For visualizing Pk1, F-actin, cell membrane, or ER domain, glass coverslips were mounted over tissue explants or dissociated cells and secured with plasticine. Time-lapse recordings used for tracking cell migration and neighbor exchange were taken on a Carl Zeiss Axiovert 200M fluorescent microscope with a 20× air-immersion objective (NA = 0.55) using an AxioCam MDm camera and AxioVision 4.8 software. Images used for measuring actin intensity in single cells were taken on a Carl Zeiss Axiovert 200M fluorescent microscope with a 20× air immersion objective (NA = 0.55) using AxioCam MDm camera and AxioVision 4.8 software. Images used for measuring tissue surface tension were taken on a Carl Zeiss Stemi SV 11 with a 5× air-immersion objective using AxioCam MRc camera and AxioVision 4.8 software. Images used for measuring tissue surface contact angle were taken on a Carl Zeiss Axiovert 200M inverted microscope with a 20× air immersion objective (NA = 0.55) using AxioCam MDm camera and AxioVision 4.8 software. All other images and time-lapses were taken on a Leica TCS SP8 laser scanning confocal microscope with a 40× oil-immersion objective (NA = 1.3) using LasAF 3.2 software. All images and time-lapses were taken at room temperature.
Axiocam mrc camera
Manufactured by Zeiss
Sourced in Germany, United States, Switzerland, United Kingdom, Japan
The AxioCam MRc is a digital camera designed for microscopy applications. It features a high-resolution sensor and is capable of capturing detailed images of specimens under a microscope. The core function of the AxioCam MRc is to provide high-quality image capture for a variety of microscopy techniques.
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Lab products found in correlation
Axiovision 4, by Zeiss (28 mentions)
Axiovision software, by Zeiss (13 mentions)
Axioskop 2 microscope, by Zeiss (10 mentions)
Axioscope 40 microscope, by Zeiss (10 mentions)
Triton x 100, by Merck Group (8 mentions)
Paraformaldehyde (pfa), by Merck Group (8 mentions)
Cell observer live imaging microscope, by Zeiss (8 mentions)
Axioskop 2 plus, by Zeiss (8 mentions)
Wga fitc, by Merck Group (8 mentions)
Axioskop2 mot plus microscope, by Zeiss (7 mentions)
Axiovert 200 microscope, by Zeiss (7 mentions)
Serum block, by Agilent Technologies (7 mentions)
Axio scope a1 microscope, by Zeiss (6 mentions)
Axio observer z1 microscope, by Zeiss (6 mentions)
Axiovision software 4, by Zeiss (6 mentions)
Axioplan 2 microscope, by Zeiss (6 mentions)
Axio imager m2 microscope, by Zeiss (6 mentions)
Methylcellulose, by Merck Group (6 mentions)
Axio observer z1, by Zeiss (6 mentions)
Axiovert 25 microscope, by Zeiss (5 mentions)
282 protocols using axiocam mrc camera
1
Microscopy techniques for cell imaging
2
Immunofluorescence Staining of CD163 in Macrophages
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MΦ, MS, and MR human macrophages grown in small culture dishes were fixed with 4% paraformaldehyde for 30 minutes, followed by washing with PBS thrice. Cells were blocked with 5% BSA for 30 minutes, and then incubated with the anti‐CD163 antibody (diluted 1:1000) at 4°C overnight. Signal was detected using Alexa Fluor 488 and 594‐conjugated secondary antibodies (Invitrogen), and the samples were imaged using the Axioskop 2 mot plus fluorescence microscope equipped with Plan Apochromat 203/0.8 NA and 403/0.95 NA objectives and the AxioCam MRc camera with AxioVision software 4.7.1 (Carl Zeiss).
3
Tol2-Mediated Transgenesis in Zebrafish
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Tol2 transposase mRNA was synthesized from a NotI-linearized pCS2-TP plasmid, a kind gift from Dr. Koichi Kawakami (National Institute of Genetics), using the SP6 mMessage mMachine kit (Ambion) then purified using MEGAclear cleanup kit (Ambion). Injection mixture consisted of 5 μl of 0.4 M potassium chloride (KCl), 1 μl of phenol red solution (Sigma-Aldrich), 1 μl of 250 ng/μl Tol2 expression construct, 1 μl of 250 ng/μl Tol2 transposase mRNA, and 2 μl of nuclease-free water (Ambion) to make up a total volume of 10 μl. An approximate volume of one nanoliter was injected into the cytoplasm of one-cell stage fertilized zebrafish embryos. Following injection, embryos were incubated in chorion water at 28.5°C. At 5 dpf, transgenic carriers expressing the Venus green fluorescent protein (GFP) in the eye lens were identified using a Zeiss LumarV12 stereoscope, and images were captured with a Zeiss Axiocam MRc camera. Animals were then raised to adulthood and bred to generate F1 generations with stable incorporation of the transgene.
