HEK293T cells were seeded in a 24-well plate (Greiner) and transfected for 24 h ~ 48 h with pAcBac1-haloTyrRS and pAcBac1-sfGFP-150TAG plasmid DNA in the presence of 0.1 mM chloroTyr, 0.1 mM bromoTyr, or vehicle using Lipofectamine 2000 (Thermo Fisher Scientific) according to the manufacturer’s protocol. PON1 expression was carried out in the presence of 1 mM chloroTyr to enhance the suppression efficiency. Fluorescence and phase contrast images were captured using the EVOS FL imaging system (Thermo Fisher) or the Keyence BZ-X710 all-in-one fluorescence microscope (Keyence Corporation, IL)
Bz x710 all in one fluorescence microscope
Manufactured by Keyence
Sourced in Japan, United States
The BZ-X710 All-in-One fluorescence microscope is a comprehensive imaging system that combines advanced optical and digital technologies. It is designed to capture high-quality fluorescence images for a wide range of applications in life science research and analysis.
Automatically generated - may contain errors
Lab products found in correlation
Lipi red, by Dojindo Laboratories (4 mentions)
Bz x viewer, by Keyence (4 mentions)
Bz x analyzer, by Keyence (4 mentions)
Fv3000 confocal laser scanning microscope, by Olympus (3 mentions)
Mitoros, by AAT Bioquest (3 mentions)
Alexa fluor 488 donkey anti rabbit igg, by Thermo Fisher Scientific (3 mentions)
Cfi plan fluor 20x, by Nikon (3 mentions)
Mitogreen, by PromoCell (3 mentions)
Bz x analyzer software, by Keyence (3 mentions)
Fluorescent mounting medium, by Agilent Technologies (3 mentions)
4 6 diamidino 2 phenylindole (dapi), by Dojindo Laboratories (3 mentions)
Vectashield mounting medium with dapi, by Vector Laboratories (3 mentions)
Opal 3 plex kit, by PerkinElmer (3 mentions)
Bz x viewer v1, by Keyence (2 mentions)
Fibronectin, by Merck Group (2 mentions)
Blocking one, by Nacalai Tesque (2 mentions)
Sybr green 1, by Thermo Fisher Scientific (2 mentions)
Propidium iodide, by Merck Group (2 mentions)
Prolong gold with dapi, by Thermo Fisher Scientific (2 mentions)
Duolink in situ mouse rabbit kit, by Merck Group (2 mentions)
170 protocols using bz x710 all in one fluorescence microscope
1
Genetic Incorporation of Noncanonical Amino Acids
2
Endogenous Localization of ADAM10 and CX3CL1 in Cardiac Tissue
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To analyze the endogenous localization of ADAM10 (1:100, AB19026, Millipore) and CX3CL1 (1:100, ab25088, Abcam), heart tissue sections (mid-ventricular short axis sections, 4% PFA fixed, paraffin embedded, 3 µm) and indicated cells were stained with specific antibodies and counterstained with CD31 (1:100, 553370, BD Biosciences), CD45 (1:100, 555480, BD Biosciences), cTnI (1:100, MAB3150, Millipore), cTnT (1:100, ab45932, Abcam) or Vimentin (1:100, 550513, BD Biosciences) where indicated. Alexa Fluor 488-coupled goat anti-rabbit (1:200, A27034, ThermoFisher Scientific), Alexa Fluor 546-couppled goat anti-mouse (1:200, A-11003, ThermoFisher Scientific) or CF 594-coupled goat anti-rat (1:200, SAB4600323, Sigma-Aldrich) and ProLong Diamant Antifade Mountant with DAPI (P36962, ThermoFisher Scientific) were used as secondary antibodies and for mounting, respectively. Image acquisition was performed using a Keyence BZ-X710 All-in-One Fluorescence Microscope (Keyence) with the BZ-X Viewer v1.03.00.05 software (Keyence). The cellular distribution of ADAM10 and CX3CL1 was assessed using CellProfiler 446 . Cardiac-TnT, CD31, CD45 and Vimentin positive cells were analyzed for ADAM10 expression. For each mouse at least 1000 cells/staining were analyzed. Cardiomyocyte cross-sectional diameter was determined using Fiji. One hundred cells/mouse were analyzed.
