All experimental procedures were approved by the Institutional Animal Care and Use Committee (IACUC) of Allen Institute for Brain Science in accordance with NIH guidelines. All characterization was done using adult mice around ages P56 or older. The mice that were characterized were in a mixed genetic background, containing 50–75% C57BL/6 background and the remainders of 129 or other backgrounds from the various Cre lines. For systematic characterization of fluorescent proteins either by their native fluorescence or IHC, perfused brains were cryosectioned using a tape transfer technique, sections were then DAPI stained directly or following antibody staining, and images were captured using automated fluorescent microscopy. Microtome sections of 100-μm thickness from perfused brains were used for confocal imaging of fluorescently labeled cells. For systematic characterization of gene expression by colorimetric ISH or DFISH, the Allen Institute established pipelines for tissue processing, probe hybridization, image capture and data processing were utilized. Informatics signal identification, mapping, and quantification used the Allen Mouse Brain Atlas spatial mapping platform24 (link), 29 . In this pipeline, image series are preprocessed (white-balanced and cropped), then registered to a three-dimensional informatics reference atlas of the C57BL/6J mouse brain28 . This registration enables data to be displayed in 2D sections or reconstructed 3D volumes.
>
Procedures
>
Laboratory Procedure
>
Microtomy
Microtomy
Microtomy is the technique of sectioning tissues into thin slices for microscopic examination.
It involves the use of specialized instruments called microtomes to precisely cut samples into thin, uniform sections.
These sections can then be stained and analyzed, allowing researchers to study the detailed structure and organization of biological tissues at the cellular and subcellular level.
Microtomy is an essential tool in fields such as histology, pathology, and developmental biology, enabling in-depth exploration of tissue morphology and facilitating the diagnosis of various medical conditions.
The process of microtomy requires careful sample preparation, instrument calibration, and sectioning technique to ensure high-quality, reproducible results.
Unlocking the power of microtomy can lead to groundbreaking discoveries and advancements in biomedical research and clinical practice.
It involves the use of specialized instruments called microtomes to precisely cut samples into thin, uniform sections.
These sections can then be stained and analyzed, allowing researchers to study the detailed structure and organization of biological tissues at the cellular and subcellular level.
Microtomy is an essential tool in fields such as histology, pathology, and developmental biology, enabling in-depth exploration of tissue morphology and facilitating the diagnosis of various medical conditions.
The process of microtomy requires careful sample preparation, instrument calibration, and sectioning technique to ensure high-quality, reproducible results.
Unlocking the power of microtomy can lead to groundbreaking discoveries and advancements in biomedical research and clinical practice.
Most cited protocols related to «Microtomy»
Acid Hybridizations, Nucleic
Adult
Brain
Cells
Colorimetry
DAPI
Fluorescence
Gene Expression
Genetic Background
Immunoglobulins
Institutional Animal Care and Use Committees
Mice, Inbred C57BL
Mice, Laboratory
Microscopy
Microtomy
Proteins
Tissues
Freshly isolated and cultivated skin samples were harvested at time points, as indicated in the figures, and fixed immediately in 7.5% paraformaldehyde (SAV Liquid Production, Flintsbach am Inn, Germany). Fixed tissues were embedded in paraffin, sectioned (5 µm) (Microtome HM 335 E–Microm, GMI, USA) and stained with H&E solution according to standardized protocols.
Full text: Click here
Microtomy
Paraffin Embedding
paraform
Skin
Tissues
Postnatal day 30 (P30) TWI mice and their WT littermates (5 for each experimental group processed in 5 different experimental sessions, every TWI with its WT littermate) and one P15 TWI mouse versus its WT littermate were perfused with a fixative solution (4% paraformaldehyde and 0.1%–1%–2.5% glutaraldehyde in phosphate buffer, pH 7.4). Sciatic nerves, spinal cords and gastrocnemius muscles were dissected and post-fixed for 4 hours at room temperature in the same fixative solution.
