agents and insure their immediate and efficient recognition. Advanced bio-selective sensors may meet the requests for isolation, concentration of the agents and their real-time detection. There is a need for robust and inexpensive affinity probes alternative to antibodies. These probes may be recruited from random peptide libraries displayed on filamentous phage. In this study, we identified from two phage-displayed random peptide libraries phage clones displaying peptides capable of specific and strong binding to P. aeruginosa cell surface. The ability of the phage clones to interact specifically with P. aeruginosa was demonstrated by using enzyme-linked immunosorbent assay (ELISA). We assessed selectivity of phage-bacteria-binding by comparing the binding ability of the selected clones to the selector bacterium and a panel of other bacterial species; we also demonstrated by dot spot and immunoblotting that the most reactive and selective phage peptide bound with high avidity the bacterial cell surface. In addition, as proof-of-concept, we tested the possibility to immobilize the affinity-selected phage to a putative biosensor surface. The quality of phage deposition was monitored by ELISA, and phage-bacterial-binding was confirmed by high-power optical phase contrast microscopy. Overall, the results of this work validate the concept of affinity-selected recombinant filamentous phages as probes for detecting and monitoring bacterial agents under any conditions that warrant their recognition, including clinical-based diagnostics and possibly biological warfare applications.
Carnazza
Santina
S
Department of Microbiological, Genetic and Molecular Sciences, University of Messina, Sal Sperone 31, Vill S Agata, Messina, Italy. santina.carnazza@unime.it
Foti
Carmela
C
Gioffrè
Giuseppina
G
Felici
Franco
F
Guglielmino
Salvatore
S
D015374:Biosensing Techniques / Q000295:instrumentation* / Q000379:methods; D004797:Enzyme-Linked Immunosorbent Assay / Q000295:instrumentation* / Q000379:methods; D004867:Equipment Design; D019544:Equipment Failure Analysis; D015336:Molecular Probe Techniques / Q000295:instrumentation*; D019151:Peptide Library; D011550:Pseudomonas aeruginosa / Q000302:isolation & purification* / Q000378:metabolism; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
D019151:Peptide Library