Livers were collected after euthanasia and fixed by immersion in 10% formalin (Thermo Fisher Scientific) for 24 hours, washed in PBS and transferred to 70% EtOH in water until they were embedded in paraffin blocks. Serial sections (4 μm thick) were cut with a microtome. Hematoxylin & Eosin (H&E) and periodic acid Schiff (PAS) staining were performed following standard protocols, and the sections were analyzed by light microscopy. EHHADH immunofluorescence to detect hepatic peroxisomes was performed as previously described (Ranea-Robles et al. 2021a (link)) using a specific anti-EHHADH antibody (dilution 1:50, GTX81126, Genetex). Microscopy images were taken with a Nikon Eclipse 80i microscope and the NIS-Elements BR 5.20.01 software (Nikon). Images were analyzed with ImageJ (Schindelin et al. 2012 (link)).
Nis elements br 5
Manufactured by Nikon
Sourced in United States
NIS-Elements BR 5.20.01 is a software suite for microscope imaging and analysis. It provides tools for image capture, processing, and measurement. The software supports a range of Nikon microscope models and accessories, enabling users to manage their microscopy workflows.
Automatically generated - may contain errors
Lab products found in correlation
Formalin, by Thermo Fisher Scientific (2 mentions)
Gtx81126, by GeneTex (2 mentions)
Eclipse 80i microscope, by Nikon (2 mentions)
Eclipse e800 microscope, by Nikon (2 mentions)
Dab substrate, by Vector Laboratories (1 mentions)
Abc reagent, by Vector Laboratories (1 mentions)
Pcna antibody, by Cell Signaling Technology (1 mentions)
Cytopro, by Wescor (1 mentions)
Anti histone h3, by Abcam (1 mentions)
4 protocols using nis elements br 5
1
Histological Analysis of Hepatic Peroxisomes
2
Histological Analysis of Liver Peroxisomes
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Livers were collected after euthanasia and fixed by immersion in 10% formalin (Thermo Fisher Scientific) for 24 hours, washed in PBS and transferred to 70% EtOH in water until they were embedded in paraffin blocks. Serial sections (4 µm thick) were cut with a microtome.
Hematoxylin & Eosin (H&E) and periodic acid Schiff (PAS) staining were performed following standard protocols, and the sections were analyzed by light microscopy. EHHADH immunofluorescence to detect hepatic peroxisomes was performed as previously described (Ranea-Robles et al. 2021a) using a specific anti-EHHADH antibody (dilution 1:50, GTX81126, Genetex). Microscopy images were taken with a Nikon Eclipse 80i microscope and the NIS-Elements BR 5.20.01 software (Nikon). Images were analyzed with ImageJ (Schindelin et al. 2012) (link).
Hematoxylin & Eosin (H&E) and periodic acid Schiff (PAS) staining were performed following standard protocols, and the sections were analyzed by light microscopy. EHHADH immunofluorescence to detect hepatic peroxisomes was performed as previously described (Ranea-Robles et al. 2021a) using a specific anti-EHHADH antibody (dilution 1:50, GTX81126, Genetex). Microscopy images were taken with a Nikon Eclipse 80i microscope and the NIS-Elements BR 5.20.01 software (Nikon). Images were analyzed with ImageJ (Schindelin et al. 2012) (link).
3
Immunohistochemical Analysis of MPO in Breast Cancer
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Immunohistochemistry was performed as described previously [51 (link)]. In brief, we stained the human breast cancer array with MPO (Abcam, Cambridge, MA, USA) antibody. Immunoreactivity was determined using ABC reagent (Vector Laboratories, Burlingame, CA, USA) and DAB substrate (Vector Laboratories). Slides were counterstained with hematoxylin. The number of MPO positive cells was counted per core. The details of the antibodies are listed in Table 2 . The representative pictures were acquired with a Nikon Eclipse E800 microscope (Nikon, Melville, NY, USA) and NIS-Elements BR 5.11.00 software (Nikon).
4
NET and PCNA Quantification in MPRO Cells
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We cultured the MPRO Clone 2.1 cells (1 × 106 cells per well of a 96 well plate) in SF media and the supernatant of Cl66, Cl66-Dox, and Cl66-Pac for 4 h for staining NET using the Anti-Histone H3 (Abcam) antibody. For PCNA staining, MPRO Clone 2.1 cells (1 × 106 cells per well of a 96 well plate) were treated with the supernatant of Cl66, Cl66-Dox, and Cl66-Pac and SF media for 24 h using the PCNA antibody (Cell Signaling, Danvers, MA, USA). From these treated cells, 100 L was cytospinned using Cytopro (Wescor) on glass slides. These slides were air-dried overnight. The cells on the air-dried glass slide were outlined using a Pap pen. Immunofluorescence was performed as described previously [55 (link)]. The representative pictures were acquired with a Nikon Eclipse E800 microscope (Nikon, Melville, NY, USA) and NIS-Elements BR 5.11.00 software (Nikon). For quantification of NET producing cells, we quantified the total number of nucleus per image, and the number of the nucleus in the vicinity of NETs was quantified as NET producing cells. The percentage was calculated using the formula: NET producing cells/Total number of the nucleus in the image × 100 = Percentage of NET producing cells.
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