Spinal cords were dissected in the lumbar region, isolating four 1-mm-thick sections in the lumbar enlargement region and the gastrocnemius muscles were cut in small portions, approximately 1 mm3 in volume. Sciatic nerves were processed without further sectioning.
The selected tissues were further treated for epoxy resin embedding as previously described43 . Briefly, the samples were deeper fixed in 2–2.5% glutaraldehyde in cacodylate buffer (0.1 M, pH 7.4). After rinsing, specimens were post-fixed with osmium tetroxide (1%)-potassium ferricyanide (1%) in cacodylate buffer, rinsed again, en bloc stained with 3% uranyl acetate in ethanol, dehydrated and embedded in epoxy resin, that was baked for 48 h at 60 °C. Thin sections were obtained with an ultramicrotome (UC7, Leica Microsystems, Vienna, Austria) and collected on G300Cu grids (EMS). Finally, sections were examined with a Zeiss LIBRA 120 plus transmission electron microscope equipped with an in-column omega filter.
Spinal cords were dissected in the lumbar region, isolating four 1-mm-thick sections in the lumbar enlargement region and the gastrocnemius muscles were cut in small portions, approximately 1 mm3 in volume. Sciatic nerves were processed without further sectioning.
The selected tissues were further treated for epoxy resin embedding as previously described43 . Briefly, the samples were deeper fixed in 2–2.5% glutaraldehyde in cacodylate buffer (0.1 M, pH 7.4). After rinsing, specimens were post-fixed with osmium tetroxide (1%)-potassium ferricyanide (1%) in cacodylate buffer, rinsed again, en bloc stained with 3% uranyl acetate in ethanol, dehydrated and embedded in epoxy resin, that was baked for 48 h at 60 °C. Thin sections were obtained with an ultramicrotome (UC7, Leica Microsystems, Vienna, Austria) and collected on G300Cu grids (EMS). Finally, sections were examined with a Zeiss LIBRA 120 plus transmission electron microscope equipped with an in-column omega filter.
Full text: Click here
Buffers
Cacodylate
Epoxy Resins
Ethanol
Fixatives
Glutaral
Hypertrophy
Lumbar Region
Mice, House
Microtomy
Muscle, Gastrocnemius
Osmium Tetroxide
paraform
Phosphates
potassium ferricyanide
Sciatic Nerve
Spinal Cord
Tissues
Transmission Electron Microscopy
Ultramicrotomy
uranyl acetate
Animals, Transgenic
Antibodies
Brain
Formalin
Immunohistochemistry
Internal Ribosome Entry Sites
LacZ Genes
Mice, Laboratory
Mice, Transgenic
Microtomy
Rosa
Spinal Cord
Animals
Animals, Laboratory
Brain
Cells
Helix (Snails)
Microscopy, Confocal
Microtomy
paraform
PEGDMA Hydrogel
Single-Cell Gene Expression Analysis
Tissues
Most recents protocols related to «Microtomy»
All specimens used for sequencing and experimentation were collected from Yunnan Province, China. All vouchers are stored in the Herpetological Museum of the Chengdu Institute of Biology, Chinese Academy of Sciences.
For the TEM experiments, two fresh skin samples (1cm×1cm) per color morph from the Asian vine snake were collected. The samples were then cut into small pieces (1 mm3) in fixative. The tissue blocks were transferred to an Eppendorf tube with fresh TEM fixative for further fixation, then washed using 0.1 M PB (pH 7.4) three times (15 min each). The samples were dehydrated in an increasing ethanol series at room temperature, followed by two changes of acetone and transfer to resin for embedding. The resin blocks were cut to 60–80-nm slices on an ultra-microtome (Leica, UC7), and the ultra-thin sections were put onto the 150-mesh cuprum grids. The cuprum grids were then stained with 2% uranium acetate-saturated alcohol solution and 2.6% lead citrate, respectively. Finally, the cuprum grids were observed under a TEM (Hitachi, HT7800/HT7700) and imaged.
