Roswell Park Memorial Institute (RPMI)-1640 medium was bought from HyClone (Logan, UT, USA). Fetal bovine serum (FBS) was obtained from Thermo Scientific (Gibco, Shanghai, China). Cell counting kit-8 (CCK-8) was obtained from Dojindo Molecular Technologies (Tokyo, Japan). Dimethyl sulfoxide (DMSO) and Hoechst 33342 staining solution were purchased from Solarbio (Beijing, China). Primary antibodies against Bcl-2, Bax and cytochrome c were supplied by Proteintech (Rosemont, IL, USA). Anti-cleaved caspase-9 and anti-cleaved caspase-3 antibodies were purchased from Cell Signaling Technology (CST, Danvers, MA, USA). Antibodies against Ki-67, PCNA, Snail, E-cadherin, Vimentin and N-cadherin were acquired from Abcam (Cambridge, MA, USA).
Hoechst 33342 staining solution
Manufactured by Solarbio
Sourced in China
Hoechst 33342 staining solution is a fluorescent dye commonly used in cell biology and biotechnology applications. It binds to the minor groove of double-stranded DNA, allowing for the visualization and quantification of DNA content in cells.
Automatically generated - may contain errors
Lab products found in correlation
Anti cleaved caspase 3, by Cell Signaling Technology (1 mentions)
Anti cleaved caspase 9, by Cell Signaling Technology (1 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions)
Proliferating cell nuclear antigen (pcna), by Abcam (1 mentions)
Vimentin, by Abcam (1 mentions)
Dimethyl sulfoxide (dmso), by Solarbio (1 mentions)
E cadherin, by Abcam (1 mentions)
Snail, by Abcam (1 mentions)
N cadherin, by Abcam (1 mentions)
Cell counting kit 8 cck 8, by Dojindo Laboratories (1 mentions)
Cytochrome c, by Proteintech (1 mentions)
Dmil microscope, by Leica (1 mentions)
Pi staining solution, by Solarbio (1 mentions)
Fsx100 microscope, by Olympus (1 mentions)
Dmi6000b, by Leica (1 mentions)
One step tunel assay kit, by Roche (1 mentions)
Tcs sp8, by Leica (1 mentions)
Reactive oxygen species kit, by Beyotime (1 mentions)
Sw480, by American Type Culture Collection (1 mentions)
T1 sam, by Nikon (1 mentions)
10 protocols using hoechst 33342 staining solution
1
Apoptosis and EMT Regulation Assays
2
Simultaneous Detection of Cell Viability
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Damping fluid (2 ml), Hoechst 33342 staining solution (10 µl) and PI staining solution (10 µl) (Solarbio Science and Technology Company, Ltd.) were added to each well and mixed lightly. Cells were kept in the dark for 20 min at 4°C. The staining solution was then discarded and the cells were rinsed with 1X PBS. Physiological saline (1 ml) was added, and fluorescent images were captured using a Leica DMIL microscope (magnification, ×200). Normal cells stained blue and the nucleolus structure was normal. Apoptotic cells stained bright blue and light red, and necrotic cells stained light blue and bright red.
3
Hoechst 33342 Nuclear Staining Protocol
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After RAW 264.7 cells were treated with or without LPS for 24 h, Hoechst 33342 staining solution (Solarbio, Peking City, China; #C0030) was added to coat the cells. After incubation at 37°C for 20 min, the cells were washed with phosphate‐buffered saline for three times and then observed with an FSX100 microscope (Olympus).
4
Pyroptosis Detection by TUNEL and PI Staining
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Pyroptosis is characterized by loss of cell membrane potential and DNA fragmentation which can be detected by TUNEL staining36 (link). The frozen sections of aortic root were permeabilized with 0.3% Triton X-100 for 15 min at room temperature. Then, the TUNEL reaction mixtures from one-step TUNEL assay kit (Roche, Switzerland) were applied for 2 h at 37 °C in the dark. Subsequently, sections were sealed with goat serum for 30 min and incubated with primary antibody MOMA2 (1:200) and the corresponding secondary antibody (1:300). Finally, DAPI was used to counterstain nucleus and the samples were visualized using Olympus fluorescence microscope. Double-fluorescent staining with Hoechst 33342 and PI was used to assess the formation of membrane pores during pyroptosis. The cells were stained with Hoechst 33342 staining solution (Solarbio, Beijing) for live cells and 2 μg/ml PI for 20 min in 5% CO2 at 37 °C. Then the cells were washed three times with PBS. The fluorescences of Hoechst 33342 and PI were detected by a Inverted fluorescence microscope (Leica DMI6000B, Germany) and the percentage of PI-positive cells was evaluated by Image J software.
