A549/A549LD and H460/H460LD cells or p120ctn siRNA-transfected cells were grown in chamber-slide glass and incubated for 24 h. The cells were then fixed in 4% formaldehyde at room temperature. After being rinsed in PBS, the cells were incubated overnight with anti-Rac, anti-Rho, or anti-p120ctn, or anti-tubulin at 4 °C. Samples were then incubated with the corresponding Alexa Fluor-488 or -546-conjugated secondary antibodies (Invitrogen-Molecular Probes or Life Technology) for 1 h followed by incubation with Alexa Fluor 555 phalloidin (Invitrogen) or 4′,6-diamidino-2-phenylindole (DAPI, Invitrogen, 1:1000) for 30 min. Samples were visualized using a ZEISS LSM 510 META/Confocor2 microscope (Carl Zeiss MicroImaging) with the pinhole set at one Airy unit using 488, 546, and/or 633 nm excitation and a 63/1.4 oil objective lens. The fluorescence intensity of target proteins was quantified by MetaMorph Microscopy Automation and Image Analysis Software (Molecular Devices, Sunnyvale, CA, USA). Fluorescence density (FD) was defined as each cell fluorescence intensity divided by each cell area (fluorescence intensity/µm2). The mean FD of 50–80 different cells in 5 ~ 8 fields at 630× was used to represent the immunofluorescence of target proteins. The proportion of inner-region lumican was determined by comparing the intensity in the inner region to the total intensity in each cell.
Metamorph microscopy automation and image analysis software
Manufactured by Molecular Devices
Sourced in United States, United Kingdom, Japan
MetaMorph Microscopy Automation and Image Analysis Software is a comprehensive software solution for controlling microscopes and analyzing microscopic images. It provides tools for automated image acquisition, multi-dimensional data management, and advanced image processing and analysis.
Automatically generated - may contain errors
Lab products found in correlation
Ti u microscope, by Nikon (3 mentions)
Csu x1 m1 confocal head, by Oxford Instruments (3 mentions)
Du 885 camera, by Oxford Instruments (3 mentions)
Prism 5, by GraphPad (3 mentions)
Coolsnap hq2, by Molecular Devices (2 mentions)
Axio observer d1, by Zeiss (2 mentions)
Deltavision microscope, by Cytiva (2 mentions)
Ix83 microscope, by Olympus (2 mentions)
Tm 1000, by Hitachi (2 mentions)
Trans retinoic acid, by Merck Group (2 mentions)
Axiovert 200m fluorescence microscope, by Zeiss (2 mentions)
Alexa fluor 488 or 546 conjugated secondary antibody, by Thermo Fisher Scientific (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (1 mentions)
Alexa fluor 555 phalloidin, by Thermo Fisher Scientific (1 mentions)
Lsm 510 meta confocor2 microscope, by Zeiss (1 mentions)
Axio imager z2 microscope, by Zeiss (1 mentions)
Axiocam 503, by Zeiss (1 mentions)
Dm4000 epifluorescence microscope, by Leica (1 mentions)
Poly l lysine (pll), by Merck Group (1 mentions)
Hoechst dye, by Thermo Fisher Scientific (1 mentions)
57 protocols using metamorph microscopy automation and image analysis software
1
Quantifying Cellular Protein Localization
2
Quantifying Cell Morphology and Protein Expression
3
Visualizing Embryonic Cell Dynamics
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Chorionated embryos that had been injected with the tbx16 cMO and/or photoactivatable tracers and spot-irradiated at 6 hpf were placed in an agarose template (560-μm × 960-μm wells) filled with E3 medium, oriented with the dorsal shield facing upward. Imaging was performed with a Leica DMI6000B inverted compound microscope equipped with a UVI 6.3x/0.13 objective, GFP and TX2 filters, and a CoolSnap HQ2 monochrome camera controlled by MetaMorph microscopy automation and image analysis software (Molecular Devices). Fluorescent micrographs were acquired at rate of 1 frame per 5 minutes, with exposure times automatically adjusted every five frames to account for progressive photobleaching. ImageJ software (NIH) was used to normalize signal brightness and generate annotated movie montages.
4
Visualizing Embryonic Cell Dynamics
5
Live Imaging of C. elegans Larvae
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Live imaging of C. elegans larvae was done by mounting larvae on 5% agarose pads in a 10 mM Tetramisole solution in M9 buffer to induce paralysis. DIC imaging was performed with an upright Zeiss AxioImager Z2 microscope using a 63 × 1.4 NA objective and a Zeiss AxioCam 503 monochrome camera, driven by Zeiss Zen software. Spinning disk confocal imaging was performed using a Nikon Ti-U microscope driven by MetaMorph Microscopy Automation and Image Analysis Software (Molecular Devices) and equipped with a Yokogawa CSU-X1-M1 confocal head and an Andor iXon DU-885 camera, using 60 × or 100 × 1.4 NA objectives. All stacks along the z-axis were obtained at 0.25 μm intervals, and all images were analyzed and processed using ImageJ(FIJI) and Adobe Photoshop. For quantifications, the same laser power and exposure times were used within experiments.
