The following reagents were purchased from Beijing Bioss Biotechnology: c-fos rabbit anti-mouse polyclonal antibody, ZO-1 rabbit anti-mouse polyclonal antibody, Occludin rabbit anti-mouse polyclonal antibody, myosin light chain kinase (MLCK) rabbit anti-mouse polyclonal antibody, P-glycoprotein (P-gp) rabbit anti-mouse polyclonal antibody, multidrug resistance associated protein 1 (MRP1) rabbit anti-mouse polyclonal antibody, multidrug resistance associated protein 2 (MRP2) rabbit anti-mouse polyclonal antibody, goat anti-rabbit immunohistochemistry kit, and hematoxylin. Other reagents used were as follows: glacial acetic acid, analytical grade (Beijing Chemical Works, Batch No. 20100603); osmic acid, acetone, embedding media, uranyl acetate stain, and lead citrate stain (supplied by the Experimental Research Center); potassium permanganate, analytical grade (Beijing Chemical Works, Batch No. 820308); toluidine blue, analytical grade (Sinopharm Chemical Reagents, Batch No. WC20050120); anhydrous ethanol, analytical grade (Beijing Chemical Works, Batch No. 20110); xylene, analytical grade (Sinopharm Chemical Reagents, Batch No. 20106); neutral gum (Sinopharm Chemical Reagents, Batch No. 20120320); hematoxylin (Beyotime Biotechnology, Batch No. C0105); ethylenediamine tetraacetic acid (EDTA), analytical grade (Beijing Chemical Works, Batch No. 840529).
Toluidine blue
Manufactured by Sinopharm
Sourced in Germany
Toluidine blue is a basic dye commonly used in laboratory settings. It is a metachromatic dye, meaning it can exhibit different colors depending on the type of material it is staining. Toluidine blue is often used for staining and visualization of various biological samples, including tissues, cells, and nucleic acids.
Automatically generated - may contain errors
Lab products found in correlation
Neutral gum, by Sinopharm (3 mentions)
Em uc7, by Leica (2 mentions)
Anhydrous ethanol, by Beijing Chemical Works (1 mentions)
Glacial acetic acid, by Beijing Chemical Works (1 mentions)
Hematoxylin, by Beyotime (1 mentions)
Ethylenediamine tetraacetic acid edta, by Beijing Chemical Works (1 mentions)
Rm2235, by Leica (1 mentions)
M165 fc stereomicroscope, by Leica (1 mentions)
Eclipse e100, by Nikon (1 mentions)
Glacial acetic acid, by Sinopharm (1 mentions)
Axio scope a1 microscope, by Zeiss (1 mentions)
Stemi 508 stereo microscope, by Zeiss (1 mentions)
Tunel assay, by Yeasen (1 mentions)
8 protocols using toluidine blue
1
Immunohistochemical Analysis of Cellular Markers
2
Ultrastructural Analysis of Cultured Cells
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Cultured fVBOrs were washed with DPBS (Life/Invitrogen) and cut into 1 mm × 1 mm small pieces. Firstly, samples were fixed with 4% PFA overnight and then were pre-fixed with 2.5% glutaraldehyde (SPI, USA), in PBS for 12 hr. After washing with PBS, samples were post-fixed with 1% OsO4 (TED PELLA) for 2 hr at 4°C. Then, dehydrated in an ascending gradual series (30–100% [v/v]) of ethanol and embedded in epoxy resin (Pon812 kit, SPI). The embedded samples were initially cut into about 500-nm-thick sections, inspected by stained with toluidine blue (Sinopharm), and finally sectioned into 70 nm by Leica EM UC7. Then sections were double-stained with uranyl acetate (SPI) and lead citrate (SPI), followed by observation with a TEM (Talos L120C) at an acceleration voltage of 80 kV.
3
Preparation of Paraffin-Embedded Plant Seed Sections
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To prepare the paraffin sections of kernels, immature seeds were fixed overnight at 4 °C in a formalin‐acetic acid‐alcohol (FAA) solution containing 50% ethanol, 5% acetic acid and 3.7% formaldehyde. The fixed materials were then dehydrated in an ethanol gradient series (50, 70, 85, 95 and 100% ethanol). Afterwards, the samples were treated with xylene, embedded in paraffin wax via infiltration and cut into 6–10 µm‐thick sections under Leica RM2235 (Germany). The sections were stained with toluidine blue (Sinopharm Chemical Reagent Co., Ltd) and examined under the Lecia M165FC stereomicroscope (Germany). The endosperm structures of mature and immature seeds of different developmental stages were observed by scanning electron microscopy (SEM) and transmission electron microscopy (TEM), respectively (Wu and Messing, 2010 ; Zhang et al., 2016 ).