4
Quantifying GFP Expression in C. elegans
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The images of living animals, anesthetized with 0.01% tetramisole in M9, were acquired on a Nikon Eclipse E600 microscope equipped with an AxioCam MRc camera and Zeiss AxioVision software. The images of gonad granules were acquired on an Olympus FV10i confocal microscope with a 60x objective lens, NA = 1.2, a 1024 × 1024 size, 8x quality, and 2.0x confocal aperture. The images used to compare levels of GFP expression of the Phsp-16.2::gfp reporter transgene were taken with identical exposures.
Quantification of GFP fluorescence used Image J (1.50b, Wayne Rasband, National Institutes of Health). The whole area of each animal was selected, and then its integrated density was measured. The corrected total fluorescence was calculated as follows: Corrected Total Fluorescence (CTF)= integrated density – (selected area × mean fluorescence of three background readings).
Quantification of GFP fluorescence used Image J (1.50b, Wayne Rasband, National Institutes of Health). The whole area of each animal was selected, and then its integrated density was measured. The corrected total fluorescence was calculated as follows: Corrected Total Fluorescence (CTF)
5
Wound Healing Assay for Cell Migration
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To study the two-dimensional migration, cells were plated at a density of 2.0 × 105 cells/well in 12-well plates and incubated at 37 °C for 24 h. Then, a straight wound was made into individual wells with a 10 µL pipette tip. This point was considered the “zero area” and was imaged using an AxioVert 25 microscope with an Axio Cam MRC camera attached (Zeiss, Oberkochen, Germany).
After obtaining the wounds, the control wells received 1 mL of medium with 1% FBS containing the different treatments (DOX, SIM, and the combination of DOX:SIM at ratios of 1:1; 1:2, or 2:1, respectively, in free or encapsulated form). The drug concentration used for treatment was 80 nM. This represents the total concentration of DOX or SIM alone or a combination of DOX: SIM at different molar ratios in the free or co-encapsulated forms. After 24 h of incubation at 37 °C, cells were fixed with 4% formaldehyde for 10 min. Images along the treated wounds were also obtained in phase contrast. The areas of all wounds were obtained using the MRI Wound Healing Tool plugin for the free version of the Image J 1.45 software (National Institutes of Health, Bethesda, CA, USA). The wound healing percentage was calculated according to the following equation:
After obtaining the wounds, the control wells received 1 mL of medium with 1% FBS containing the different treatments (DOX, SIM, and the combination of DOX:SIM at ratios of 1:1; 1:2, or 2:1, respectively, in free or encapsulated form). The drug concentration used for treatment was 80 nM. This represents the total concentration of DOX or SIM alone or a combination of DOX: SIM at different molar ratios in the free or co-encapsulated forms. After 24 h of incubation at 37 °C, cells were fixed with 4% formaldehyde for 10 min. Images along the treated wounds were also obtained in phase contrast. The areas of all wounds were obtained using the MRI Wound Healing Tool plugin for the free version of the Image J 1.45 software (National Institutes of Health, Bethesda, CA, USA). The wound healing percentage was calculated according to the following equation:
6
Verification of Equine Dermal Fibroblasts
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Equine dermal fibroblasts (PriFi1 and PriFi2) were verified by indirect immunofluorescence staining applying a modified reported protocol [58 ], except for the secondary antibody and antibody-dilutions. Briefly, a monoclonal mouse anti-vimentin antibody (Clone V-9, Sigma-Aldrich, dilution 1:200) was used. Samples incubated with a monoclonal mouse anti-cytokeratin antibody (C-11, Invitrogen, Rockford, US, dilution 1:100) and those incubated without primary antibody served as negative controls. F(ab’)2 goat anti-mouse IgG-FITC antibody (Bio-Rad Laboratories GmbH, Munich, Germany, dilution 1:200) was used for the visualization of the signals. Cells were evaluated and photographed at 546 nm and a 20 fold magnification with a Leica fluorescence microscope (Leica Microsystems, Wetzlar, Germany) and an AxioCam MRc camera (Zeiss Microscopy GmbH, Jena, Germany).
