Tumor at each time point for biodistribution analysis of 131I-prohy group were cut into 5 μm frozen sections and examined using a fluorescence microscopy (Vert A1, ZEISS, Gottingen, Germany) to acquire fluorescence images as prohy can emit red fluorescence. Fluorescence microscopy parameters: Rhod, filter set 20, excitation BP 546/12, beam splitter FT 560, emission BP 575–640. Afterwards, these sections were stained with H&E. The relative fluorescence density of the necrotic region and the normal region of tumor, which can be distinguished based on the histological examination of H&E stain, was calculated with ZEN software (Vert A1, ZEISS, Gottingen, Germany) by drawing the regions of interest.
Vert a1
Manufactured by Zeiss
Sourced in Germany, United States
The Vert. A1 is a high-precision laboratory microscope designed by Zeiss. It features a vertical optical system, providing a stable and ergonomic viewing experience. The Vert. A1 is engineered to deliver reliable performance and accurate results in various scientific and research applications.
Automatically generated - may contain errors
Lab products found in correlation
Mito tracker red cmxros, by Beyotime (4 mentions)
Jc 1 assay kit, by Beyotime (4 mentions)
Mitotracker red cmxros, by Zeiss (3 mentions)
S0033, by Beyotime (3 mentions)
Reactive oxygen species assay kit, by Beyotime (2 mentions)
Goat anti rabbit or anti mouse igg, by Abbkine (2 mentions)
Col 2, by Novus Biologicals (2 mentions)
Celltiter glo 3d cell viability assay, by Promega (2 mentions)
Basic fibroblast growth factor fgfb, by Thermo Fisher Scientific (2 mentions)
Poly hema, by Kasvi (2 mentions)
Facscanto 2, by BD (1 mentions)
Tunel assay kit, by Elabscience (1 mentions)
Alexa flour 594, by Thermo Fisher Scientific (1 mentions)
Hoechst 33342, by Beyotime (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Beyotime (1 mentions)
Phalloidin ifluor 555, by Abcam (1 mentions)
Osteo assay surface 96 well multiple well plate, by Corning (1 mentions)
Tmr red tunel cell apoptosis detection kit, by Wuhan Servicebio Technology (1 mentions)
48 well cell culture plate, by Corning (1 mentions)
Lsm 880, by Zeiss (1 mentions)
46 protocols using vert a1
1
Fluorescence Microscopy Analysis of Tumor Biodistribution
2
Epithelial Cell Infection by PAO1 Biofilm
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CFBE41o- or Calu-3 cells were used for the infection experiment, respectively, after 7 or 12 days of seeding. The cultures at ALI condition were infected with a preformed biofilm of PAO1 (CFU/ml of biofilm ~ 4.6 × 108). The biofilms (pre-grown for 24 h in a 24-well plate) were gently scratched off and transferred with a pipette to the apical side of the epithelial cells. Biofilms were allowed to attach to cell monolayers for 1 h. After that time, unattached bacteria and excess medium were removed from the apical side, and the basolateral medium changed to MEM, which was supplemented with 0.4% arginine. TEER measurement was assessed as described before. Biofilm formation, epithelial cell viability, and bacteria survival were further determined after 4 and 24 h infection, as described below. During the experiment, the epithelial cell monolayers’ integrity was checked with a light microscope (ZEISS, Vert.A1).
3
Intracellular ROS Assessment of CdTe QDs
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KUP5 cells (5 × 104) were cultured in 2 mL in wells of a 6-well plate and incubated with 0, 5, 50, and 500 nM CdTe QDs for 24 h. Intracellular ROS levels were measured using fluorescence microscopy (VERT.A1, ZEISS AG, Germany) and flow cytometry (FACSCANTO II, BD Bioscience) using the Reactive Oxygen Species Assay Kit (Beyotime Technology, Shanghai). In brief, KUP5 cells exposed to CdTe QDs for 24 h were washed with PBS twice, then diluted with serum-free medium and DCFH-DA at a ratio of 1:2000 and incubated at 37 °C for 30 min. Finally, the fluorescence signal could be photographed after cleaning with PBS. The laser axis positioned at an angle of about 90° with a laser beam is called side scattered light (SSC), and the size of SSC is proportional to the complexity of the particle composition of the cell. The fluorescence values of the SSC and DCF in different dose groups were normalized to evaluate the production of intracellular ROS and the uptake of intracellular particulate matter. The excitation wavelength and emission wavelengths were 488 nm and 525 nm, respectively. The data was evaluated using FlowJo, version 10.4.