3
Histological Analysis of Myocardial Infarction
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For histological analysis, mid-ventricular short axis sections were fixed in 4% PFA for 24 h before dehydration using an ethanol gradient. Subsequently the samples were embedded in paraffin, sectioned at 3 μm thickness and stained with sirius red (Sigma-Aldrich). Image acquisition was performed using the Keyence BZ-X710 All-in-One Fluorescence Microscope (Keyence) with the BZ-X Viewer v1.03.00.05 software (Keyence). Using Fiji, the epicardial and endocardial infarct length and circumference was measured. Infarct size was calculated using the following equation: [(epicardial infarct length/epicardial circumference) + (endocardial infarct length/endocardial circumference)/2] * 10047 (link).
4
Chemoattractant Evaluation for Cell Migration
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To generate chemoattracting media for in vitro migration assays, HL-1 cells were plated in 0.5% fibronectin, 0.02% gelatin (all Sigma-Aldrich) coated 6-well plates. After 4 d of incubation at 37 °C in a humidified atmosphere containing 5% CO2, cells were treated with eighter DMSO or GI254023X for 24 h. Subsequently, cells were washed with fresh medium and subjected to 3 h of normoxic and hypoxic (1% O2) conditioning. Cells were lysed for Western Blot analysis and supernatants were used as chemoattractant in the lower chamber of 24-well Transwell plates with 8-µm pore size (ThermoFisher Scientific). Following isolation (neutrophils) or differentiation (macrophages), bone marrow derived cells were suspended in RPMI without FBS, plated in each upper chamber of the Transwell plates and incubated for 12 h at 37 °C in a humidified atmosphere containing 5% CO2. Following incubation, cells were fixed with 4% PFA for 15 min, DAPI stained and mounted on microscopy slides using ProLong Diamant Antifade Mountant (ThermoFisher Scientific). Image acquisition was performed using the Keyence BZ-X710 All-in-One Fluorescence Microscope (Keyence). Cell counts per field were analyzed using CellProfiler 4.046 .
5
Comprehensive post-exposure evaluation of mice
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After the exposure period and SHIRPA evaluation, mice were anesthetized with 0.5–2% isoflurane (Patterson Veterinary, Greeley, CO, USA) via face mask in oxygen. Pre-sacrifice blood was withdrawn in a subset of animals (n = 5) via terminal left ventricular puncture for evaluation of serologic markers of renal and hepatic injury, coagulopathy, as well as arterial blood gas analysis. In all animals, all major organs were then removed, formalin-fixed, stained by hematoxylin and eosin, and evaluated by light microscopy (Keyence BZ-X710 All-in-One Fluorescence Microscope, Keyence, Itasca, IL, USA) by a pathologist blinded to treatment allocation. In order to examine for any subtle damage (i.e., invisible to light microscopy) to the airways, samples of the mid-trachea were fixed, embedded, sectioned (80 nm-thick), and examined by electron microscopy (Tecnai G2 Spirit BioTWIN Electron Microscope, FEI Company, Hillsboro, OR, USA) by a pathologist blinded to treatment allocation in a subset of animals (n = 5 per group).
6
Histopathological Analysis of Mouse Lung Tissue
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For histological analysis, mouse lung tissue was fixed in 4% paraformaldehyde overnight. Subsequent paraffin embedding, sectioning (5 µm) and staining (hematoxylin/eosin and picrosirius red) was performed by the histology facility at the Center for Molecular and Cellular Bioengineering (CMCB) Dresden. For immunohistochemical staining deparaffinization in serial solutions of xylene, ethanol and water were followed by antigen retrieval via steaming in 10 mM citric acid (pH 6.0) for 20 min and a subsequent cooling period of 20 min at room temperature. Samples were washed twice with PBS for 10 min and subsequently blocked with 10% FCS for 60 min. Following, samples were incubated with anti-α-SMA antibody overnight at 4 °C in a humidified chamber. After washing twice with PBS, the samples were incubated with the secondary antibody (Alexa fluor 546; Thermo Fisher Scientific, Waltham, MA USA) and DAPI for 60 min. Fluorescence and brightfield images (for picrosirius red) of randomly chosen areas were acquired with a Keyence BZ-X710 All-in-One Fluorescence Microscope (Keyence Corporation of America, Itasca, IL USA). Fibrosis and fluorescence intensity were quantified using FIJI 1.52n software (Wayne Rasband, National Institutes of Health, Bethesda, MD USA).
