The polyclonal rabbit anti-human HER2 primary antibody (Dako) was used for the tumour tissue biopsies, diluted 1:50 from the stock solution (10 μg ml−1), and incubated at 4°C overnight. Goat anti-rabbit secondary antibody labeled with Alexa Fluor 680 (Invitrogen) was incubated for 1 h at room temperature at a protein concentration of 250 nM. Antibody conjugates were fixed in place with 2% paraformaldehyde. The slides were mounted with Fluoromount-G and covered with a coverslip for microscopy. Serial sections were stained with hematoxylin and eosin (H&E) using an autostainer (Leica). NIR fluorescence microscopy was performed on a Nikon TE300 with a 4-channel Nikon TE300. The excitation and emission filters used were 650/45 nm and 710/50 nm for Alexa 680 imaging, and 750/50 nm and 810/40 nm for NIR targeted tumour imaging, respectively.
Autostainer
Manufactured by Leica
Sourced in Germany, United States
The Leica Autostainer is a automated slide staining system designed for efficient and consistent sample preparation. It automates the staining process, allowing for precise control of staining protocols and reagent dispensing. The Autostainer is a core laboratory equipment piece that facilitates efficient slide processing.
Automatically generated - may contain errors
Lab products found in correlation
Tissue processor, by Leica (3 mentions)
Bz x700 microscope, by Keyence (3 mentions)
Rabbit anti human her2 primary antibody, by Agilent Technologies (2 mentions)
Alexa fluor 680, by Thermo Fisher Scientific (2 mentions)
Te300, by Nikon (2 mentions)
Nanozoomer scanner, by Hamamatsu Photonics (2 mentions)
Lsm 700, by Zeiss (2 mentions)
Hematoxylin, by Thermo Fisher Scientific (2 mentions)
Eosin y, by Thermo Fisher Scientific (2 mentions)
Paraffin wax, by Leica (1 mentions)
Tp1020, by Leica (1 mentions)
Dp71 camera, by Olympus (1 mentions)
Hematoxylin and eosin, by Merck Group (1 mentions)
Immpress polymerized reporter enzyme staining system, by Vector Laboratories (1 mentions)
Rotary microtome, by Leica (1 mentions)
Thunder imager tissue, by Leica (1 mentions)
Citrate buffer ph 6, by Merck Group (1 mentions)
Aperio scanscope system, by Leica (1 mentions)
Axiovision 4, by Zeiss (1 mentions)
Bond polymer refine detection kit, by Leica (1 mentions)
38 protocols using autostainer
1
Immunofluorescence Imaging of HER2 Expression
2
Deltamethrin Exposure Effects on Cockroach Tissues
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For histopathological evaluation, cockroach species P. americana was exposed to 0.25% (group I; N = 5), 0.5% (group II; N = 5), and 1% (group III; N = 5) of deltamethrin for 1 h. Whereas the insects exposed to acetone (group IV; N = 5) were taken as control. Brain, ovary and gut were dissected in saline solution (0.13 M NaCl; 0.01 M Na2HPO4; 0.02 M KH2PO4; pH 7.2) post 24 h of exposure and fixed in 10% neutral buffered formalin solution for another 24 h at room temperature. After fixation and overnight washing in distilled water, the extracted tissues were processed for dehydration in graded series of alcohol and toluene in an auto-tissue processor (Leica TP-1020, Germany). Processed tissues were embedded in paraffin wax (Leica, Germany). Multiple sections of 2.5–3 μm thickness from each block were cut on a rotatory microtome (Thermo Scientific, Microm, United States), mounted on pre-coated glass slide and air dried overnight. The sections were deparaffinised and stained with haematoxylin and eosin (McMannus and Mowry, 1965 ) in auto-stainer (Leica, Germany) and cover slipped by auto cover slipper (Leica, Germany). After drying, sections were photographed under a light microscope (Leica DMLB, Germany) using DM-500 camera (Leica, Germany).
