Cells grown on glass coverslips were fixed with 4% paraformaldehyde/ PBS for 30 minutes, followed by permeabilization in 0.5% Triton X-100/PBS for 20 minutes, or fixed with methanol at −20°C. Paraffin-embedded breast tissue sections were cut, dewaxed, and rehydrated with xylene and graded alcohols and then underwent antigen retrieval and inactivation of endogenous peroxidase. Cells or tissues were blocked with 2% bovine serine albumin in PBS and incubated with primary antibodies and then FITC- or rhodamine-conjugated secondary antibodies, followed by staining with DAPI as described [27 (link)]. For visualization of microfilaments, cells were stained with rhodamine-conjugated phalloidin for 30 minutes. Coverslips were then mounted and examined with an Axio Observer A1 fluorescence microscope (Carl Zeiss) as described previously [28 (link)].
Axio observer a1 fluorescence microscope
Manufactured by Zeiss
Sourced in Germany
The Axio Observer A1 is a fluorescence microscope designed for routine observation and imaging of fluorescently labeled samples. It features an inverted optical design, allowing for convenient sample handling and analysis. The microscope is equipped with a range of illumination options, including LED or mercury lamp, to enable fluorescence excitation.
Automatically generated - may contain errors
Lab products found in correlation
4 6 diamidino 2 phenylindole (dapi), by Merck Group (4 mentions)
Enspire multimode plate reader, by PerkinElmer (3 mentions)
Axiocam ccd camera, by Zeiss (3 mentions)
Axiovision software 4, by Zeiss (2 mentions)
Thermopol buffer, by New England Biolabs (2 mentions)
Plan apochromat 40 na 1.4 oil objective, by Zeiss (2 mentions)
Egfp luciferase sars cov 2 spike s1 lentivirus vector, by GenScript (2 mentions)
Axio vert a1 microscope, by Zeiss (1 mentions)
Axiocam icc3 camera, by Zeiss (1 mentions)
Normal goat serum (ngs), by Vector Laboratories (1 mentions)
Tnnt2, by Abcam (1 mentions)
Alp substrate kit 3, by Vector Laboratories (1 mentions)
Nanog, by Abcam (1 mentions)
Carboxy h2dcfda, by Thermo Fisher Scientific (1 mentions)
Propidium iodide, by Thermo Fisher Scientific (1 mentions)
Macsquant analyzer 10, by Miltenyi Biotec (1 mentions)
Varioskan microplate reader, by Thermo Fisher Scientific (1 mentions)
Quartz cuvette, by Agilent Technologies (1 mentions)
Cary 50 bio uv vis spectrophotometer, by Agilent Technologies (1 mentions)
Sodium nitrite, by Merck Group (1 mentions)
27 protocols using axio observer a1 fluorescence microscope
1
Immunofluorescence Microscopy of Cells and Tissues
2
Fluorescence Microscopy Imaging Protocols
3
Isolation and Purification of Mouse Primary Hepatocytes
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Mouse primary hepatocytes were isolated from the female WT or Glt25d1+/− mice using a two-step collagenase perfusion procedure as described previously (14 (link)). The hepatocytes were purified further using 60% percoll (1.076 g/ml) (15 (link),16 (link)). The mean viability and purity of mouse primary hepatocytes following isolation were identified by trypan blue exclusion and periodic acid Schiff (PAS) staining, respectively. All the images were acquired and analyzed using an Axio Observer A1 Fluorescence Microscope (Zeiss AG). Low activity or dead hepatocytes were stained blue by trypan blue staining. Six fields (original magnification, ×100) were selected randomly and the ratio of hepatocellular viability was calculated according to the following formula: Cell viability=1-(number of blue cells/total number of cells) ×100%. The hepatocytes were subjected to PAS staining, which specifically stains the hepatocyte cytoplasm red, in order to assess their purity. Six fields (original magnification, ×100) were selected randomly and the ratio of hepatocellular purity was calculated according to the following formula: Hepatocellular purity=(number of red cells/total number of cells) ×100%.