7
Fluorescence Microscopy Imaging Protocol
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Images of H&E, IF and IHC samples were taken with the Keyence BZ-X710 All-in-One fluorescence microscope (Keyence, Osaka, Japan). Standard filters were used to image DAPI, Alexa Fluor 488 and Alexa Fluor 594. Images were taken using the 10×, 20×, and 40× objectives. Images were captured using Keyence BZ-X Viewer version 01.03 software.
8
Quantifying csrA Promoter Activity
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PCR fragments containing the csrA promoter were amplified using pcsrA aaaagatctCTGATTGCAGGCGTATCTAAGG and pcsrAR aaatctagaAAAGATTAAAAGAGTCGGGTCTCTCTGTATCC primer pair from both E. coli K12 and C strains and cloned into the BglII/XbaI site of the pAG136 plasmid [55 (link)] or the SmaI site of the pPROBE-GFP [LVA] promoter probe vector [56 (link)]. All constructs were verified by DNA sequencing. Plasmids were introduced into both the E. coli K12 and C strains by a TSS transformation [66 (link)]. GFP activity (OD480–520) was measured using BioTek Synergy HT (BioTek) or Tecan InfiniteM200 Pro (Tecan) plate readers and normalized to the optical density of the culture (OD600), yielding relative fluorescence units (RFU; FL480–520/OD600). For quantification of promoter activities in late stationary phase, single colonies were inoculated into 5 mL of LB broth, vortexed and 5 μL of cell suspension was spotted on LB Miller agar plates. Plates were incubated at 30 °C or 37 °C. At the specific time points, plates were scanned with a Typhoon 9400 Variable Mode Imager using 532/526-nm excitation/emission wavelengths (GE Healthcare). Scans were analyzed using the ImageQuant TL software (GE Healthcare). Student t-test was used to compare results and check statistical significance. Fluorescence microscopy was done with a Keyence BZ-X710 All-in-One Fluorescence microscope (Keyence).
9
Sensitive Detection of Induced Pluripotent Stem Cells
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hiPSCs at a concentration of 1000 cells/mL were prepared by serial dilution and 10 cells (10 µL) or 100 cells (100 µL) of hiPSCs were added to 1 × 107 immortalized NPCs. The cells were seeded onto laminin-521-coated 100-mm dishes in mTeSR1 medium and transduced with the Ad vector at 3 pfu/cell for 24 h, followed by adding 10 nM AP1903. Twenty-four hours after treatment, one tenth of the untreated 0.001% hiPSC group and the 0.0001% hiPSC group treated with either the Ad vector or AP1903, and all of the 0.0001% hiPSC group treated with both the Ad vector and AP1903 were seeded onto laminin-521-coated 12-well plates. Four days after seeding, cells were fixed with 4% paraformaldehyde in PBS for 15 min, permeabilized with 0.1% Triton X-100 in PBS for 10 min, and then blocked with Blocking One (Nacalai Tesque) for 2 h at 15–25 °C. The fixed cells were then incubated with an anti-TRA-1-60 mouse monoclonal antibody (Merck, Darmstadt, Germany, #MAB4360; 1:100) for 1 h at 15–25 °C, followed by staining with a goat anti-mouse IgM conjugated with Alexa Fluor 488 (Thermo Fisher Scientific, #A21042; 1:100) for 1 h at 15–25 °C. The stained cells were visualized using a Keyence BZ-X710 All-in-one Fluorescence Microscope (Keyence, Osaka, Japan).
10
Histological Analysis of Skin Samples
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Skin samples were fixated in 4% paraformaldehyde overnight for histological analysis. Subsequently, the samples were embedded in paraffin, sectioned (5-µm layer thickness), and stained (hematoxylin/eosin (H&E) and picrosirius red) by the staff of the histology facility at the Center for Molecular and Cellular Bioengineering (CMCB), Dresden. The immunohistochemical staining was performed as described previously (Kant et al. 2021 (link)). Fluorescence and brightfield images (for H&E and picrosirius red) were acquired with a Keyence BZ-X710 All-in-One Fluorescence Microscope (Keyence Corporation of America, USA) and a Zeiss Axio Observer Z1 microscope (Carl Zeiss AG, Germany). To quantify tissue collagen accumulation and tissue αSMA fluorescence intensity, FIJI 1.52n software (Schindelin et al. 2012 (link)) was used to quantify stained areas in relation to the total image area.
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