3
Histopathological Tissue Analysis
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Frozen tissues kept at −80 °C were fixed in 70% ethanol. Fixed tissues were stained with hematoxylin and eosin (H&E) by Leica Autostainer (Histopathology Platform in Research Institute of MUHC). Hematoxylin and eosin-stained permanent sections were examined using an Olympus IX51 microscope (Olympus, Tokyo, Japan) equipped with an Olympus DP71 camera (Olympus).
4
Histological Analysis of Gonad Tissue in Sterile and Fertile Salmon
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Formalin‐fixed gonad tissues from sterile and fertile males and females (n = 6–10 per group) were processed overnight in Tissue Processor (Logos; Milestone). Paraffin embedded tissues were sectioned (2 μm) using a rotary microtome (Leica) and stained with hematoxylin and eosin (Merck) in auto‐stainer (Leica) at the Norwegian Veterinary Institute, Harstad. All slides were then analyzed at Nofima, Tromsø, using light microscopy and the QuPath (Quantitative Pathology & Bioimage Analysis) software. Germ cells were detected by immunohistochemical analysis of a subset of gonads using polyclonal rabbit antibodies against Atlantic salmon Vasa (Škugor et al., 2016 (link)) as primary antibodies and the ImmPRESS polymerized reporter enzyme staining system (Vector) with horseradish peroxidase anti‐rabbit immunoglobulin G as secondary antibody according to manufacturer's protocol. Sections treated without the primary antibody were used as negative controls.
5
Histological Analysis of Pancreatic Samples
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Pancreas samples were cut (1 cm thickness), fixed in 10% neutral buffer formalin, and embedded in paraffin wax to form sample blocks. A microtome was used to cut the wax blocks into 3–5 µm thickness sections, which were later mounted on labeled glass slides. A Leica Auto Stainer was then used to stain the slides with Hematoxylin and Eosin. Afterward, a light microscope was used to examine the pancreatic samples [25 ].
6
Liver Tissue Analysis Protocol
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Livers were fixed for 24 h in 10% neutral buffered formalin, dehydrated, embedded in paraffin and cut into 3 µm-thick sections. Tissue sections were stained with hematoxylin &eosin & saffron (HES) with Leica autostainer for preliminary analysis. Tumor differentiation and characteristics were reviewed by an expert pathologist (BR). Slides were digitally processed using the Nanozoomer scanner (Hamamatsu).
Immunofluorescence assays were performed after deparaffinization and antigen retrieval in citrate buffer pH6.0 (Sigma Aldrich). The Click-iT AF488 (ThermoFisher Scientific) EdU detection reaction was performed according to the manufacturer’s instructions. Labeling of c-Myc-positive cells was performed with human c-Myc antibody (Abcam rabbit ab#32072 dilution 1:200) and anti-rabbit secondary antibody (Thermofisher anti-rabbit IgG AF633, dilution 1:1000). Images were acquired using the Thunder Imager Tissue (Leica) microscope. Quantitative image analysis was performed using the QuPath v0.4.4 software (Bankhead et al, 2017 ). Foci were hand drawn and positive cell detection were performed using nuclear staining as basis of cell recognition.
Immunofluorescence assays were performed after deparaffinization and antigen retrieval in citrate buffer pH6.0 (Sigma Aldrich). The Click-iT AF488 (ThermoFisher Scientific) EdU detection reaction was performed according to the manufacturer’s instructions. Labeling of c-Myc-positive cells was performed with human c-Myc antibody (Abcam rabbit ab#32072 dilution 1:200) and anti-rabbit secondary antibody (Thermofisher anti-rabbit IgG AF633, dilution 1:1000). Images were acquired using the Thunder Imager Tissue (Leica) microscope. Quantitative image analysis was performed using the QuPath v0.4.4 software (Bankhead et al, 2017 ). Foci were hand drawn and positive cell detection were performed using nuclear staining as basis of cell recognition.
