The growth of brigatinib-treated cells was determined using the MTT assay. Cells were plated in 96-well plates at 5,000 cells per well and subjected to different treatments. The detailed procedure has been previously described 22 (link). For the colony formation assay, cells were cultured in 24-well plates with different treatments. The colonies were counterstained with Giemsa after 2 weeks, then washed 3 times by PBS. The visible colonies were recorded using a Molecular Imager Gel Do XR+ System (BIO-RAD) and counted using Image J software (NIH). EdU Cell Proliferation Assay Kit (Ribobio, Guangzhou, China) was used to perform the 5-ethynyl-20-deoxyuridine (EdU) labeling assay. The detailed procedure has been previously described 23 (link).
Molecular imager gel do xr system
Manufactured by Bio-Rad
Sourced in United States
The Molecular Imager Gel Do XR+ System is a lab equipment product by Bio-Rad. It is designed for the detection and analysis of nucleic acids and proteins in gel electrophoresis applications.
Automatically generated - may contain errors
9 protocols using molecular imager gel do xr system
1
Cell Growth and Proliferation Assays
2
Colony Formation Assay for HyFS
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Cells were seeded at a density of 300 cells/well in 24-well plates and treated with the indicated concentration of HyFS or vehicle control. The medium was changed every 3 days. After 14 days, the colonies were stained with Giemsa for 15 min and washed three times. Visible colonies were photographed by a Molecular Imager Gel Do XR+ System (Bio-Rad) and counted using Image J software (NIH).
3
Cell Growth, Colony Formation, and Proliferation Analysis
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The MTT assay was used to detect the cell growth rate. Briefly, cells were plated in 96-well plates (4 x 103 per well) and received different concentrations of drug treatment.
The detailed procedures have been described by Dou et al. (2016) (link). The colony formation assay, cells were cultured in 24-well plates (800 cells/well) and treated with different treatments. After 2 weeks, the 4% paraformaldehyde was adopted for fixation and crystal violet was used for dyeing. The visible colonies were captured by Molecular Imager Gel Do XR + System (Bio-Rad, Hercules, CA, USA) and calculated applying ImageJ software (National Institutes of Health, Bethesda, MD, USA). The detailed procedures have been previously described by Wang et al. (2011) (link). The 5-ethynyl-20-deoxyuridine (EdU) labeling analysis was carried out in 24-well plates (2 x 104 cells) using the EdU cell Proliferation Assay Kit (Ribobio, Guangzhou, China). After different concentrations of drug treatment, 10 μmol/L EdU was added to the cells, and the cells were incubated for 12 h at 37°C. Cells were fixed with 4% paraformaldehyde. Then DAPI was utilized for nuclear staining and fluorescence microscopy was applied for imaging.
The detailed procedures have been described by Dou et al. (2016) (link). The colony formation assay, cells were cultured in 24-well plates (800 cells/well) and treated with different treatments. After 2 weeks, the 4% paraformaldehyde was adopted for fixation and crystal violet was used for dyeing. The visible colonies were captured by Molecular Imager Gel Do XR + System (Bio-Rad, Hercules, CA, USA) and calculated applying ImageJ software (National Institutes of Health, Bethesda, MD, USA). The detailed procedures have been previously described by Wang et al. (2011) (link). The 5-ethynyl-20-deoxyuridine (EdU) labeling analysis was carried out in 24-well plates (2 x 104 cells) using the EdU cell Proliferation Assay Kit (Ribobio, Guangzhou, China). After different concentrations of drug treatment, 10 μmol/L EdU was added to the cells, and the cells were incubated for 12 h at 37°C. Cells were fixed with 4% paraformaldehyde. Then DAPI was utilized for nuclear staining and fluorescence microscopy was applied for imaging.
4
Cell Viability and Colony Formation Assays
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Cell sensitivity to the treatments was determined using a WST‐1 cell viability assay. Cells were plated in 96‐well plates (5,000 cells per well) and subjected to different treatments as reported previously (Shen et al, 2019 (link), 2020 (link)). For the colony formation assay, cells were cultured in 24‐well plates with different treatments for 10 days. Giemsa was used to counterstain the colonies, then washed cells three times with cold PBS. Molecular Imager Gel Do XR+ System (BIO‐RAD) and Image J software (NIH), respectively, were used to record or count the visible colonies. For soft agar colony formation assay, 1 ml of 1% agar‐mixed medium and 1 ml of 0.5% agar‐mixed medium with 1 × 103 cells were successively added to the 12‐well plates. Cells were cultured for 2–3 weeks at 37°C, and were observed and captured using a phase contrast microscope (Olympus IX73). For observation of cell morphology, cells were cultured in six‐well plates and treated with different agents or vehicle. Cell morphology was observed and captured using a phase contrast microscope (Olympus IX73).