5
Cellular Uptake of ICG-loaded Nanoparticles
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2 × 105 4T1 cells (purchased from Hanyin Biotechnology Co., Ltd.) were seeded on a special culture dish for confocal laser (CLSM) (Diameter/size: 35mm/15mm, Catalog No. 801002, Nestle Biotechnology Co., Ltd., Wuxi, China), and cultured in a cell incubator under normal conditions (37°C, 5% CO2, 95% humidity), when the cell density reached 70–80%, the original medium was discarded, and 10 µL of ICG@SANPs-cRGD and ICG@SANPs were added to incubate for 1 h, and then the liquid was discarded, washed twice with PBS, 10μg/mL Hoechst 33342 Staining Solution (1mg/mL, Solarbio Technology Co., Ltd. Beijing, China) was added into each dish. After staining for 5min, PBS buffer was used to clean the stain for 3 times under dark conditions, 5min each time. The uptake of ICG@SANPs-cRGD and ICG@SANPs by 4T1 cells was observed under a confocal laser scanning microscope (CLSM) (Leica TCS SP8, Germany) with an excitation wavelength of 750 nm and an emission wavelength of 817nm.
6
Detecting ROS and Apoptosis in IBV Infection
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The reactive oxygen species kit (Beyotime, China) was used to detect the changes of reactive oxygen species in cells caused by IBV infection. The apoptotic changes after IBV infection were observed with the Hoechst 33,342 staining solution (Solarbio, China). HD11 cells were cultured in six-well plates. After 1 h of IBV infection in HD11 cells, the supernatant was discarded and washed with PBS buffer. The samples were incubated with 10 μM, 25 μM, 50 μM APL, and 2 mM NAC for 2 h, washed with PBS buffer, and added with 2% maintenance medium for 36 h. After the supernatant was discarded, 1 mL DEFH-DA was added to each well for 20 min at 37°C. The supernatant was discarded, washed three times with DMEM, and then 1 mL Hoechst 33342 was added into each well. After incubation at 37°C for 20 min, the results were observed under the fluorescence microscope.
7
Hoechst Staining for Cell Apoptosis
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Hoechst staining was conducted immediately after the respective treatments to observe cell apoptosis.
Myotubes were washed twice with precooled sterile PBS to remove debris. Then, 4% paraformaldehyde was added to the six-well plate for 15 min to x the cells. After that, PBS was used to wash the cells 2-3 times. Hoechst 33342 staining solution (Solarbio, Beijing, China, C0030) was added to six-well plates at a concentration of 10 µg/ml. Staining at room temperature was carried out in a dark environment for 15 min. The maximum excitation wavelength of Hochest 33342 is 461 nm, and the maximum emission wavelength is 460 nm. After staining, myotubes were washed with PBS 2-3 times for 3 minutes each time. The cells were observed and imaged under a uorescence microscope.
Myotubes were washed twice with precooled sterile PBS to remove debris. Then, 4% paraformaldehyde was added to the six-well plate for 15 min to x the cells. After that, PBS was used to wash the cells 2-3 times. Hoechst 33342 staining solution (Solarbio, Beijing, China, C0030) was added to six-well plates at a concentration of 10 µg/ml. Staining at room temperature was carried out in a dark environment for 15 min. The maximum excitation wavelength of Hochest 33342 is 461 nm, and the maximum emission wavelength is 460 nm. After staining, myotubes were washed with PBS 2-3 times for 3 minutes each time. The cells were observed and imaged under a uorescence microscope.
8
Oxaliplatin Resistance in Colorectal Cancer
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Human colon cancer cell line SW480 and human embryonic kidney cell line 293T were purchased from ATCC. To induce OXA resistant SW480 cells, the cells were seeded onto 24 well plate before chemotherapeutics treatment. Next, SW480 cells were cultured in the presence of 0.5 mM oxaliplatin (OXA) for 24 hours followed by three days in fresh medium without a drug. This procedure was continued for six months with drug concentration increase 0.4μM per month, and the nal concentration is 2.5 mM.
