Madin-Darby bovine kidney (MDBK) cells and baby hamster kidney (BHK 21) cells were purchased from Sigma-Aldrich Co. (St. Louis, USA). The BHK-21 cells stably expressing human Bcl-2 (BHK 21/Bcl 2) were described previously [22 (link)]. Cells were cultured in Dulbecco’s modified eagle medium (DMEM) supplemented with 10% fetal bovine serum (FBS). One day before each experiment, cells were seeded in 6-cm cell culture dishes with 1 × 106 cells, and incubated at 37 °C with 5% CO2. The 2004/TW/TN1 strain of BEFV was propagated in BHK-21 cells. When 70–80% cytopathic effect (CPE) was shown, the supernatants of BEFV-infected cells were harvested, condensed by PEG 6000 precipitation, dialyzed and resuspended in phosphate-buffered saline (PBS), and finally stored at −70 °C before use.
Bhk 21
Manufactured by Merck Group
Sourced in United States
BHK-21 is a cell line derived from baby hamster kidney cells. It is commonly used for the cultivation of various viruses and the production of biological products. The BHK-21 cell line provides a reliable and well-established in vitro system for a range of applications in the field of virology and biotechnology.
Automatically generated - may contain errors
Lab products found in correlation
Dulbecco modified eagle medium (dmem), by Merck Group (3 mentions)
Pen strep, by Thermo Fisher Scientific (1 mentions)
Pen strep, by Merck Group (1 mentions)
C6 36, by Merck Group (1 mentions)
Bhk 21, by Health Science Research Resources Bank (1 mentions)
Elisa reader, by Agilent Technologies (1 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions)
Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (1 mentions)
Fetal bovine serum (fbs), by Merck Group (1 mentions)
Fetal bovine serum (fbs), by Cytiva (1 mentions)
Penicillin, by Cytiva (1 mentions)
Streptomycin, by Cytiva (1 mentions)
Non essential amino acids (neaa), by Cytiva (1 mentions)
5 protocols using bhk 21
1
Cultivation and Propagation of Cell Lines
2
Virus Isolation using Mammalian and Mosquito Cells
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Two mammalian cell lines, Vero [Japanese Collection of Research Bioresources (JCRB), Osaka, Japan] and BHK-21 (JCRB), and one mosquito cell line, C6/36 (Health Science Research
Resources Bank, Osaka, Japan), were used for virus isolation. Vero and BHK-21 cells were cultured in Dulbecco’s modified eagle’s medium (DMEM; Sigma-Aldrich, St. Louis, MO, USA)
containing heat-inactivated 5% fetal bovine serum (FBS; Hyclone Laboratories, South Logan, UT, USA) and 1% penicillin and streptomycin (Pen-Strep; Life Technologies, Carlsbad, CA,
USA), and C6/36 cells were cultured in Eagle’s minimum essential medium (Sigma-Aldrich) containing heat-inactivated 10% FBS and 1% Pen-Strep. Cell lines were maintained at 37°C for
mammalian cells and 28°C for mosquito cells under 5% CO2.
Resources Bank, Osaka, Japan), were used for virus isolation. Vero and BHK-21 cells were cultured in Dulbecco’s modified eagle’s medium (DMEM; Sigma-Aldrich, St. Louis, MO, USA)
containing heat-inactivated 5% fetal bovine serum (FBS; Hyclone Laboratories, South Logan, UT, USA) and 1% penicillin and streptomycin (Pen-Strep; Life Technologies, Carlsbad, CA,
USA), and C6/36 cells were cultured in Eagle’s minimum essential medium (Sigma-Aldrich) containing heat-inactivated 10% FBS and 1% Pen-Strep. Cell lines were maintained at 37°C for
mammalian cells and 28°C for mosquito cells under 5% CO2.
3
Cytotoxicity Evaluation of Materials
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Potential Cytotoxicity of the studied material was examined using the method of Skehan et al., [25 (link)] in the Clinical Pharmacy Department at the National Cancer Institute, Egypt, to obtain the IC50 value which is the half-maximal inhibitory concentration to measure the potency of a substance in inhibiting a specific biological or biochemical function. Normal cell line (baby hamster kidney cells, BHK-21, Sigma-Aldrich, Germany) was plated in 96-multiwell plate (104 cells/well) for 24 h before application of the tested material to allow attachment of the cells to the wall of the plate. Different concentrations of the attested material (0, 1, 2.5, 5, and 10 μg/ml) were added to the cell monolayer triplicate wells prepared for each dose. Monolayer cells were incubated with the material for 24 h at 37°C and in an atmosphere of 5% CO2. After 48 h, cells were fixed, washed, and stained with sulforhodamine B stain. Excess stain was washed with acetic acid, and the attached stain was removed with Tris-EDTA buffer. Colour intensity was measured in an enzyme-linked immunosorbent assay reader (ELISA reader, BioTek, USA). The relation between surviving fraction and material concentration is plotted to get the survival curve of each cell line after specified compound concentration.
4
Chikungunya virus replicon cell line protocol
5
Establishment and Characterization of Chikungunya Virus Replicon Cell Line
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BHK-21 cells were purchased from The Global Bioresource Center (ATCC) and maintained in Dulbecco's modified Eagle's medium (DMEM, Sigma-Aldrich) supplemented with 100U/mL of penicillin (Hyclone Laboratories), 100 mg/mL of streptomycin (Hyclone Laboratories), 1% dilution of stock of non-essential amino acids (Hyclone Laboratories) and 10% of fetal bovine serum (FBS, Hyclone Laboratories) in a humidified 5% CO2 incubator at 37 °C. BHK-21-Gluc-nSP-CHIKV-99659 cell line, harboring a replicative CHIKV replicon expressing Gaussia luciferase (Gluc) as a reporter gene, was maintained in DMEM 10% FBS with 500 µg/ml G418 (Sigma-Aldrich). The CHIKV replicon construct includes a T7 bacteriophage promotor followed by the viral 5' UTR region, the nsp1-4 coding sequence, the CHIKV subgenomic promoter (Sg) followed by the GLuc sequence and the expression cassette containing a ubiquitination sequence (Ubi) and the neomycin phosphotransferase gene (Neo-resistance gene), and the viral 3' UTR region. This construction and the development of this replicon cell line will be described elsewhere. The CHIKV expressing nanoluciferase reporter (CHIKV-nanoluc) used for the antiviral assays is based on the CHIKV isolate LR2006OPY1 (East/Central/South African genotype) and was produced, rescued, and titrated as previously described40 ,41 (link).
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