Detection of Smallpox by the Laboratory Method of ELISA Immunoassay after Its Inactivation with Formaldehyde

The evolutionary variability of the monkeypox virus requires improvement of differential diagnostics. In the Russian Federation, the virus is classified as one of the most dangerous pathogens. A complex procedure for delivering samples to the laboratory for identification is required. On-site analysis or sample inactivation and preservation without loss of specific analytes can simplify the procedure. Formaldehyde treatment is considered reliable. The ELISA method can confirm the results of laboratory detection of viral DNA, provided that it is adapted to the analysis of treated samples. The antigen sample was prepared by infecting a monolayer of Vero cells with the hMpxV/Russia/St.Petersburg-02/2022 strain, followed by inactivation of one aliquot with 8% formaldehyde solution. To detect the antigen in the native and inactivated sample, an experimental test system containing a peroxidase conjugate of monoclonal antibodies to the A29L protein as the main reagent was used in noninstrumental and instrumental formats. The virus was detected without use of instruments in the native sample and using instruments in the inactivated sample without any difference in sensitivity. Formaldehyde inactivation reduced the detection sensitivity of the treated antigen in the noninstrumental ELISA format by 100 times with an equal increase in the detection sensitivity of the native antigen in the instrumental format. For differential diagnostic purposes, cases of monkeypox can be confirmed by ELISA on-site of the native antigen or delivered to the laboratory a formaldehyde-treated antigen preparation.