Both of SW480 and SW480/OXA cells were cultured in Dulbecco Modi ed Eagle Medium (DMEM) with 10% fetal bovine serum (FBS) and 1% penicillin/streptomycin solution in a cell incubator at 37 °C, 5% CO 2 , with saturated humidity.
Hoechst 33342 staining Cells were seeded onto bronectin coated 12 well plate, after treatment, 5 mg/ml Hoechst 33342 staining solution (Solarbio, Beijing, China) was added to cells and incubated for 20 mins at room temperature. Then cells were washed with PBS for three time. Images were collected by Nikon T1-SAM.
Both of SW480 and SW480/OXA cells were cultured in Dulbecco Modi ed Eagle Medium (DMEM) with 10% fetal bovine serum (FBS) and 1% penicillin/streptomycin solution in a cell incubator at 37 °C, 5% CO 2 , with saturated humidity.
Hoechst 33342 staining Cells were seeded onto bronectin coated 12 well plate, after treatment, 5 mg/ml Hoechst 33342 staining solution (Solarbio, Beijing, China) was added to cells and incubated for 20 mins at room temperature. Then cells were washed with PBS for three time. Images were collected by Nikon T1-SAM.
9
Hoechst Staining for Cell Apoptosis
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Hoechst staining was conducted immediately after the respective treatments to observe cell apoptosis.
Myotubes were washed twice with precooled sterile PBS to remove debris. Then, 4% paraformaldehyde was added to the six-well plate for 15 min to x the cells. After that, PBS was used to wash the cells 2-3 times. Hoechst 33342 staining solution (Solarbio, Beijing, China, C0030) was added to six-well plates at a concentration of 10 µg/ml. Staining at room temperature was carried out in a dark environment for 15 min. The maximum excitation wavelength of Hochest 33342 is 461 nm, and the maximum emission wavelength is 460 nm. After staining, myotubes were washed with PBS 2-3 times for 3 minutes each time. The cells were observed and imaged under a uorescence microscope.
Myotubes were washed twice with precooled sterile PBS to remove debris. Then, 4% paraformaldehyde was added to the six-well plate for 15 min to x the cells. After that, PBS was used to wash the cells 2-3 times. Hoechst 33342 staining solution (Solarbio, Beijing, China, C0030) was added to six-well plates at a concentration of 10 µg/ml. Staining at room temperature was carried out in a dark environment for 15 min. The maximum excitation wavelength of Hochest 33342 is 461 nm, and the maximum emission wavelength is 460 nm. After staining, myotubes were washed with PBS 2-3 times for 3 minutes each time. The cells were observed and imaged under a uorescence microscope.
10
Hoechst Staining for Cell Apoptosis
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Hoechst staining was conducted immediately after the respective treatments to observe cell apoptosis.
Myotubes were washed twice with precooled sterile PBS to remove debris. Then, 4% paraformaldehyde was added to the six-well plate for 15 min to x the cells. After that, PBS was used to wash the cells 2-3 times. Hoechst 33342 staining solution (Solarbio, Beijing, China, C0030) was added to six-well plates at a concentration of 10 µg/ml. Staining at room temperature was carried out in a dark environment for 15 min. The maximum excitation wavelength of Hochest 33342 is 461 nm, and the maximum emission wavelength is 460 nm. After staining, myotubes were washed with PBS 2-3 times for 3 minutes each time. The cells were observed and imaged under a uorescence microscope.
Myotubes were washed twice with precooled sterile PBS to remove debris. Then, 4% paraformaldehyde was added to the six-well plate for 15 min to x the cells. After that, PBS was used to wash the cells 2-3 times. Hoechst 33342 staining solution (Solarbio, Beijing, China, C0030) was added to six-well plates at a concentration of 10 µg/ml. Staining at room temperature was carried out in a dark environment for 15 min. The maximum excitation wavelength of Hochest 33342 is 461 nm, and the maximum emission wavelength is 460 nm. After staining, myotubes were washed with PBS 2-3 times for 3 minutes each time. The cells were observed and imaged under a uorescence microscope.
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