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Mitochondrial Content Quantification
8
Mouse Brain Growth Measurements
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All measurements of brain growth were conducted as previously described (Chakrabarti et al., 2007 (link); Goodliffe et al., 2016 (link)). Briefly, embryos were imaged using an Olympus MVX10 brightfield microscope coupled with a Zeiss AxioCam MRc camera. Somatic and gross brain measurements were determined using Axiovision software (Zeiss). All embryo crown-rump lengths were measured from the top of the head to the base of the tail. For gross brain measurements, brains were removed and cleared of all other tissue and the maximal rostrocaudal and mediolateral lengths of each telencephalon were measured.
The number of mice per group were as follows: (1) Ts1Cje – 11 euploid, 13 trisomic mice; (2) T65Dn – 20 euploid, seven trisomic mice; (3) Dp(16)1/Yey – 19 euploid, 26 trisomic mice; (4) Ts1Cje B6C3Sn hybrid background – 18 euploid, six trisomic mice; (5) Dp(16)1/Yey B6C3Sn hybrid background – nine euploid, seven trisomic mice.
The number of mice per group were as follows: (1) Ts1Cje – 11 euploid, 13 trisomic mice; (2) T65Dn – 20 euploid, seven trisomic mice; (3) Dp(16)1/Yey – 19 euploid, 26 trisomic mice; (4) Ts1Cje B6C3Sn hybrid background – 18 euploid, six trisomic mice; (5) Dp(16)1/Yey B6C3Sn hybrid background – nine euploid, seven trisomic mice.
9
Quantifying Skin Inflammation in K14-VEGFC Mice
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Chronic dorsal skin inflammation was induced in 6-8 week old hemizygous K14-VEGFC [20 (link)] and littermate WT mice with TPA application (5 μg dissolved in acetone 1x/week for 5 weeks). Experimentation was undertaken in accordance with protocols approved by the Kantonales Veterinäramt Zürich. Tissue for analysis was harvested two days after the final TPA treatment. Protein was isolated from back skin of known weight as described [5 (link)]. VEGF-C and TGF-β1 protein was quantified using VEGF-C and TGF-β1 ELISA Kits (R&D Systems), and was normalised to tissue weight and total protein amount within the lysate. Immunofluorescence stainings were performed using standard techniques as described in the Supplementary Methods . Stained sections were examined on an Axioscope Mot Plus microscope (Carl Zeiss) equipped with an Axiocam MRc camera (Carl Zeiss). Images were acquired using Axio-Vision software Version 4.7.1 (Carl Zeiss). ImageJ was used for image analysis. To quantify immune cell infiltration and the area covered by lymphatic vessels in the skin of mice, ten images/skin section/mouse at a 20x magnification were taken. Foxp3+CD4+ cells were quantified by manually counting double positive cells in skin sections from three control and four inflamed mice for each genotype.
10
Measuring Tissue Surface Tension
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In homotypic and heterotypic cell pairs and clusters in MBS, contact angle components θA and θB were measured with the ImageJ angle tool and used for calculating relative cortical tensions. To calculate absolute cortical tensions, a group of five to seven PCM explants were fused into an aggregate and allow to round up for an hour in MBS and assume a drop shape on plasticine for a further 2 h. Aggregates were fixed with 4% formaldehyde before obtaining aggregate profiles by imaging aggregates from the side in a 45° mirror using a stereomicroscope and AxioCam MRC camera (Zeiss). Tissue surface tension was determined using ADSA (Río and Neumann, 1997 (link); Luu et al., 2011 (link), 2015 (link)) to fit the aggregate profiles thus obtained to theoretical drop-shape curves. The drop shape of a tissue aggregate represents an equilibrium state of balanced tissue surface tension σ and gravity, which rounds up and flattens the tissue, respectively. The ADSA program generates theoretical drop shapes of different surface tensions using the Laplace equation and finds the best fit to measured aggregate outlines. After ADSA measurements, aggregates were cut in half and laid flat in glass-bottom dishes for measuring tissue surface contact angles using the Axiovision angle tool.
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