4
Quantifying Spinal Cord Motor Neurons
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We used Nissl staining to identify spinal cord motor neurons. A decrease in the number of Nissl bodies and abnormal motor neuron morphology indicated that nerve cells may be damaged. Paraffin sections of the lumbosacral enlargements of the spinal cord were deparaffinized and rehydrated, and the sections were stained with toluidine blue (Sinopharm Chemical Reagent Co., Ltd., Shanghai, China) for 5 minutes, washed with water, and differentiated with 1% glacial acetic acid (Sinopharm Chemical Reagent Co., Ltd.). The reaction was terminated with tap water. The degree of stain differentiation was controlled by observation under an upright microscope (ECLIPSE E100, Nikon). After washing with water, the sections were dried in an oven (Sinopharm Chemical Reagent Co. Ltd.), cleared with xylene for 5 minutes, and mounted with neutral gum (Sinopharm Chemical Reagent Co., Ltd.). Serial coronal slides of the paraffin-embedded lumbosacral enlargements of the spinal cord were obtained and analyzed at intervals of six sections. The section thickness was 5 μm. The slides were scanned using an upright microscope (ECLIPSE E100, Nikon) with a 40× objective lens. The number of motor neurons in the lateral nucleus of the anterior horn of the spinal cord was manually calculated in three non-overlapping areas (220 × 350 μm2).
5
Paraffin Sectioning and Staining for Plant Samples
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Paraffin sections of 10-DAP kernels were prepared according to a previously described protocol [73 (link)]. The samples were treated with xylene, embedded in paraffin wax via infiltration, and cut into 10 μm-thick sections with a Thermo Scientific HM 325 rotary microtome. The sections were stained with toluidine blue (Sinopharm Chemical Reagent Co., Ltd) and examined under a ZEISS Stemi 508 Stereo Microscope (Germany) and a ZEISS Axio Scope A1 Microscope (Germany).
6
Hippocampal Apoptosis in Cardiac Arrest
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Rats were sacrificed 24 h after the ROSC. Hippocampal tissues were carefully removed and immersed in paraformaldehyde overnight. Then, all tissues were embedded in paraffin and sliced into 5-μm sections. For histologic assessment, sections were stained with 0.01% Toluidine blue (Sinopharm Group, Shanghai, China) and mounted. Terminal deoxynucleotide transferase-mediated dUTP-biotin nick-end labeling (TUNEL) assay (Yeasen Biotech, Shanghai, China) was performed using an in-situ apoptosis detection kit, as per the manufacturer's instruction. Three fields of vision in the hippocampal CA area were randomly selected, and the viable neurons and TUNEL-stained neuronal cells were counted under an optical microscope.
7
Toluidine Blue Staining Nissl Bodies
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After the tissue paraffin sections were dewaxed, hydrated, and washed with distilled water, they were stained with 0.25% toluidine blue (Sinopharm Group; 71041284) at 60°C for 3 h. The sections were then washed with 95% ethanol, dehydrated with absolute ethanol, made transparent with xylene (Sinopharm Group; 10023418), and eventually assembled with neutral gum (Sinopharm Group; 10004160). The sections were observed and images were captured under a vertical fluorescent microscope (Olympus BX53, Tokyo, Japan). The average optical density of Nissl bodies was analyzed by ImageJ 5.0 software (Rawak Software Inc., Stuttgart, Germany) by three researchers who were blinded to the treatments.
8
Transmission Electron Microscopy of fVBOr
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Cultured fVBOrs were washed with DPBS (Life/Invitrogen) and cut into 1 mm 1 mm small pieces. Firstly samples were fixed with 4% paraformaldehyde overnight and then were pre-fixed with 2.5% glutaraldehyde (SPI, USA), in PBS for 12 hr. After washing with PBS, samples were post-fixed with 1% OsO4 (TED PELLA) for 2 hr at 4°C. Next, followed by dehydrated in an ascending gradual series (30%-100% (v/v)) of ethanol, and embedded in epoxy resin (Pon812 kit, SPI, USA). The embedded samples were initially cut into about 500 nm-thick sections, inspected by stained with toluidine blue (Sinopharm), and finally sectioned into 70-nm by Leica EM UC7. Then sections were double-stained with uranyl acetate (SPI, USA) and lead citrate (SPI, USA), followed by observation with a Transmission Electron Microscopy (Talos L120C) at an acceleration voltage of 80 kV.
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