The Chinese Hamster Lung cell line was purchased from the American Type Culture Collection (ATCC, U.S.A.) on Nov. 24, 2011. The modal chromosome number of this cell line is 25. The doubling time is approximately 15 h. Cells were cultured in Eagle’s Minimum Essential Medium (EMEM, Lonza Walkersville, U.S.A.) supplemented with 10% fetal bovine serum (FBS, Gibco, U.S.A.) and incubated in a 5% CO2 incubator (MCO-20AIC, SANYO, Japan) at 37 °C. The cell line was evaluated for contamination of mycoplasma with the Hoechst Stain Kit (MPBIOMEDICALS, Japan). Sub-culturing was done approximately twice a week.
Hoechst stain kit
Manufactured by MP Biomedicals
Sourced in Japan
The Hoechst Stain Kit is a fluorescent dye used to detect and quantify DNA in biological samples. It binds to the minor groove of DNA, emitting a fluorescent signal upon excitation. The kit provides a simple and reliable method for DNA labeling and analysis.
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3 protocols using hoechst stain kit
1
Cultivation of Chinese Hamster Lung Cells
2
Cryopreservation of CHL/IU Cell Line
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The CHL/IU cell line was selected due to its high detection sensitivity and frequent use in chromosomal aberration tests [15 (link)]. CHL/IU cells were purchased from American Type Culture Collection (USA) on November 24, 2011, placed in a 75 cm2 flask (Nunc, Denmark) containing Eagle’s Minimum Essential medium (EMEM, Lonza Walkersville Inc., USA) and 10% fetal bovine serum (FBS, Gibco, USA), and cultured in a 37℃ incubator (MCO-20AIC, SANYO, Japan) supplemented with 5% CO2. CHL/IU cells were checked for mycoplasma contamination using the Hoechst Stain Kit (MPBIOMEDICALS, Japan), and 0.25% Trypsin-EDTA solution was added to the culture flask to separate the cells from the bottom of the flask. After centrifugation of the cell suspension for five minutes at 1,000 rpm, the supernatant was discarded. FBS was added to a concentration of 1 × 106 cells/mL, DMSO was added to a final concentration of 10%, and the cells were aliquoted into a tube for freezer storage.
3
CHL/IU Cell Line Cultivation Protocol
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Chinese hamster lung (CHL/IU) cell (ATCC, CRL-1935) line was obtained from American Type Culture Collection (ATCC, U.S.A.). Cells were maintained in Eagle’s Minimum Essential Medium (EMEM, Lonza Walkersville Inc., U.S.A.) supplemented with 10% heat-inactivated fetal bovine serum (FBS, Invitrogen, U.S.A.), 100 units/ml penicillin and 100 μg/ml streptomycin (Invitrogen, U.S.A.)4 (link). Then, cells were incubated to 70–80% confluent at 37 °C with 5% CO2 prior to subculture. Mycoplasma contamination was regularly evaluated by Hoechst Stain Kit (MPBIOMEDICALS, Japan). Cells within 13 passages were routinely used to detect chromosomal aberration.
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