scholarly journals PB-02Universal design of a diamond knife for ultra-thin sectioning

Microscopy ◽  
2018 ◽  
Vol 67 (suppl_2) ◽  
pp. i32-i32
Author(s):  
Hiroshi Takase ◽  
Makoto Sugiura-Nakazato ◽  
Kaori Yamaguchi
Keyword(s):  
Thin Sectioning ◽  
Diamond Knife

Practical methods for TEM

1996 ◽  
Vol 54 ◽  
pp. 628-629
Author(s):  
K. Chien ◽  
R. Gonzalez ◽  
R. C. Heusser ◽  
H. Shiroishi ◽  
M. L. Heathershaw
Keyword(s):  
Needle Biopsy ◽  
Long Distance ◽  
Hot Plate ◽  
Razor Blade ◽  
Thin Sectioning ◽  
Block Face ◽  
Single Block ◽  
The Difference ◽  
Glass Knife ◽  
Diamond Knife

Multiple sectioning: Two or more core specimens from a kidney needle biopsy can be embedded in single block using an 8-facet silicone rubber embedding mold. Tissue blocks are softened by placing them face down on an 85°C hot plate for one minute and then trimming with a Teflon coated razor blade. This prevents the block face from being deeply fractured which can affect section quality. If glomeruli are found within different cores in a single block, the block face can be retrimmed into multiple mesas and simultaneously thin-sectioned. This method greatly increases the efficiency of both thick and thin sectioning which is extremely helpful when handling more than 1600 cases of renal biopsies in our EM laboratory each year.Knife Spacer: Due to the difference in thickness between a glass and diamond knife, and also depending on which portion of the diamond knife is being used, the knife stage has to be moved laterally over a long distance when changing from one knife type to the other. Placing a L-shaped spacer, 3 to 5 mm in thickness, on the left side of the stage when sectioning a glass knife will greatly reduce this lateral adjustment.

An inexpensive water volume control device for ultramicrotomy

1990 ◽  
Vol 48 (3) ◽  
pp. 732-733
Author(s):  
E.G. Kokko ◽  
B. Gowen ◽  
C.R. Jahnke
Keyword(s):  
Three Dimensional ◽  
Control Device ◽  
Volume Control ◽  
Water Volume ◽  
Thin Sections ◽  
Precise Control ◽  
Transmission Electron ◽  
Thin Sectioning ◽  
Main Disadvantage ◽  
Diamond Knife

Numerous techniques and related devices have been reported for preparing serial thin-sections with an ultramicrotome for transmission electron microscopy. The section pickup method, a continuous serial thin-sectioning technique developed by Farenbach (1984), is helpful for: (a) three-dimensional EM reconstruction (Todd et.al. 1989) and (Farenbach 1984), (b)section thickness determinations of thick (LM) or thin (EM) sections using the “section piling - resectioning” technique of Sakai (1980), (c) insuring that important or rare EM sections (such as from a reembedded 0.5 urn LM section) are picked up, or (d) for routine section pickup on formvar-coated slot grids. Farenbach’s technique makes use of a pedestal, located within a diamond knife trough, to hold the slot grids. It calls for a peristaltic pump to maintain precise control of the water level within the trough. The main disadvantage of his technique is the relatively high cost (approx. $ 500 US) for a pump capable of these accurate and two-way flow adjustments of small volumes of liquid.

A Rapid Diamond Knife Alignment Procedure

1980 ◽  
Vol 38 ◽  
pp. 648-649
Author(s):  
Robert C. Heusser
Keyword(s):  
Tissue Culture ◽  
Restricted Range ◽  
Alignment Procedure ◽  
Critical Orientation ◽  
Thin Sectioning ◽  
Flat Embedding ◽  
Diamond Knife

The purpose of this paper is to present a rapid and accurate means of accomplishing two of the more difficult manipulations in diamond knife thin sectioning, namely precise block align¬ment and advancement of the knife to a point of contact. In routine thin sectioning, the desire for serial continuity from a previously sectioned block is universal. Specialized embedding procedures such as: tissue culture monolayers, re-sectioning of large plastic sections and flat embedding molds for specially oriented tissue require extremely accurate block ori¬entation due to an unusually restricted range of tissue thick¬ness and critical orientation.

