When it was noticed that 5D-8.1 may cross-react with human proteins, doubt was casted on the significance of the above histopathologic findings. To address this issue, properties of EV antibodies 5D-8.1 and 9D5 have been investigated using peptide microarrays, peptide substitution scanning, immunofluorescence of EV-infected cells, EV neutralization assays, bioinformatics analysis. Evidence indicates that the two antibodies bind to distinct non-neutralizing linear epitopes in VP1 and are specific for a vast spectrum of EV types (not for other human viruses). However, their epitopes may align with a few human proteins at low expected values. When tested by immunofluorescence, high concentrations of 5D-8. Several recent reports point to enterovirus (EV) infections as key environmental triggers of type 1 diabetes (T1D) 1-7 . Conclusions are based on multiple proofs that include the histopathologic detection of EV antigens/ genome in the islets of Langerhans of diabetics at different stages of the disease 8-12 . Recent findings suggest that EVs are causing chronic low-level infection in the islet cells of newly diagnosed T1D patients 12 . EVs are small, non-enveloped, single-strand positive-sense RNA viruses belonging to the family Picornaviridae. The EV capsid is composed of four structural proteins named VP1, VP2, VP3, and VP4. The main structural differences among VP1, VP2, and VP3 lie in the loops that connect the beta-strands with the N-and C-terminal sequences extending from the beta-barrel domain 13 . These amino acid (AA) sequences give each EV its distinct morphology and antigenicity. The VP4 component lies on the inner surface of the shell and is essential for virion stability. Evolution has resulted in a large number of antigenically distinguishable members that have been categorized as EV "serotypes" 13 . Not considering human rhinoviruses, the EV genus contains human agents of the A, B, C, D, and unnamed species that, together, comprise 109 different types. Each of the serotypes correlates with a specific immune response of the host and protection from disease. The serotype-specific protective immune response is directed to the capsid proteins VP1, VP2, and VP3, as the VP4 has no role in the interaction with neutralizing antibodies (i.e., those directed to variable regions of surface capsid proteins). Non-neutralizing antibodies recognizing VP regions that are conserved among different EV types are also produced. The significance of the non-neutralizing antibody response is currently under investigation 14 . Immunization with different EV types allowed to produce a variety of monoclonal antibodies (MAbs) that are either type-specific (i.e., responsible of virus neutralization) or directed to conserved regions of capsid proteins 15-18 . Reactivity of the latter antibodies may be limited to sets of different EV types or may be directed to a wider range of EV types. Though molecular methods are held to be more informative than classical serologic methods for virus identification 19 , "pan-EV antibodies" capable of reacting with all or with the majority of EV types remain desirable reagents for detecting these agents both in the diagnosis of infectious diseases 20-23 and in immunohistochemistry 24 .
Output: Properties of Two Enterovirus Antibodies that are Utilized in Diabetes Research OPEN