Aptamers that bind filovirus GP : a new frontier for detection and neutralization of Ebola and Marburg virus

Abstract

Aptamers are single chained, nucleic acid-based affinity probes that bind to their targets with strong affinity and specificity. They are made through an in vitro combinatorial selection method, wherein large libraries of nucleic acids with randomized sequences are subjected to an iterative process of affinity enrichment, partitioning, and amplification. Evolved libraries are sequenced, and the individual sequences are screened and characterized for their structure and function. Aptamers have been developed to target many molecules, including small molecules, purified proteins, whole cells, bacteria, and viruses. They have been developed for therapeutics and as research and diagnostic probes. Aptamers that have an affinity for virus surfaces are excellent probes for developing low-cost biosensors and potentially antiviral therapeutics. In this work, I present the development of aptamers that have an affinity for filovirus surfaces. I first describe the development of an improved method for purifying highly lytic vesicular stomatitis virus-based filovirus GP displayed surrogate viruses. Filoviruses are highly pathogenic and thus require highly secured containment facilities for their studies. The use of attenuated surrogates facilitates filovirus research at biosafety level 2 facilities. This work outlines the steps required to propagate and generate pure virus particles to be used as selection targets. I then describe the development of aptamer probes that differentially recognize GPs from MARV and EBOV. This work represents the first step in the development of aptamer based-low-cost point of care devices for filovirus disease diagnostics. Finally, this work describes the use of a hybrid selection approach that combines two different selection platforms to generate aptamers that bind to EBOV surfaces.