Cell Culture Protocols for Virus Research

SW13, HEK293T, Vero E6 (all from ATCC), Huh7, HUVEC (provided by National Virus Resource Center, Wuhan, China) and DLD1 (provided by Dr. Youjun Li, Wuhan University) cells were cultured at 37 °C in Dulbecco’s modified Eagle’s medium (DMEM) (GIBCO) supplemented with 10% (v/v) fetal bovine serum (FBS) and 1% (v/v) penicillin-streptomycin (Hyclone). The primary mouse hepatocytes and MLF cells were isolated and cultured as previously described.^{55 (link),56 (link)} Primary human PBMCs were isolated from whole blood of healthy donors with SepMate™-15 (STEMCELL Technologies) according to the manufacturer’s instructions and cultured in RPMI 1640 medium supplemented with 10% (v/v) fetal bovine serum (FBS) and 1% (v/v) penicillin-streptomycin. The isolation of human PBMCs and its application for virus infection were approved by Institutional Review Board, Wuhan Institute of Virology, Chinese Academy of Sciences (Approval Number: WIVH31202301). FreeStyle 293F cells (provided by Dr. Bing Yan, Wuhan Institute of Virology, Chinese Academy of Sciences) were cultured at 37 °C in SMM 293-TII Expression Medium (SinoBiological) on an orbital shaker platform rotating at 120 rpm. All cell lines were tested and found to be free of mycoplasma contamination using the MycoBlue Mycoplasma Detector kit (#D101, Vazyme).

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Xu Z.S., Du W.T., Wang S.Y., Wang M.Y., Yang Y.N., Li Y.H., Li Z.Q., Zhao L.X., Yang Y., Luo W.W, & Wang Y.Y. (2024). LDLR is an entry receptor for Crimean-Congo hemorrhagic fever virus. Cell Research, 34(2), 140-150.

Publication 2024

DMEM penicillin-streptomycin SepMate™-15 SMM 293-TII Expression Medium MycoBlue Mycoplasma Detector kit

Corresponding Organization :

Other organizations : Wuhan Institute of Virology, Chinese Academy of Sciences

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Variable analysis

independent variables

Cell lines used (SW13, HEK293T, Vero E6, Huh7, HUVEC, DLD1, primary mouse hepatocytes, MLF cells, primary human PBMCs, FreeStyle 293F cells)

dependent variables

Virus infection

control variables

Culturing conditions (37 °C, Dulbecco's modified Eagle's medium (DMEM) supplemented with 10% (v/v) fetal bovine serum (FBS) and 1% (v/v) penicillin-streptomycin)
Mycoplasma contamination testing (found to be free of mycoplasma contamination using the MycoBlue Mycoplasma Detector kit)

positive controls

Not explicitly mentioned

negative controls

Not explicitly mentioned

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