Tissues were fixed in 4% paraformaldehyde, embedded in paraffin wax, and sectioned at 5 µm intervals. Slides were stained with hematoxylin using standard protocols, as previously described (Herriges et al., 2014 (link); Swarr et al., 2019 (link)). Immunofluorescence staining was performed using the following antibodies: anti-NKX2.1 (guinea pig, Seven Hills, 1:500), anti-SOX9 (rabbit, Millipore, 1:100), anti-KI67 (mouse, BD Pharmigen, 1:100), anti-SCGB1A1 (rabbit, Seven Hills, 1:500), anti-TUB1A1 (mouse, Sigma Aldrich, 1:1000), and anti-SOX2 (mouse, Santa Cruz, 1:100). Immunofluorescence for phosphorylated-ATK (pAKT) was performed with anti-pAKT (rabbit, Cell Signaling, 1:100), followed by multi-step detection with the Biotin-Tyramide signal amplification kit (Perkin Elmer) with a 15-min exposure time. TUNEL staining was performed using the TACS-XL Blue Label kit, per manufacturer’s instructions (Trevigen). All slides were mounted with Prolong Gold Antifade medium (Invitrogen), and were then imaged on either a Nikon Eclipse 90i widefield, or Nikon A1R GaAsP Inverted Confocal Microscope. Cell type quanitification based on immunofluorescence microscopy images was performed using the Nikon Elements Advanced Analysis software suite.
Tacs xl blue label kit
Manufactured by Bio-Techne
Sourced in United States
The TACS XL Blue Label kit is a lab equipment product designed for end-labeling of DNA and RNA for use in various molecular biology applications. It provides the necessary reagents and components to perform end-labeling procedures.
Automatically generated - may contain errors
Lab products found in correlation
Prolong gold antifade medium, by Thermo Fisher Scientific (1 mentions)
Anti sox9, by Merck Group (1 mentions)
Anti p akt, by Cell Signaling Technology (1 mentions)
Anti sox2, by Santa Cruz Biotechnology (1 mentions)
A1r gaasp inverted confocal microscope, by Nikon (1 mentions)
Optical cutting temperature compound, by Sakura Finetek (1 mentions)
Dp72 digital camera, by Olympus (1 mentions)
Dfc425, by Leica (1 mentions)
Dfc420 digital camera, by Leica (1 mentions)
Paraplast plus, by Merck Group (1 mentions)
5 protocols using tacs xl blue label kit
1
Quantitative Immunohistochemical Analysis
2
Apoptosis Analysis in Tumor Tissues
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Tumor masses of the mice were harvested at day 14 following treatment. Tumor tissues were embedded in optical cutting temperature compound (Sakura Finetek USA, Inc., Torrance, CA, USA), frozen in liquid nitrogen and maintained at −80°C. Tumor tissues were then cryosectioned into 10-µm sections for apoptosis, and hematoxylin and eosin (Boster Biotechnology Co., Ltd., Wuhan, China) staining. Level of apoptosis was determined via a terminal deoxynucleotidyl transferase dUTP nick end-labeling assay using a TACS XL Blue Label kit (Trevigen, Gaithersburg, MD, USA) according to the manufacturer's protocol. For each slide, images from random six fields were captured using an Olympus digital camera (IX53; Olympus Corporation, Tokyo, Japan). Apoptosis results were analyzed using the apoptotic index, which was defined as the number of apoptotic cells/total number of cells in each field.
3
Apoptosis Quantification in Tumor Sections
4
Quantifying Cell Death in Brain Sections
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Cell death was determined on 6-μm paraffin-embedded brain sections using the TACS XL Blue Label kit (Trevigen, Gaithersburg, MD), following the manufacturer’s protocol. For positive control, samples were treated with TACS-nuclease to generate DNA breaks in cells. Slides were examined under a Nikon light microscope and images were acquired by a Leica DFC425 camera. Image analysis was conducted by the LAS V4.0 program (Leica Microsystems, Inc., Buffalo Grove, IL) by counting the positively stained cells per mm2 from 10 different fields from each of the three slide replicates for a total of 30 fields per group.
5
Ovule TUNEL Assay for Cell Death
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Isolated ovules were fixed overnight at 4°C in 4% (w/v) buffered paraformaldehyde (PBS, pH 7.4); then they were dehydrated in ethanol, embedded in Paraplast Plus (Sigma-Aldrich, Milan, Italy) at 60°C for 48 hr, sliced in 10 µm sections and finally collected on poly-lysine coated slides. Deparaffinized sections were rehydrated and subjected to TUNEL assay (Terminaldeoxynucleotidyl transferase (TdT)-mediated dUTPnick-end labelling), by using TACS•XL Blue Label Kit (Trevigen, Gaithersburg, MD, USA) according to Bartoli et al. (2015) . The sections were air-dried, mounted with DPX, observed with a LEITZ DIAPLAN light microscope (Wetzlar, Germany) and then captured using a Leica DFC 420 digital camera (Leica Microsystems, Heerbrugg, Germany). The assay was repeated at least three times.
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