Development and Characterization of an HPV Type-16 Specific Modified DNA Aptamer for the Improvement of Potency Assays

Abstract

Measuring vaccine potency is critical for vaccine release and is often accomplished using antibody-based ELISAs. Antibodies can be associated with significant drawbacks that are often overlooked including lot-to-lot variability, problems with cell-line maintenance, limited stability, high cost, and long discovery lead times. Here, we address many of these issues through the development of an aptamer, known as a slow off-rate modified DNA aptamer (SOMAmer), which targets a vaccine antigen in the human papillomavirus (HPV) vaccine Gardasil. The aptamer, termed HPV-07, was selected to bind the Type 16 virus-like-particle (VLP) formed by the self-assembling capsid protein L1. It is capable of binding with high sensitivity (EC of 0.1 to 0.4 μg/mL depending on assay format) while strongly discriminating against other VLP types. The aptamer competes for binding with the neutralizing antibody H16.V5, indicating at least partial recognition of a neutralizing and clinically relevant epitope. This makes it a useful reagent for measuring both potency and stability. When used in an ELISA format, the aptamer displays both high precision (intermediate precision of 6.3%) and a large linear range spanning from 25% to 200% of a typical formulation. To further exploit the advantages of aptamers, a simplified mix and read assay was also developed. This assay format offers significant time and resource reductions compared to a traditional ELISA. These results show aptamers are suitable reagents for biological potency assays, and we expect that their implementation could improve upon current assay formats.

Advances in detection of infectious agents by aptamer-based technologies

“…Tuberculosis [9] H63SL2-M6 HspX Aptamer linked immobilized sorbent assay and electrochemical sensor Tuberculosis [18] Thiolated MPT64 aptamer MPT64 Combination of aptamers and electrochemical impedance spectroscopy Tuberculosis [19] MAA I and MAA II MPT64 A sandwich-type electrochemical aptasensor Salmonella [10] LA27 LPS Truncated aptamer LA27 combined with GO-based FP assays Salmonella [20] Apt-E. coli Apt-S. typ E. coli S. typ Aptamer modified by fluorescent magnetic multifunctional nanoprobe (APT-FMNPs) S. aureus [21] Chimeric [33] HCVcp ssDNA aptamer HCVcp A plasmonic nanoplatform Human papillomavirus [34] HPV16 L1 ssRNA aptamer HPV16 L1 protein The laser desorption/ionization mass spectrometry method Human papillomavirus [35] HPV-07 HPV16 virus-like particle (VLP) Slow abscission rate-modified DNA aptamer Influenza [37] NP-specific aptamer, CTnI-specific aptamer Influenza nucleoprotein (NP) and cardiac troponin I (cTnI).…”

“…Several problems of antibodies like batch variability, limited stability, and long discovery lead time were overcome by developing slow abscission rate-modified DNA aptamer (SOMAmer), which targets antigens in the HPV vaccine [35]. An HPV-07 aptamer was selected to bind to the type 16 virus-like particle (VLP) formed by the self-assembled capsid protein L1.…”

Chemical Modification of Aptamers for Increased Binding Affinity in Diagnostic Applications: Current Status and Future Prospects

“…In a similar approach, a SOMAmer named HVP-07 was selected against the Type 16 virus-like particle (VLP) and used in a sandwich-like mix and read assay. The use of SOMAmers in this assay greatly improved the simplicity of the system (no wash steps required) and allowed for a fast read-out [ 124 ].…”

Aptamers for Anti-Viral Therapeutics and Diagnostics

“…When this aptamer was used in ELISA, the aptamer showed very high specificity. They demonstrated that this assay format offered more time and required fewer resources than traditional ELISA [10]. Aspermair et al developed a reduced graphene oxide-based field-effect transistor (rGO-FET) for detecting the HPV E7 protein in saliva using RNA aptamers.…”