7
Histopathologic Evaluation of GA Aerosol Effects
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Morphological changes caused by GA aerosols were evaluated at T5 and PT7. Sample preparation and histopathology staining were performed as described previously [12 (link),13 (link)]. Hematoxylin and eosin (H&E) staining was conducted using a Leica Autostainer (Buffalo Grove, IL, USA); histopathologic changes were evaluated and scored by a board-certified pathologist. Digital images of the immunohistochemistry (IHC)-stained sections were obtained by scanning with the Aperio Scanscope System (Leica Biosystems, Vista, CA, USA). The percentages of cleaved caspase-3-positive apoptotic bodies, i.e., apoptotic index (AI), and periodic acid-Schiff (PAS)-stained goblet cells were calculated semi-automatically; the total numbers of nuclei were quantified automatically with the Nuclear Algorithm (Aperio Scanscope System), which counts the numbers of positive (brown) and negative (blue) nuclei in the section under examination, and the apoptotic bodies and PAS-stained goblet cells were counted manually. The percentage of Ki-67-positive nuclei, i.e., proliferative index (PI), was evaluated automatically with the Nuclear Algorithm.
8
Histopathological Diagnosis of Tuberculosis
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The specimen was transported to the TASH pathology lab in 10% formalin. After gross description by a pathologist, it was transferred into plastic cassettes and processed using Tissue Tek II VIP processor and then embedded in paraffin. After sectioning, it was stained by the Hematoxylin & Eosin (H&E) stain using a Leica autostainer. The Presence of caseating granulomas surrounded by epitheloid cells, lymphocytes, plasma cells and giant cells were diagnostic of tuberculosis.
9
Immunofluorescence Imaging of HER2 Expression
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The polyclonal rabbit anti-human HER2 primary antibody (Dako) was used for the tumour tissue biopsies, diluted 1:50 from the stock solution (10 μg ml−1), and incubated at 4°C overnight. Goat anti-rabbit secondary antibody labeled with Alexa Fluor 680 (Invitrogen) was incubated for 1 h at room temperature at a protein concentration of 250 nM. Antibody conjugates were fixed in place with 2% paraformaldehyde. The slides were mounted with Fluoromount-G and covered with a coverslip for microscopy. Serial sections were stained with hematoxylin and eosin (H&E) using an autostainer (Leica). NIR fluorescence microscopy was performed on a Nikon TE300 with a 4-channel Nikon TE300. The excitation and emission filters used were 650/45 nm and 710/50 nm for Alexa 680 imaging, and 750/50 nm and 810/40 nm for NIR targeted tumour imaging, respectively.
10
Immunohistochemical Localization of VEGF-A
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During dissection, 750 μl formalin (4%) was instilled in the lungs with a catheter. Thereafter, lungs were completely submerged in formalin (4%) for 24 h. Paraffin sections were pre-treated with citrate (pH 6). Subsequently, endogenous peroxidase activity of the lung sections were blocked for 5 min with a peroxide block (Bond Polymer Refine Detection kit, Leica, Diegem, Belgium). The sections were further incubated for 30 min at room temperature (RT) with monoclonal rabbit anti-mouse VEGF-A (dilution 1/100; EP1176Y, Abcam, Cambridge, UK) in Bond Primary Antibody Diluent (Leica), HRP-labeled goat-anti rabbit (8 min at RT), 3,3′-diaminobenzidine tetrahydrochloride hydrate (DAB) chromogen (10 min, Bond Polymer Refine Detection kit, Leica), and hematoxyline (5 min, Bond Polymer Refine Detection kit, Leica), using an autostainer (Leica). Transmitted light images were taken through a 10x/0.25 or a 40x/0.65 N Plan objective of a Leica DM2000 microscope. Image background adjustments were performed with the AxioVision 4.6 software (Zeiss, Zaventem, Belgium).
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