Newborn pigs were sedated using isoflurane and viral vector formulated with NaCl (0.9% or 5% final) and aerosolized intratracheally using a MADgic Laryngo-Tracheal Mucosal Atomization Device (Teleflex, Morrisville, NC). Doses for each vector for each pig were as follows: 2.8×1010 infectious genomic units (IGU) of helper-dependent Ad-CMV-GFP and 7×108 TU of GP64 HIV-CMV-GFP. 1 week later, pigs were humanely euthanized and lungs were analyzed for GFP expression. Lungs were systematically divided and fixed in 4% paraformaldehyde, subjected to a sucrose gradient, and embedded in OCT for cryosectioning. Sectioned lung slides were mounted using DAPI and imaged using a Keyence All-in-one Fluorescence Microscope BZ-X series (Osaka, Japan).
Bz x series all in one fluorescence microscope
Manufactured by Keyence
Sourced in Japan
The BZ-X series All-in-One Fluorescence Microscope is a compact, all-in-one microscope system designed for high-resolution fluorescence imaging. It features automated functions and advanced imaging capabilities to facilitate various microscopy applications.
Automatically generated - may contain errors
Lab products found in correlation
Sm2010r microtome, by Leica (2 mentions)
Thio s stain, by Merck Group (2 mentions)
Foxp3 transcription factor staining buffer set, by Thermo Fisher Scientific (1 mentions)
Live dead stain, by Thermo Fisher Scientific (1 mentions)
Attune nxt flow cytometer, by Thermo Fisher Scientific (1 mentions)
H 3300, by Vector Laboratories (1 mentions)
Pk 4001, by Vector Laboratories (1 mentions)
Sk 4100, by Vector Laboratories (1 mentions)
5 protocols using bz x series all in one fluorescence microscope
1
Porcine Intratracheal Viral Vector Delivery
2
Quantification of GFP Expression in Cells
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GFP images were acquired using a Keyence All-in-one Fluorescence Microscope BZ-X series (Osaka, Japan). 0.33 cm2 transwells were imaged at 2X magnification. GFP expression was quantified by flow cytometry as previously reported39 (link), 41 (link). Briefly, cells were stained with a fixable LIVE/DEAD stain (Thermo Fisher Scientific, Waltham, MA), lifted in Accutase at 37°C for 30 minutes, and run through an Attune NxT Flow Cytometer (Thermo Fisher Scientific, Waltham, MA). Cells were treated with the Foxp3/Transcription Factor Staining Buffer Set (Thermo Fisher Scientific, Waltham, MA, USA) according to manufacturer’s recommendations and stained for 1 hour at 4°C with the following antibodies: NGFR (345110; 1:600, BioLegend, San Diego, CA, USA), α-tubulin (NB100–69AF405, 1:300, Novus, Centennial, CO, USA) and CD66c (12-0667-42, 1:600, Invitrogen, Waltham, MA). Expression was gated on live cells.
3
Immunohistochemical Analysis of PTEN in Mouse Prostate
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Mouse prostate tissues were fixed in 4% of formaldehyde for 48 h, and paraffin-embedded through Molecular Pathology and Imaging Core at UPenn. Paraffin-embedded sections from the prostate of Pten floxed control and Pten knockout mice were deparaffinized with 3 changes of xylene for 5 min each. Slides were then rehydrated in 100% alcohol for 10 min, 95% alcohol twice for 10 min each, and 70% alcohol and distilled water for 10 min each. Slides were subjected to citrate-based (pH 6.0) antigen retrieval (Vector, H-3300) at 95 °C for 30 min and followed by blocking endogenous peroxidase activity with 3% H2O2 for 5 min. After three times 5 min TBST washing, slides were blocked with blocking buffer (1.25% of goat serum) at room temperature for 1 h. Slides were applied with diluted primary antibody and incubated in a humidified chamber at 4 °C overnight. After three 10 min TBST washes, slides were incubated with secondary antibody (Vector, PK-4001) for 30 min at room temperature the color of the antibody staining was revealed by peroxidase-based detection (Vector, SK4100) and the sections were counterstained with hematoxylin (Millipore Sigma, MHS1). Representative photographs were taken on a Keyence BZ-X Series All-in-one Fluorescence Microscope with 20x and 40x objectives. Images were assessed and quantified in ImageJ.
4
Thioflavin-S Staining of Hippocampal Slices
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At 6 months of age, mice were anesthetized with isoflurane and transcardially perfused with ice-cold PBS. The whole brain was extracted and fixed in 4% paraformaldehyde for ~24 h at 4 °C, and then cryoprotected in 15% sucrose in PBS for ~24 h followed by 30% sucrose in PBS for another ~24 h at 4 °C. The brains were then sectioned into 40 μm slices using a Leica SM 2010R microtome. Hippocampal slices were mounted on glass slides with 8–10 slices per slide. The slices were then allowed to dry completely before staining (about 2–3 h). Thioflavin-S staining followed a standard staining procedure with a 7-chamber staining apparatus. The slides were first washed with 70% EtOH for one minute followed by a wash in 80% EtOH for one minute. The slides were then incubated in filtered Thio-S stain (Sigma-Aldrich, 1% in 80% EtOH) for 15 min. The slides were protected from light. Following the staining, the slides were washed in 80% EtOH for one minute and then 70% EtOH for one minute. Finally, the slices were washed twice with distilled water. The slices were allowed to dry in a light-tight container for at least two hours then coverslipped using VECTASHEILD HardSet Antifade Mounting Medium with DAPI. Thio-S-stained beta-pleated sheets were visualized using a Keyence BZ-X series All-in-One Fluorescence Microscope.
5
Thioflavin-S Staining of Hippocampal Slices
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