For the TEM experiments, two fresh skin samples (1cm×1cm) per color morph from the Asian vine snake were collected. The samples were then cut into small pieces (1 mm3) in fixative. The tissue blocks were transferred to an Eppendorf tube with fresh TEM fixative for further fixation, then washed using 0.1 M PB (pH 7.4) three times (15 min each). The samples were dehydrated in an increasing ethanol series at room temperature, followed by two changes of acetone and transfer to resin for embedding. The resin blocks were cut to 60–80-nm slices on an ultra-microtome (Leica, UC7), and the ultra-thin sections were put onto the 150-mesh cuprum grids. The cuprum grids were then stained with 2% uranium acetate-saturated alcohol solution and 2.6% lead citrate, respectively. Finally, the cuprum grids were observed under a TEM (Hitachi, HT7800/HT7700) and imaged.
Full text: Click here
Acetone
Asian Persons
Chinese
Citrates
Copper
Ethanol
Fixatives
Microtomy
Morphine
Resins, Plant
Skin
Snakes
Tissues
uranyl acetate
Protocol full text hidden due to copyright restrictions
Open the protocol to access the free full text link
Animals
Brain
Cortex, Cerebral
Ferrets
Microscopy
Microtomy
Oxidase, Cytochrome-c
Tissues
White Matter
Protocol full text hidden due to copyright restrictions
Open the protocol to access the free full text link
Antibodies
Antigens
Calvaria
CTSK protein, human
Microtomy
Paraffin Embedding
Serum Albumin, Bovine
Sodium Citrate
Tissues
The ablated veins harvested after autopsy were irrigated with normal saline. They were immersed and shaded in a 2% 2,3,5-triphenyltetrazolium chloride (TTC) (Sigma) solution and incubated for 1 hour at 40 ℃. After the staining was completed, the veins were sectioned longitudinally and completely unfolded. The exposed ablated site was macroscopically checked, and photographs were taken.
TTC-stained femoral/cephalic veins were fixed in 10% neutral-buffered formalin. Each fixed tissue was rinsed in tap water for 24 hours to completely remove the fixative from the tissue. For tissue dehydration, the tissue was gradually dehydrated using high-concentration ethanol of 70%–100%, and then a paraffin block was produced by clearing with xylene. The prepared block was cut to a thickness of 5 µm using a microtome to prepare slides. The slides were stained with H&E for microscopic evaluation.
Verifying the nonstained area in the vein subjected to TTC staining identified the surviving and damaged areas in the venous endothelium, making it easier to select the area to be examined under the microscope. The part that was not stained with TTC was assessed as the part where vein injury occurred through ablation.
The vessel injury score analyzed based on H&E staining was also used to objectively evaluate the ablating effect. Vessel injury scores were measured at 3 sites per harvested ablated vein. After scanning the entire tissue made of slides with a scanner, the damaged area was visually checked. This method was applied by modifying that of a previous study [3 (link)]. The criteria were assigned according to injury severity from 1 (least injury) to 4 (most injury): 1, endothelial cell coverage; 2, medial smooth muscle cell loss; 3, internal and external elastic lamina disruption; and 4, adventitia disruption. Scoring was comprehensively performed by a pathologist through evaluating the damaged area that each criterion had inflicted on the tissue.
TTC-stained femoral/cephalic veins were fixed in 10% neutral-buffered formalin. Each fixed tissue was rinsed in tap water for 24 hours to completely remove the fixative from the tissue. For tissue dehydration, the tissue was gradually dehydrated using high-concentration ethanol of 70%–100%, and then a paraffin block was produced by clearing with xylene. The prepared block was cut to a thickness of 5 µm using a microtome to prepare slides. The slides were stained with H&E for microscopic evaluation.
Verifying the nonstained area in the vein subjected to TTC staining identified the surviving and damaged areas in the venous endothelium, making it easier to select the area to be examined under the microscope. The part that was not stained with TTC was assessed as the part where vein injury occurred through ablation.