Ultrastructure of Some Planktonic Formainifera

1974 ◽  
Vol 32 ◽  
pp. 190-191
Author(s):  
William H. Zucker
Keyword(s):  
Cell Biology ◽  
Water Mass ◽  
Planktonic Foraminifera ◽  
Uranyl Acetate ◽  
Ethanol Dehydration ◽  
Globigerinoides Ruber ◽  
Light Microscope Level ◽  
Globigerina Bulloides ◽  
Glutaraldehyde Solution ◽  
Diamond Knife

Planktonic foraminifera are widely-distributed and abundant zooplankters. They are significant as water mass indicators and provide evidence of paleotemperatures and events which occurred during Pleistocene glaciation. In spite of their ecological and paleological significance, little is known of their cell biology. There are few cytological studies of these organisms at the light microscope level and some recent reports of their ultrastructure.Specimens of Globigerinoides ruber, Globigerina bulloides, Globigerinoides conglobatus and Globigerinita glutinata were collected in Bermuda waters and fixed in a cold cacodylate-buffered 6% glutaraldehyde solution for two hours. They were then rinsed, post-fixed in Palade's fluid, rinsed again and stained with uranyl acetate. This was followed by graded ethanol dehydration, during which they were identified and picked clean of debris. The specimens were finally embedded in Epon 812 by placing each organism in a separate BEEM capsule. After sectioning with a diamond knife, stained sections were viewed in a Philips 200 electron microscope.

Preparatory Procedures for Electron Microscopic Analysis at the Molecular Level of Resolution

1978 ◽  
Vol 36 (3) ◽  
pp. 516-520
Author(s):  
F.J. Sjostrand
Keyword(s):  
Electron Microscope ◽  
Molecular Level ◽  
Microscopic Analysis ◽  
Resolving Power ◽  
Cellular Structures ◽  
Electron Microscopic ◽  
Systematic Analysis ◽  
Technical Developments ◽  
Thin Sectioning ◽  
Obvious Reason

In the 1940's and 1950's electron microscopy conferences were attended with everybody interested in learning about the latest technical developments for one very obvious reason. There was the electron microscope with its outstanding performance but nobody could make very much use of it because we were lacking proper techniques to prepare biological specimens. The development of the thin sectioning technique with its perfectioning in 1952 changed the situation and systematic analysis of the structure of cells could now be pursued. Since then electron microscopists have in general become satisfied with the level of resolution at which cellular structures can be analyzed when applying this technique. There has been little interest in trying to push the limit of resolution closer to that determined by the resolving power of the electron microscope.

Lead aspartate: a new en bloc stain for electron microscopy

1978 ◽  
Vol 36 (2) ◽  
pp. 34-35
Author(s):  
J.R. Walton
Keyword(s):  
Electron Microscopy ◽  
Calcium Ion ◽  
Enzyme Reaction ◽  
Lead Citrate ◽  
Reaction Products ◽  
En Bloc ◽  
Thin Sections ◽  
Lead Salts ◽  
Thin Sectioning ◽  
High Alkalinity

In electron microscopy, lead is the metal most widely used for enhancing specimen contrast. Lead citrate requires a pH of 12 to stain thin sections of epoxy-embedded material rapidly and intensively. However, this high alkalinity tends to leach out enzyme reaction products, making lead citrate unsuitable for many cytochemical studies. Substitution of the chelator aspartate for citrate allows staining to be carried out at pH 6 or 7 without apparent effect on cytochemical products. Moreover, due to the low, controlled level of free lead ions, contamination-free staining can be carried out en bloc, prior to dehydration and embedding. En bloc use of lead aspartate permits the grid-staining step to be bypassed, allowing samples to be examined immediately after thin-sectioning.Procedures. To prevent precipitation of lead salts, double- or glass-distilled H20 used in the stain and rinses should be boiled to drive off carbon dioxide and glassware should be carefully rinsed to remove any persisting traces of calcium ion.

Albumins as Embedding Media for Electron Microscopy

1969 ◽  
Vol 27 ◽  
pp. 422-423 ◽  
Author(s):  
J. L. Farrant ◽  
J. D. McLean
Keyword(s):  
Electron Microscopy ◽  
Electron Microscope ◽  
High Temperature ◽  
Biological Material ◽  
Globular Proteins ◽  
The Past ◽  
Antibody Staining ◽  
Thin Sectioning ◽  
Embedding Medium

For electron microscope techniques such as ferritin-labeled antibody staining it would be advantageous to have available a simple means of thin sectioning biological material without subjecting it to lipid solvents, impregnation with plastic monomers and their subsequent polymerization. With this aim in view we have re-examined the use of protein as an embedding medium. Gelatin which has been used in the past is not very satisfactory both because of its fibrous nature and the high temperature necessary to keep its solutions fluid. We have found that globular proteins such as the serum and egg albumins can be cross-linked so as to yield blocks which are suitable for ultrathin sectioning.

