2.4. Indirect immunofluorescence test (iIFT)

To confirm the results of both ELISAs, indirect immunofluorescence test (iIFT) was performed. This assay was conducted for samples (n = 32), which yielded either positive or inconclusive results in one of the two ELISAs. Due to a lack of material, one of the respective sera could not be confirmed by iIFT. A recently developed assay was used (Schlottau et al., 2020). Vero E6 cells were infected with the 2019_nCoV Muc‐IMB‐1 SARS‐CoV‐2 isolate at a multiplicity of infection (MOI) of 0.1. Twenty‐four hours after infection, cells were fixed (4% paraformaldehyde) and permeabilized with 0.5% Triton‐X‐100. Heat inactivated and serially diluted sera were added. The cells were incubated for 1 hr. A washing step was conducted afterwards and an anti‐cat‐IgG‐FITC (Sigma‐Aldrich) antibody diluted 1/600 was applied. This was followed by washing the cell monolayer one more time. After adding a fluorescence preservation buffer, the assay was evaluated by fluorescence microscopy. Values ≥1/8 were considered as positive.