HNE1 cells were diluted to 5 × 103 cells/mL in 0.4% noble agar solution in DMEM with 20% FBS. The cell suspension was added to each well of a 24-well plate on an underlayer of 0.8% noble agar in DMEM with 20% FBS at 37℃. Fresh DMEM containing 0.2% FBS with or without HGF (10 ng/mL) was added to the top of the soft agar and then cultured at 37℃ for 21 days. The culture medium was changed twice per week with or without HGF. The colonies were stained with a 0.05% crystal violet solution (dissolved in 20% methanol) and the stained colonies were counted using the MetaMorph NX image software (Molecular Devices, Sunnyvale, CA, USA). Each experiment was performed in triplicate.
Metamorph nx
Manufactured by Molecular Devices
Sourced in United States
Metamorph NX is a comprehensive software package for controlling microscope hardware and acquiring, processing, and analyzing images. It provides a flexible and customizable platform for a wide range of imaging applications.
Automatically generated - may contain errors
Lab products found in correlation
Inverted microscope, by Zeiss (1 mentions)
Axiovision 4, by Zeiss (1 mentions)
Bovine serum albumin (bsa), by Merck Group (1 mentions)
Formamide, by Avantor (1 mentions)
Axio imager z1 fluorescent microscope, by Zeiss (1 mentions)
Confocal tcs sp5, by Leica (1 mentions)
Imagej, by Adobe (1 mentions)
8 protocols using metamorph nx
1
Anchorage-independent Growth of HNE1 Cells
2
Soft Agar Colony Formation Assay
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HNE1 cells were diluted to 5 × 103 cells/ml in 0.4% Noble agar solution in DMEM with 20% FBS. The cell suspension was added to each well of a 24-well plate on an underlayer of 0.8% Noble agar in DMEM with 20% FBS at 37°C. Fresh DMEM containing 0.2% FBS with or without HGF (30 ng/ml) was added to the top of the soft agar and then cultured at 37°C for 21 days. The culture medium was changed twice per week with or without HGF. The colonies were stained with a 0.05% crystal violet solution (dissolved in 20% methanol) and the stained colonies were counted using the Metamorph NX image software (Molecular Devices, Sunnyvale, CA, USA). Each experiment was performed in triplicate.
3
Imaging Cell Migration from Wound Edge
4
Time-lapse Imaging of Cell Migration
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Cell migration was monitored with an inverted microscope (Carl Zeiss, Oberkochen, Germany) equipped with an XY motorized stage, time-lapse imaging software (Metamorph NX; Molecular Device, Sunnyvale, USA), and a Carl Zeiss incubation system. The microscope system was able to capture images of multiple locations. For recording cell sheets’ collective migration, 9 to 12 regions were recorded for each cell sheet allowing about 20% percent overlap for each region. Images were then stitched together using ImageJ software from the National Institutes of Health. All experiments were conducted using 10× phase contrast objectives. Images are taken at 5 min intervals.
5
Quantifying Liver Lipid Content
6
Automated Wound Healing Analysis
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Wound healing assays were performed as previously described [6 (link)]. Briefly, the monolayer was scratched with a sterile pipette tip (1000 μl), followed by extensive washing to remove cellular debris. The remaining cells were treated with LTP and incubated at 37 °C for 24 h. The wound on the captured image was automatically recognized and evaluated via Metamorph® NX image software (Molecular Devices, Sunnyvale, CA, USA), and the eluate of crystal violet (cat) staining was examined using light microscopy (EVOS FL Auto).
7
Non-Thermal Plasma Wound Healing Assay
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For wound healing assasy, inducible MSKQLL1 and SCC1483 cells were plated on 12-well plates and grown to confluency (>90%), followed by serum starvation for 24 h. Wounds were generated by using a sterile 200-mL pipette tip and washing with PBS. The cells were then exposed to various gases (N2, Ar, and He) of NTP for 3 min. Each experiment was performed in triplicate. The wound on the captured image was automatically recognized and measured by Metamorph® NX image software (Molecular Devices, Sunnyvale, CA, USA) and eluate of crystal violet staining was measured under a light microscopy.
8
Cytosolic dsDNA and DNA-RNA Hybrid Staining
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Cytosolic dsDNA and DNA-RNA staining were performed as described previously (16). Briefly, 50% formamide (VWR International, USA) diluted in PBS was added for 10 mins at room temperature, followed by incubation for 15 mins at 75°C to denature dsDNA. Cells were washed with PBS and incubated with blocking buffer (1% BSA (Sigma Aldrich), 2% goat serum (Hyclone) in PBS) for 1 h to prevent non-specific binding of antibodies. Cells were then stained with dsDNA (1:200, Sigma-Aldrich, USA) or S9.6 DNA-RNA hybrids (1:100, Kerafast, Boston, USA) antibodies overnight in 4°C. Cells were subsequently washed with PBST (0.1% Tween) thrice followed by anti-mouse IgG coupled with Cy3 (Millipore, Singapore) or anti-rabbit IgG coupled to Alexa Fluor 488 (Invitrogen, Singapore). Finally, DNA fluorophore DAPI (0.5 μg/ml in PBS, KPL Inc., USA) was added for 10 min. Slides were washed once in PBS before mounted with Da-/- fluorescent mounting medium (Da-/-, UK). Confocal images of staining were captured using a Zeiss Axio Imager Z1 fluorescent microscope equipped with AxioVision 4.8 software (Carl Zeiss MicroImaging, USA) or confocal TCS SP5 (Leica, Singapore). Images were analyzed using Photoshop CS4 (Adobe, USA) or ImageJ. Colocalisation of AIM2 with DNA or other proteins were quantified using Metamorph (Metamorph NX, version 8.12, Molecular Devices, USA).
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