Effects of Vibration on the Preparation of Ultrathin Sections

1973 ◽  
Vol 31 ◽  
pp. 358-359
Author(s):  
Joseph M. Blum ◽  
Edward P. Gargiulo ◽  
J. R. Sawers
Keyword(s):  
Cutting Speed ◽  
Ground Level ◽  
Environmental Vibration ◽  
Block Face ◽  
Ultrathin Sections ◽  
Embedding Medium ◽  
Thickness Variations ◽  
Building Walls ◽  
Cutting Direction ◽  
Diamond Knife

It is now well-known that chatter (Figure 1) is caused by vibration between the microtome arm and the diamond knife. It is usually observed as a cyclical variation in “optical” density of an electron micrograph due to sample thickness variations perpendicular to the cutting direction. This vibration might be induced by using too large a block face, too large a clearance angle, excessive cutting speed, non-uniform embedding medium or microtome vibration. Another prominent cause is environmental vibration caused by inadequate building construction. Microtomes should be installed on firm, solid floors. The best floors are thick, ground-level concrete pads poured over a sand bed and isolated from the building walls. Even when these precautions are followed, we recommend an additional isolation pad placed on the top of a sturdy table.

Structural modifications of intercellular junctions.

1978 ◽  
Vol 36 (2) ◽  
pp. 330-331
Author(s):  
J. Metz ◽  
M. Merlo ◽  
W. G. Forssmann
Keyword(s):  
Gap Junctions ◽  
Intercellular Junctions ◽  
Cell Isolation ◽  
Extrahepatic Cholestasis ◽  
Structural Modifications ◽  
Pancreatic Duct Ligation ◽  
Pancreatic Cells ◽  
Duct Ligation ◽  
Thin Sectioning ◽  
And Function

Structure and function of intercellular junctions were studied under the electronmicroscope using conventional thin sectioning and freeze-etch replicas. Alterations of tight and gap junctions were analyzed 1. of exocrine pancreatic cells under cell isolation conditions and pancreatic duct ligation and 2. of hepatocytes during extrahepatic cholestasis.During the different steps of cell isolation of exocrine pancreatic cells, gradual changes of tight and gap junctions were observed. Tight junctions, which formed belt-like structures around the apex of control acinar cells in situ, subsequently diminished, became interrupted and were concentrated into macular areas (Fig. 1). Aggregations of membrane associated particles, which looked similar to gap junctions, were intermixed within tight junctional areas (Fig. 1). These structures continously disappeared in the last stages of the isolation procedure. The intercellular junctions were finally separated without destroying the integrity of the cell membrane, which was confirmed with porcion yellow, lanthanum chloride and horse radish peroxidase.

Surface Structure in Microtomed Sections Caused by Glass and Diamond Knives

1971 ◽  
Vol 29 ◽  
pp. 456-457
Author(s):  
J. Temple Black ◽  
William G. Boldosser
Keyword(s):  
Plastic Deformation ◽  
Epoxy Resin ◽  
Carbon Coating ◽  
Surface Damage ◽  
Body Angle ◽  
Normal Manner ◽  
Bottom Side ◽  
Cutting Direction ◽  
Made In ◽  
Diamond Knife

Ultramicrotomy produces plastic deformation in the surfaces of microtomed TEM specimens which can not generally be observed unless special preparations are made. In this study, a typical biological composite of tissue (infundibular thoracic attachment) infiltrated in the normal manner with an embedding epoxy resin (Epon 812 in a 60/40 mixture) was microtomed with glass and diamond knives, both with 45 degree body angle. Sectioning was done in Portor Blum Mt-2 and Mt-1 microtomes. Sections were collected on formvar coated grids so that both the top side and the bottom side of the sections could be examined. Sections were then placed in a vacuum evaporator and self-shadowed with carbon. Some were chromium shadowed at a 30 degree angle. The sections were then examined in a Phillips 300 TEM at 60kv.Carbon coating (C) or carbon coating with chrom shadowing (C-Ch) makes in effect, single stage replicas of the surfaces of the sections and thus allows the damage in the surfaces to be observable in the TEM. Figure 1 (see key to figures) shows the bottom side of a diamond knife section, carbon self-shadowed and chrom shadowed perpendicular to the cutting direction. Very fine knife marks and surface damage can be observed.

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