4
Stem Cell Characterization Protocol
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Alkaline phosphatase (ALP) activity was analysed by using an ALP substrate kit III (Vector Laboratories, USA) according to the manufacturer's instructions. Immunostaining assays were performed according to the protocol described previously [24 (link)]. Briefly, cells were fixed with 4% paraformaldehyde, permeabilized in 0.3% Triton X-100, blocked in 10% normal goat serum (Vector Laboratories), and then incubated with primary antibodies against OCT4 (1 : 200, Abcam, USA), NANOG (1 : 200, Abcam, USA), and TNNT2 (1 : 500; Abcam, USA) in 4°C overnight and detected by DyLight 488- or DyLight 549-conjugated secondary antibodies. Nuclei were stained with DAPI (Sigma, USA). A Zeiss Axio Observer A1 fluorescence microscope was used for slide observing and image capture.
5
Glutamate-Induced Oxidative Stress Assay
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HT-22 cells were seeded in 5 mM dishes and allowed to adhere overnight. The cells were then treated with 5 mM glutamate with or without B.monnieri extracts (10 μg/ml) or in the case of the control 0.5% DMSO and incubated for 18 h. The cells were then washed with PBS and stained with carboxy-H2DCFDA (Invitrogen) and incubated at 37 °C/5% CO2 for 30 min. The cells were then washed three times with PBS and imaged using Axio Observer A1 fluorescence microscope (Carl Zeiss, Jena, Germany). The images were then analyzed for mean cellular fluorescence using image J and the data represented as AU.
6
Immunofluorescence Staining and Flow Cytometry
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Cells were fixed with 4% paraformaldehyde (w/v), permeabilized with Triton X-100 blocking solution and stained by standard protocols using primary antibodies, listed in Table S2 , and appropriate secondary antibodies. Incubation with primary antibodies and corresponding isotype controls was performed overnight at 4°C. Staining of living cells for extracellular markers was performed for 1 h. Secondary antibody staining was performed afterward. Cells were counterstained with DAPI (Sigma) for the analysis with an Axio Observer A1 fluorescence microscope and Axiovision software 4.71 (Zeiss) or propidium iodide (PI) (final concentration 1 μg/mL) (Thermo Fisher Scientific) for the analysis using a MACSQuant Analyzer 10 (Miltenyi Biotech). Flow cytometric data evaluation was performed with FlowJo 7.6.5 software (Celeza).
7
Cellular Uptake of GSH-CdTe QDs
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The cells were treated with GSH-CdTe QDs for 1 h. Afterward, the QDs were removed and washed with PBS for three times. The fluorescence images of cells were then observed under Axio Observer A1 fluorescence microscope (ZEISS, Germany).
8
Characterization of Fluorescent Compounds
9
Photochromic Material Characterization Protocol
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D230 was supplied by Huntsman Corporation, USA. Sodium nitrite was purchased from Sigma-Aldrich. All other chemicals were purchased from China National Pharmaceutical Group Co., Ltd. (Shanghai, China) and used without further purification. 1H NMR spectroscopy was tested on a 400 MHz Bruker Spectrometer (Bruker, Germany). Absorption of UV-Vis spectra was measured on a Cary 50-Bio UV-vis spectrophotometer in a quartz cuvette (Varian). Isomerisation was tested on an EXFO Acticure 4000 light source (working wavelength: 365 nm). The visible light intensity (12.1 mW/cm2). UV light intensity (3.8 mW/cm2). SEM was performed on a FEI Quanta 400F ESEM (FEI Co., Ltd., USA). The samples were examined at 5.0 kV accelerating voltage. The surfaces were observed using an Axio Observer A1 fluorescence microscope (Carl Zeiss Inc., Germany). Rheology was conducted on a rheometer with a temperature controller to maintain the temperature (TA Instruments, USA).
10
Immunofluorescence Staining Protocol
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Cells were fixed with 4% paraformaldehyde (w/v) and stained by standard protocols using primary antibodies, as listed in Table S2 , and appropriate secondary antibodies (DyLight 488_donkey_anti-mouse_IgG and IgM, DyLight 488_donkey_anti-goat_IgG, DyLight 594_donkey_anti-mouse_IgG; 1:200; Jackson ImmunoResearch Laboratories). The corresponding isotype antibodies were used for negative control staining. Cells were counterstained with DAPI (Sigma) and analyzed with an AxioObserver A1 fluorescence microscope and Axiovision software 4.71 (Zeiss).
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