6
Humoral Immunity Impact on HCMV Infection
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
To measure the impact of humoral immunity on HCMV infection, all viruses were preincubated with pooled sera from seropositive or seronegative donors for 1 h at dilutions from 1:5 to 1:800. After 1 h, the cells were infected with these HCMV preparations and fixed and stained 24 hpi for IE gene expression as a marker of lytic infection.
For studies with IFN, cells were pretreated with a combination of IFN-α/β (Peprotech) for 3 h and then infected with HCMV at an MOI of 1. After 16 h, the cells were fixed and stained for IE gene expression as a marker of lytic infection.
Cells stained for viral IE protein expression were quantified by automated fluorescence microscopy and image recognition. A Hermes WiScan (IDEA Bio-Medical) automated microscope was used to image the central 25% of each well (20 images/well), and the resultant images were processed with MetaMorph microscopy automation and image analysis software (Molecular Devices).
For studies with IFN, cells were pretreated with a combination of IFN-α/β (Peprotech) for 3 h and then infected with HCMV at an MOI of 1. After 16 h, the cells were fixed and stained for IE gene expression as a marker of lytic infection.
Cells stained for viral IE protein expression were quantified by automated fluorescence microscopy and image recognition. A Hermes WiScan (IDEA Bio-Medical) automated microscope was used to image the central 25% of each well (20 images/well), and the resultant images were processed with MetaMorph microscopy automation and image analysis software (Molecular Devices).
7
Quantitative Fluorescent Imaging of MBP+ Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Ten images at 10x magnification were captured at 485 nm (FITC channel, MBP-positive cells) and 358 nm (DAPI channel, nuclei), at identical locations on each stained cover glass, with a Leica DM4000 epifluorescence microscope (Leica Microsystems UK Ltd., Milton Keynes, UK) interfaced with the Metamorph® microscopy automation and image analysis software (Molecular Devices, LLC, San Jose, CA, USA). Cell quantification was performed with the ImageJ software (Rasband, W.S., NIH, Bethesda, Maryland) using scripts developed by Prof. John V. Priestley. The data obtained from each field of view were the number of nuclei and MBP-positive cells bearing membranous sheaths (which was computed as a percentage of the total number of cells per field of view and then normalized to and expressed as a relative percentage of the fields of view-averaged value of the vehicle-treated control wells), and the area covered by the cells bearing sheaths.
8
Analysis of Mitotic Defects in Norad-Deficient MEFs
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Primary Norad+/+ and Norad–/– MEFs (three MEF lines per genotype, P<4) were grown on Lab-Tek Chambered Coverglass slides (Thermo Fisher) that were coated with poly-L-lysine (Sigma-Aldrich). Prior to the analysis, DNA was visualized by adding 50 ng/mL Hoechst dye (Invitrogen) to the growth medium. Mitoses were monitored by taking fluorescence images every 5 min for ~48 hr on a Leica inverted microscope equipped with a temperature and CO2-controlled chamber, a 63X oil objective, an Evolve 512 Delta EMCCD camera, and the MetaMorph Microscopy Automation and Image Analysis Software (Molecular Devices, LLC). Videos were generated from the acquired time-lapse images and analyzed for the occurrence of mitotic defects including anaphase bridges and lagging chromosomes.
9
Intracellular cAMP Dynamics Measurement by FRET
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
To evaluate intracellular cAMP levels, fluorescence resonance energy transfer (FRET) experiments were performed. Briefly, HCT8 cells were seeded onto 12 mm diameter glass coverslips at 37°C, 5% CO2, and transiently transfected with a plasmid encoding the H96 sensor (Klarenbeek et al., 2011 (link)) (a gift from Dr. K. Jalink) containing the cAMP-binding sequence of Epac1 sandwiched between ECFP (donor) and EYFP (acceptor). In detail, cells were transfected with 0.5 μg of DNA/cm2 using TransFectin™ Lipid Reagent (1.5 μL/cm2) according to the protocol provided by the manufacturer (Bio-Rad Laboratories, Inc., Hercules, CA, United States) and left under basal condition or treated as mentioned before. Experiments were performed 48 h after transfection. Visualization of ECFP- and/or EYFP-expressing cells and detection of FRET was performed on an inverted microscope (Nikon Eclipse TE2000-S) controlled by Metamorph® Microscopy Automation and Image Analysis Software (Molecular Devices, LLC, San Jose, CA, United States). Each image was corrected and analyzed as previously shown (Di Mise et al., 2019 (link)).
10
Modeling Huntington's Disease in SH-SY5Y Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
SH-SY5Y TBP/Q36~79-GFP cells were seeded in six-well (2×105/well) plates in medium containing all-trans retinoic acid (10 μM; Sigma). The next day, doxycycline (5 μg/mL) was added to induce TBP/Q36~79-GFP expression for 7~21 days. After that, cells were stained with Hoechst 33342 (0.1 μg/mL; Sigma), and aggregation percentage was assessed by a high content analysis system at 482 (excitation)/536 (emission) wavelengths. The morphologic differentiation of TBP/Q36~79 cells, including processes and branches, was assessed by using Metamorph microscopy automation and image analysis software (Molecular Devices). For testing the aggregation reduction potential of GGA, indole, and NC001-8, cells were pretreated with GGA, indole, or NC001-8 (0.1 μM) for 8 hours before inducing TBP/Q36~79-GFP expression for 7 days.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!