The vessel injury score analyzed based on H&E staining was also used to objectively evaluate the ablating effect. Vessel injury scores were measured at 3 sites per harvested ablated vein. After scanning the entire tissue made of slides with a scanner, the damaged area was visually checked. This method was applied by modifying that of a previous study [3 (link)]. The criteria were assigned according to injury severity from 1 (least injury) to 4 (most injury): 1, endothelial cell coverage; 2, medial smooth muscle cell loss; 3, internal and external elastic lamina disruption; and 4, adventitia disruption. Scoring was comprehensively performed by a pathologist through evaluating the damaged area that each criterion had inflicted on the tissue.
Adventitia
Autopsy
Endothelial Cells
Endothelium
Ethanol
Fixatives
Formalin
Injuries
Microscopy
Microtomy
Myocytes, Smooth Muscle
Normal Saline
Paraffin
Pathologists
Tissues
triphenyltetrazolium chloride
Vascular System Injuries
Vein, Femoral
Veins
Xylene
According to the previous studies on avian species and a chick brain atlas (Kuenzel and Masson, 1988 ; Kang et al., 2009 (link); Kang et al., 2020 (link)), two major 5-HTergic regions in the brainstem, DRN and CRN, and VTA regions were dissected in a cryostat microtome. The dimensions of the dissected section are as follows: 2.5–3 mm (W) x 1–1.5 mm (H) x 2.5–3.0 mm (L) for DRN; 2–2.5 mm (W) x 1–1.2 mm (H) x 2.5–3.0 mm (L) for CRN; and 3–3.5 mm (W) x 2–3 mm (H) x 1–1.2 mm (L) for VTA. The thickness (W, H, and L) of the dissected brain tissue block was proportionally increased from young birds to older birds based on the brain size and structure. Inside the cryostat, the brain areas shown as rectangles were dissected from each flattened brain section using a scalpel handle and blade (#11) and were quickly transferred to TRIzol reagent and then stored at -80°C until total RNA extraction.
Full text: Click here
Aves
Brain
Brain Stem
Microtomy
Tissues
trizol
Top products related to «Microtomy»
Sourced in Germany, United States, Japan, China, United Kingdom, Italy, Israel, Australia, France, Canada, Switzerland, Austria, Brazil, Sweden
A Microtome is a precision instrument used to cut extremely thin sections of material, typically for microscopic examination. It operates by moving a sample through a sharp blade, producing uniform slices of the desired thickness. The core function of a Microtome is to enable the preparation of high-quality samples for various microscopic techniques.
Sourced in Germany, United States, Japan, Canada, Australia, Spain, Israel
The Freezing Microtome is a laboratory instrument used to prepare thin, uniform sections of frozen biological samples for microscopic examination. It accurately slices the frozen specimen into thin sections, allowing researchers to study the internal structure and composition of the sample.
Sourced in Germany, United States, China, Italy, United Kingdom, Japan, Brazil
The RM2235 is a rotary microtome designed for sectioning paraffin-embedded tissue samples. It features a vertical specimen feed and a high-quality steel knife or disposable blade holder. The RM2235 allows users to obtain consistent, high-quality tissue sections for further analysis and investigation.
Sourced in Germany, United States, China, Switzerland, Canada, France, Italy, Japan, United Kingdom
The RM2255 is a rotary microtome designed for sectioning a wide range of paraffin-embedded tissue samples. It features a vertical specimen feed with a stroke of 70 mm and a section thickness range of 0.5 to 100 μm. The RM2255 is equipped with a precision feed mechanism and a high-quality steel knife holder for consistent and accurate sectioning.
Sourced in United States, Germany, United Kingdom, China, Italy, France, Macao, Australia, Canada, Sao Tome and Principe, Japan, Switzerland, Spain, India, Poland, Belgium, Israel, Portugal, Singapore, Ireland, Austria, Denmark, Netherlands, Sweden, Czechia, Brazil
Paraformaldehyde is a white, crystalline solid compound that is a polymer of formaldehyde. It is commonly used as a fixative in histology and microscopy applications to preserve biological samples.
