Infection of Huh7 and SW13 Cells with CCHFV

Huh7 and SW13 cells were infected with the CCHFV in triplicate, as described by us previously (Appelberg et al., 2020 (link); Krishnan et al., 2021 (link)). Briefly, Huh7 cells were infected with CCHFV IbAr10200 at a multiplicity of infection (MOI) of 1. After 1 hr of incubation (37 °C, 5% CO₂) the inoculum was removed, the cells were washed with PBS, and 2 ml DMEM supplemented with 5% heat-inactivated FBS was added to each well. Samples were collected in triplicate at 24 and 48hpi along with controls. Due to high permissiveness, we restricted the SW13 infection of 1 MOI for 24 hr only. The infection in Huh7 24hpi was confirmed by immunofluorescence staining of CCHFV nucleoprotein-protein. The cells were fixed in ice-cold acetone-methanol (1:1) and stained using a rabbit polyclonal anti-CCHFV nucleocapsid antibody (home-made) followed by a fluorescein isothiocyanate (FITC)-conjugated anti-rabbit antibody (Thermo Fisher Scientific, US) and DAPI (Roche, US).

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Neogi U., Elaldi N., Appelberg S., Ambikan A., Kennedy E., Dowall S., Bagci B.K., Gupta S., Rodriguez J.E., Svensson-Akusjärvi S., Monteil V., Vegvari A., Benfeitas R., Banerjea A., Weber F., Hewson R, & Mirazimi A. (2022). Multi-omics insights into host-viral response and pathogenesis in Crimean-Congo hemorrhagic fever viruses for novel therapeutic target. eLife, 11, e76071.

Publication 2022

FITC)-conjugated anti-rabbit antibody DAPI

Acetone Anti antibody Cells Cold Dapi Fluorescein Immunofluorescence Infection Isothiocyanate Methanol Nucleocapsid Nucleoprotein Protein Rabbit

Corresponding Organization :

Other organizations : Karolinska Institutet, Manipal Academy of Higher Education, Sivas Cumhuriyet Üniversitesi, Public Health Agency of Sweden, Public Health England, Oxford Brookes University, Science for Life Laboratory, Stockholm University, National Institute of Immunology, University of Giessen, London School of Hygiene & Tropical Medicine, Swedish Veterinary Agency

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Variable analysis

independent variables

CCHFV infection in Huh7 cells at a multiplicity of infection (MOI) of 1
CCHFV infection in SW13 cells at a multiplicity of infection (MOI) of 1

dependent variables

Viral infection at 24 and 48 hours post-infection (hpi) in Huh7 cells
Viral infection at 24 hours post-infection (hpi) in SW13 cells

control variables

Incubation of infected cells at 37 °C, 5% CO2
Washing of infected Huh7 cells with PBS after 1 hour of infection
Addition of 2 ml DMEM supplemented with 5% heat-inactivated FBS to each well of infected Huh7 cells

controls

Uninfected Huh7 and SW13 cells as negative controls

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