Third-instar larvae of H. armigera strains LF, LF5, and LF60 were starved for 24 h, then allowed to feed on an artificial diet with 1.2 µg/g of Vip3AcAa, 1.2 µg/g of Cry1Ac toxin, or 0.01 M PBS buffer (control) or on non-Bt cotton Coker312, NuCOTN33B, or CV163 for 96 h. Excised midguts were fixed in 4% v/v paraformaldehyde in PBS buffer, dehydrated in a graded ethanol series, and then rinsed in toluene and embedded in paraffin wax. Four-micrometer sections were cut using a Thermo Scientific Finesse ME+ microtome and placed on microscope slides coated with a mixture of 1.5% egg albumin and 3% glycerol in distilled water. Sections were deparaffinized in 100% toluene, then stained with hematoxylin–eosin according to the protocol of Ruiz et al. (2004) . The strained tissues were observed using an optical microscope (Optika, XDS-3FL4) and photographed.
Finesse me microtome
Manufactured by Thermo Fisher Scientific
Sourced in United States
The Finesse ME+ microtome is a specialized laboratory instrument designed for the precise sectioning of samples for microscopic analysis. It features a precise, motorized mechanism that allows for the controlled and accurate cutting of thin specimens, enabling detailed examination and study of the sample's structure and composition.
Automatically generated - may contain errors
Lab products found in correlation
Histostar embedding workstation, by Thermo Fisher Scientific (3 mentions)
Citadel 2000 tissue processor, by Thermo Fisher Scientific (3 mentions)
Ctm6 coverslipper, by Thermo Fisher Scientific (2 mentions)
Gemini as automated slide stainer, by Thermo Fisher Scientific (2 mentions)
Xds 3fl4, by Optika (1 mentions)
Application suite, by Leica (1 mentions)
Dm2000 microscope, by Leica (1 mentions)
Axiocam mrc5 camera, by Zeiss (1 mentions)
Perfection v600 scanner, by Epson (1 mentions)
Histostar workstation, by Thermo Fisher Scientific (1 mentions)
Histomix, by BioVitrum (1 mentions)
Microm stp 120, by Thermo Fisher Scientific (1 mentions)
Vitrogel, by BioVitrum (1 mentions)
8 protocols using finesse me microtome
1
Histological Analysis of Bt Toxin Effects
2
Histomorphometric Analysis of Intestinal Tissues
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After fixation, intestinal tissues were dehydrated through an ethanol series and xylene substitute (Citadel 2000 Tissue Processor, Thermo Fisher Scientific, Waltham, MA, USA), then embedded in synthetic paraffin. After paraffin block formation (Histo Star Embedding Workstation, Thermo Fisher Scientific), 4-µm sections were cut (Finesse ME+ Microtome, Thermo Fisher Scientific), stained with hematoxylin and eosin (Panreac Química SLU) and mounted and examined under a light microscope (Leica, Wetzlar, Germany).
A histomorphometric study was performed using an image analyzer (Leica Application Suite, Leica), which measured different parameters of the duodenum and the cecum in each animal in five fields at 40× magnification. In each case, the number of duodenal villi and duodenal and cecal crypts were counted. Duodenal villi height was measured from the top of the villus to the villus–crypt junction; duodenal and cecal crypt depth was measured from the villus–crypt junction to the muscularis mucosae. A minimum of seven well-oriented villi and 14 crypts were measured from different sections of each hen.
A histomorphometric study was performed using an image analyzer (Leica Application Suite, Leica), which measured different parameters of the duodenum and the cecum in each animal in five fields at 40× magnification. In each case, the number of duodenal villi and duodenal and cecal crypts were counted. Duodenal villi height was measured from the top of the villus to the villus–crypt junction; duodenal and cecal crypt depth was measured from the villus–crypt junction to the muscularis mucosae. A minimum of seven well-oriented villi and 14 crypts were measured from different sections of each hen.
3
Histological Staging of Bovine Tuberculosis
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Tissue samples collected during necropsy were fixed 24 hours, trimmed, dehydrated (Citadel 2000 Tissue Processor, Thermo Fisher Scientific, Waltham, MA), embedded in paraffin (Histo Star Embedding Workstation, Thermo Fisher Scientific), sectioned (Finesse ME + Microtome, Thermo Fisher Scientific) and stained with hematoxylin-eosin (HE) (Gemini AS Automated Slide Stainer, Thermo Fisher Scientific). Finally, samples were mounted in glass slides (CTM6 Coverslipper, Thermo Fisher Scientific). HE-stained sections were evaluated and classified according to a published staging of granulomas in M. bovis-infected cattle (Wangoo et al. 2005 (link)): stage I (initial), II (solid), and III (minimal necrosis). Stage IV granulomas were not included due to their structural similarity to stage III granulomas, despite a larger size and multicentric nature.
4
Histological Analysis of Epididymal WAT
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Epididymal white adipose tissues (eWATs) were fixed in 10% buffered formalin and embedded in paraffin. Using a Finesse ME Microtome (Thermo, Waltham, MA, USA), the samples were cut at 3 μm thickness and stained with hematoxylin and eosin for histological examination.