5
Evaluating HPV16E7 Inhibitor Efficacy
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The SiHa, CaSki, and C666-1 cells were respectively seeded in 6-well plates at a density of 2000 cells per well and exposed to 30 μm of ZHPV16E7384 in complete media for 14 days. The colonies were fixed by 4% paraformaldehyde for 15 min and stained with crystal violet staining solution (Beyotime Biotechnology, Shanghai, China) for 10 min and washed with ultrapure water three times. Visible colonies were photographed by a Molecular Imager Gel Do XR+ System (Bio-Rad, CA, USA) and counted using ImageJ software (NIH, Bethesda, MD, USA). SiHa, CaSki, and C666-1 cells treated with ZWT or PBS were used as the controls.
6
Colony Formation Assay of SW480 Cells
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The SW480 cells were seeded (1×103/well) into 24-well plates and treated with indicated concentrations of TCG or DMSO (or 3-MA according to each experiment requires). Following 7 days of incubation, the colonies were stained with Giemsa for 15 min at 37°C, then washed three times with PBS. The visible colonies were visualized using a Molecular Imager Gel Do XR+ system (Bio-Rad Laboratories, Inc.) and counted using ImageJ 1.47 software (National Institutes of Health).
7
Colony Formation Assay for Cell Viability
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Cells were seeded in 24-well cell culture plates (200 cells/well) and continuously cultured for 14 days. Meanwhile, cells were subjected to the indicated treatments. Clones were stained with crystal violet for 15 minutes and washed three times. The Molecular Imager Gel Do XR + System (BIO-RAD) and Image J software (NIH) were used to photograph or count the visible colonies. This experiment was repeated three times.
8
Evaluation of Sertaconazole's Effects on Cell Growth
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The short‐term effects of sertaconazole on cell growth were evaluated by MTT assay. Briefly, cells were plated in 96‐well plates at 5000 cells peer well and treated for 24 or 48 h, and then incubated with 5 mg/ml MTT (MilliporeSigma, M2128) for 4 h and dissolved in DMSO. The absorbance was measured at 570 nm with a spectrophotometer.
For colony formation assay, NSCLC cells were plated in 24‐well plates (500 cells/well) and then subjected to the indicated concentrations of drugs. After 2 weeks, the colonies were fixed using 4% paraformaldehyde and stained with 0.1% crystal violet, then photographed using a Molecular Imager Gel Do XR+ System (BIO‐RAD). The clone numbers were counted using Image J software.
EdU labeling was performed in 96‐well plates (4000 cells/well) to measure cell proliferation using the EdU Cell Proliferation Assay Kit. After indicated treatments, 10 μM EdU was added to the cells. Then the cells were incubated for 24 h at 37°C and fixed with 4% paraformaldehyde. DAPI was then added for nuclear staining followed by imaging with a fluorescence microscope (Axio Observer 7, ZEISS).
For colony formation assay, NSCLC cells were plated in 24‐well plates (500 cells/well) and then subjected to the indicated concentrations of drugs. After 2 weeks, the colonies were fixed using 4% paraformaldehyde and stained with 0.1% crystal violet, then photographed using a Molecular Imager Gel Do XR+ System (BIO‐RAD). The clone numbers were counted using Image J software.
EdU labeling was performed in 96‐well plates (4000 cells/well) to measure cell proliferation using the EdU Cell Proliferation Assay Kit. After indicated treatments, 10 μM EdU was added to the cells. Then the cells were incubated for 24 h at 37°C and fixed with 4% paraformaldehyde. DAPI was then added for nuclear staining followed by imaging with a fluorescence microscope (Axio Observer 7, ZEISS).
9
Quantification of Colony Formation
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Cells (300 cells/well) were seeded in 24-well plates and treated with the indicated concentration of TCO. After 2 weeks, the colonies were stained with Giemsa for 15 min and washed three times by PBS. The visible colonies were photographed by a Molecular Imager Gel Do XR+ System (Bio-Rad) and counted using Image J software (NIH).
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