4
Mitochondrial Activity Detection with MitoTracker Red
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MitoTracker Red CMXRos (C1049B, Beyotime) was used to detect mitochondrial activity. According to the manufacturer’s instructions, the cells were incubated with a culture medium containing 20 nM MitoTracker Red CMXRos for 30 min at 37 ℃ in the dark and then observed and captured with a fluorescence microscope (ZEISS Vert. A1) after changing the fresh culture medium.
5
Immunofluorescence Analysis of ATF6 and Col-2
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The cells were treated as indicated, and after 48 h, the cells were fixed with 4% paraformaldehyde for 20 min. After being permeabilized with 0.2% TritonX-100 for 20 min, the samples were blocked by BSA at 37 ℃ for 1 h. Then, the cells were incubated with antibodies against ATF6 (1:100, Proteintech) and Col-2 (1:500, Novus) at 4 ℃ overnight. The next day, the cells were incubated with fuorescently labelled goat anti-rabbit or anti-mouse IgG (1:100, Abbkine) for 1 h at 37 ℃. The nuclei were stained with DAPI. The images were taken using a fuorescence microscope (ZEISS Vert. A1) and analysed with the ImageJ software.
6
Mitochondrial Membrane Potential Assay
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The mitochondrial membrane potential changes were detected with a JC-1 assay kit (C2006, Beyotime). Based on the manufacturer’s instructions, the cells were stained with the JC-1 staining solution at 37 ℃ for 20 min free from light. Then, the cells were washed twice with JC-1 staining buffer, and the images were observed and captured using a fuorescence microscope (ZEISS Vert. A1).
7
Mitochondrial Activity Monitoring
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MitoTracker Red CMXRos (C1049B, Beyotime Biotechnology) was used to detect mitochondrial activity and membrane potential. According to the manufacturer’s instructions, the cells were incubated with a culture medium containing 20 nM MitoTracker Red CMXRos for 30 min at 37°C in the dark and then observed and captured with a fluorescence microscope (ZEISS Vert. A1) after changing the fresh culture medium.
8
Quantifying Cellular Apoptosis via TUNEL
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Human NP cells on coverslips were stained with the TUNEL assay Kit (E-CK-A331, Elabscience) to examine the cellular apoptosis of each experimental group. All procedures were performed in accordance with the manufacturer’s instructions. The images were captured using a fluorescence microscope (ZEISS Vert. A1).
9
Immunofluorescence Analysis of Tumor Spheroids
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Tumor spheroids were placed in 4% paraformaldehyde and then processed with Triton X-100 for 20 min and incubated respectively with 5% bovine serum albumin and anti-PFKFB3 (ABclonal, A3934) and anti-p-AMPK (ABclonal, AP1002) antibody overnight. Next day, the spheroids were rinsed thrice and cultivated in secondary antibodies (Invitrogen, Alexa flour-594, Invitrogen) for 2 h at indoor temperature. The cell nucleus was used by using Hoechst33342 (Beyotime, 1810898). Images were acquired using a confocal microscope (Zeiss, Vert.A1).
10
Immunofluorescence Analysis of ATF6 and Col-2
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The cells were treated as indicated, and after 48 h, the cells were fixed with 4% paraformaldehyde for 20 min. After being permeabilized with 0.2% TritonX-100 for 20 min, the samples were blocked by BSA at 37 ℃ for 1 h. Then, the cells were incubated with antibodies against ATF6 (1:100, Proteintech) and Col-2 (1:500, Novus) at 4 ℃ overnight. The next day, the cells were incubated with fuorescently labelled goat anti-rabbit or anti-mouse IgG (1:100, Abbkine) for 1 h at 37 ℃. The nuclei were stained with DAPI. The images were taken using a fuorescence microscope (ZEISS Vert. A1) and analysed with the ImageJ software.
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