5
Perfusion-Fixation of Mouse Brain
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Once the mouse is unconscious under respiratory anesthesia, make an incision between the ribs and abdomen to access the heart. Once visualized, insert the needle into the convex convergence at the tip of the heart to inject phosphate buffered saline (PBS) via syringe. Once the needle is inserted, slowly inject PBS to achieve perfusion. Once perfusion is achieved, the brain is obtained through the same incision and the tissue is fixed by complete immersion in 4% paraformaldehyde. The fixed brain tissue was sectioned to a thickness of approximately 2–3 mm and subjected to tissue processing (STP120 Spin tissue Processor; Especialidades Médicas MYR, S.L., Tarragona, Spain). The processed tissue was sectioned using a sectioning machine (Finesse ME Microtome; Thermo Fisher Scientific, Waltham, MA, USA) to a thickness of approximately 3–4 µm, and the sections were attached to slides for drying, deparaffinized in xylene for 3 steps/3 minutes, hydrated in alcohol for 4 steps/2 minutes, hematoxylin for 10 minutes, water for 3 minutes, eosin for 1 minute 40 seconds, alcohol for 4 steps/1 minute, cleared in xylene for 3 steps/3 minutes, and embedded to prepare H&E sections.
6
Histopathological Evaluation of ASF Infection
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Animals were necropsied and post-mortem evaluation was performed. Macroscopic changes observed in experimentally ASF-infected wild boars were evaluated according to previous protocols (26 (link), 30 (link)). A wide variety of tissue samples from various organs were taken (26 (link)), fixed in 10% neutral formalin and routinely processed for histopathological examination. After fixation, samples were trimmed, dehydrated (Citadel 2000 Tissue Processor, Thermo Fisher Scientific, Waltham, MA), and embedded in paraffin (Histo Star Embedding Workstation, Thermo Fisher Scientific) following standard procedures. Each tissue sample block was sectioned (Finesse ME + Microtome, Thermo Fisher Scientific) and stained with hematoxylin–eosin (HE) (Gemini AS Automated Slide Stainer, Thermo Fisher Scientific). Selected sections of the IPAS were additionally stained with Masson’s trichrome (MT) for the detection of collagen fibres. Finally, samples were mounted in glass slides (CTM6 Coverslipper, Thermo Fisher Scientific) and evaluated for histopathological alterations under a Leica DM2000 microscope (Leica Microsystems, Wetzlar, 162 Germany).
7
Cardiac Tissue Preparation for Light and Electron Microscopy
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For light microscopy, hearts fixed in 10% neutral-buffered formalin were embedded in paraffin wax and sectioned using a Finesse ME+ microtome (Thermo Fisher, Waltham, MA, USA). Transverse sections (T/S) were stained with haematoxylin and eosin.
For transmission electron microscopy (TEM), 1 mm3 cubes of left ventricular tissue were fixed in 3% glutaraldehyde and 4% formaldehyde in 0.1 M PIPES and post-fixed with 1% osmium tetroxide in 0.1 PIPES. Samples were taken from the left ventricular free wall of three wild-type and three EcsitN209I/N209I males at 16 weeks of age. After serial dehydration in an increasing concentration of ethanol, the tissue was embedded in epoxy resin (TAAB) and polymerized overnight at 60°C. Golden ultrathin sections (70–80 nm) were cut with a diamond knife and collected on copper/palladium grids. To improve contrast, blocks were stained with 2% uranyl acetate and grids were stained with lead citrate.
Images were collected at the Wolfson bioimaging facility at the University of Bristol using a Tecnai 12 Biotwin electron microscope.
For transmission electron microscopy (TEM), 1 mm3 cubes of left ventricular tissue were fixed in 3% glutaraldehyde and 4% formaldehyde in 0.1 M PIPES and post-fixed with 1% osmium tetroxide in 0.1 PIPES. Samples were taken from the left ventricular free wall of three wild-type and three EcsitN209I/N209I males at 16 weeks of age. After serial dehydration in an increasing concentration of ethanol, the tissue was embedded in epoxy resin (TAAB) and polymerized overnight at 60°C. Golden ultrathin sections (70–80 nm) were cut with a diamond knife and collected on copper/palladium grids. To improve contrast, blocks were stained with 2% uranyl acetate and grids were stained with lead citrate.
Images were collected at the Wolfson bioimaging facility at the University of Bristol using a Tecnai 12 Biotwin electron microscope.
8
Assessing Lung Pathology in Mice
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Lungs from each mouse were fixed in 10% neutral buffered formalin (Soluformtm, JLS-Chemical, Russia) at +4°C, dehydrated in isoprepe (BioVitrum, Russia) using Microm STP 120 (Thermo Scientific, USA), and embedded in HISTOMIX (BioVitrum, Russia) using HistoStar workstation (Thermo Scientific, USA). 5-μm thick sections (10 per mouse) were cut using a Finesse ME+ microtome (Thermo Scientific, USA), stained with haematoxylin and eosin and mounted in Vitrogel (all BioVitrum, Russia). Pictures were obtained either using an Epson Perfection V600 scanner (9600 dpi) or using an Imager. Z1 microscope with an AxioCam MRc 5 camera (all ZEISS, Germany) at 20х and 63х magnification. Pathological changes were assessed using three parameters: (1) the acute lung injury (ALI) score, showing mostly the immunopathology in the lung parenchyma (from 0 to 1); (2) the peribronchiolar infiltration score, estimating the degree of inflammation of the tissue around the bronchioles (from 0 to 5); and (3) the perivascular infiltration score, estimating the degree of inflammation of the tissue around vessels (from 0 to 5). The exact parameters of tissue scoring systems are provided in the Supplementary Materials (p. 9).
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