Proof of principle for a sensitive, real time and label-free detection of poxviruses using optical fiber biosensors

Reliable, affordable, rapid and sensitive point-of-care testing is essential for preparedness towards pandemic viruses that could emerge from zoonoses, laboratory accidents or undercover spread of biological weapons. Although successfully eradicated, variola virus, the causative agent of smallpox, is a category A agent stored in laboratories in USA and Russia, and a possible threat to humanity. Outbreaks of monkeypox worldwide emphasized the need for efficient detection of poxviruses in body fluids and the environment, also considering the fact that the human population is largely unprotected. We produced the ectodomains of vaccinia virus A27, A33 and L1 (highly homologous to smallpox and monkeypox proteins) and established their label-free detection by specific antibodies immobilized by silanization on the surface of optical fiber sensors. Each protein was detected using a fiber-optic sensor based on a ball-resonator assisted by a tilted fiber Bragg grating, which ensure sensitivity > 70 dB/RIU and real-time detection. Fabrication of the sensors was carried out through a CO₂ laser splicer, inscribing a spherical tip with ∼500 µm size, and then splicing a previously inscribed tilted grating. The overall spectrum shows a quasi-random pattern due to the ball resonator, modulated by the cladding modes of the grating, in which the intensity of each mode depends on the analyte concentration. Each sensor shows detection limits of 1 fM – 20 pM in serum (<65 aM in phosphate-buffered saline), wide-range response up to 1 μM and a high specificity. This proof of principle for poxvirus detection conceptualizes a general real time pandemic virus preparedness.