Sourced in Germany, United States, Japan, China, United Kingdom, Australia, Switzerland, France, Netherlands, Spain, Ireland
The Leica CM1950 is a cryostat designed for sectioning frozen tissue samples. It features a temperature range of -10°C to -35°C and a specimen size of up to 55 x 55 mm. The instrument is equipped with a motorized specimen feed and a high-performance cooling system.
Sourced in Germany, United States, France, United Kingdom, Israel, Japan, Switzerland, Canada, Belgium
The VT1000S is a vibratome, a precision instrument used for sectioning biological samples, such as tissues or organs, into thin slices for microscopic examination or further processing. The VT1000S provides consistent and accurate sectioning of samples, enabling researchers to obtain high-quality tissue sections for a variety of applications.
Sourced in Japan, United States, Germany, United Kingdom, China, France
The Hitachi H-7650 is a transmission electron microscope (TEM) designed for high-resolution imaging of materials. It provides a core function of nanoscale imaging and analysis of a wide range of samples.
Sourced in Japan, United States, Germany, Italy, Denmark, United Kingdom, Canada, France, China, Australia, Austria, Portugal, Belgium, Panama, Spain, Switzerland, Sweden, Poland
The BX51 microscope is an optical microscope designed for a variety of laboratory applications. It features a modular design and offers various illumination and observation methods to accommodate different sample types and research needs.
More about "Microtomy"
Microtomy: Unlocking the Secrets of Tissue Sectioning Microtomy, the art of precisely slicing biological samples into thin sections, is a fundamental technique in fields like histology, pathology, and developmental biology.
This meticulous process enables researchers to delve into the intricate structures and organizations of tissues at the cellular and subcellular level.
At the heart of microtomy lies the microtome, a specialized instrument designed to cut uniform, high-quality sections with remarkable accuracy.
From the manual rotary microtome to the automated freezing microtome (like the RM2235 and RM2255), these cutting-edge tools are essential for preparing samples for microscopic examination.
Proper sample preparation is crucial for successful microtomy.
Fixation with chemicals like paraformaldehyde (used in the CM1950 cryostat) helps preserve the tissue's structure, while dehydration and embedding techniques, such as those employed in the VT1000S vibrating microtome, ensure the samples are ready for sectioning.
Once the samples are prepared, the microtoming process begins.
Skilled technicians use instruments like the H-7650 ultramicrotome to carefully slice the tissue into thin, uniform sections, often just a few micrometers thick.
These sections are then stained and mounted on slides, ready for analysis under powerful microscopes like the BX51.
Unlocking the full potential of microtomy requires a deep understanding of the techniques, instruments, and sample preparation methods.
By leveraging the latest advancements in this field, researchers can push the boundaries of biomedical discovery and enable groundbreaking insights into the intricate world of cellular and subcellular biology.
This meticulous process enables researchers to delve into the intricate structures and organizations of tissues at the cellular and subcellular level.
At the heart of microtomy lies the microtome, a specialized instrument designed to cut uniform, high-quality sections with remarkable accuracy.
From the manual rotary microtome to the automated freezing microtome (like the RM2235 and RM2255), these cutting-edge tools are essential for preparing samples for microscopic examination.
Proper sample preparation is crucial for successful microtomy.
Fixation with chemicals like paraformaldehyde (used in the CM1950 cryostat) helps preserve the tissue's structure, while dehydration and embedding techniques, such as those employed in the VT1000S vibrating microtome, ensure the samples are ready for sectioning.
Once the samples are prepared, the microtoming process begins.
Skilled technicians use instruments like the H-7650 ultramicrotome to carefully slice the tissue into thin, uniform sections, often just a few micrometers thick.
These sections are then stained and mounted on slides, ready for analysis under powerful microscopes like the BX51.
Unlocking the full potential of microtomy requires a deep understanding of the techniques, instruments, and sample preparation methods.
By leveraging the latest advancements in this field, researchers can push the boundaries of biomedical discovery and enable groundbreaking insights into the intricate world of cellular and subcellular biology.