scholarly journals Vapor Coating: A Simple, Economical Procedure for Preparing Difficult Specimens for Scanning Electron Microscopy

2007 ◽  
Vol 15 (3) ◽  
pp. 44-45 ◽  
Author(s):  
E. Ann Ellis ◽  
Michael W. Pendleton
Keyword(s):  
Osmium Tetroxide ◽  
Petri Dish ◽  
Specimen Preparation ◽  
Fume Hood ◽  
Imaging Center ◽  
User Facility ◽  
Small Container ◽  
Set Up ◽  
Preparation Techniques ◽  
Scanning Electron

The Microscopy and Imaging Center at Texas A&M University is a multi-user facility involved with preparation and analysis of many different biological and materials sciences projects. Vapor stabilization and coating is an important part of our specimen preparation methodology for difficult biological and materials, especially polymer, samples. The procedure for all our vapor preparation techniques is done in a simple, economical apparatus set up in a properly functioning fume hood with a flow rate of at least 100 ft/min (Fig. 1). The apparatus is made from a glass petri dish or a glass petri dish for the bottom and an appropriate size beaker for the top. Specimens, mounted on stubs, are placed inside the chamber and the fixative (osmium tetroxide, ruthenium tetroxide or acrolein) is placed in a small container (plastic bottle cap) near the specimens.

Electron Microscopic Observation of Biological Specimens in Their Native State by Employing Cryogenic Techniques

1972 ◽  
Vol 30 ◽  
pp. 410-411 ◽  
Author(s):  
Tokio Nei ◽  
Haruo Yotsumoto ◽  
Yoichi Hasegawa ◽  
Yuji Nagasawa
Keyword(s):  
Electron Microscopic Observation ◽  
Surface Structures ◽  
Specimen Preparation ◽  
Native State ◽  
Electron Microscopic ◽  
Biological Specimen ◽  
Specimen Holder ◽  
Cold Stage ◽  
Preparation Techniques ◽  
Scanning Electron

In order to observe biological specimens in their native state, that is, still containing their water content, various methods of specimen preparation have been used, the principal two of which are the chamber method and the freeze method.Using its recently developed cold stage for installation in the pre-evacuation chamber of a scanning electron microscope, we have succeeded in directly observing a biological specimen in its frozen state without the need for such conventional specimen preparation techniques as drying and metallic vacuum evaporation. (Echlin, too, has reported on the observation of surface structures using the same freeze method.)In the experiment referred to herein, a small sliced specimen was place in the specimen holder. After it was rapidly frozen by freon cooled with liquid nitrogen, it was inserted into the cold stage of the specimen chamber.

Comparison of freeze-fractured yeast replicas using conventional TEM and low-voltage field-emission SEM

1989 ◽  
Vol 47 ◽  
pp. 74-75
Author(s):  
William P. Wergin ◽  
Eric F. Erbe ◽  
Terrence W. Reilly
Keyword(s):  
Electron Microscope ◽  
Scanning Electron Microscope ◽  
Field Emission ◽  
Low Voltage ◽  
Objective Lens ◽  
Specimen Preparation ◽  
Lens System ◽  
Air Drying ◽  
Preparation Techniques ◽  
Scanning Electron

Although the first commercial scanning electron microscope (SEM) was introduced in 1965, the limited resolution and the lack of preparation techniques initially confined biological observations to relatively low magnification images showing anatomical surface features of samples that withstood the artifacts associated with air drying. As the design of instrumentation improved and the techniques for specimen preparation developed, the SEM allowed biologists to gain additional insights not only on the external features of samples but on the internal structure of tissues as well. By 1985, the resolution of the conventional SEM had reached 3 - 5 nm; however most biological samples still required a conductive coating of 20 - 30 nm that prevented investigators from approaching the level of information that was available with various TEM techniques. Recently, a new SEM design combined a condenser-objective lens system with a field emission electron source.

A Model Outreach Program for Teaching Scanning Electron Microscopy Technology: Successes and Failures

1998 ◽  
Vol 4 (S2) ◽  
pp. 40-41
Author(s):  
N.R. Smith ◽  
R.A. Quinta
Keyword(s):  
Electron Microscopy ◽  
Community College ◽  
Scanning Electron Microscopy ◽  
Outreach Program ◽  
Specimen Preparation ◽  
State University ◽  
Learning Module ◽  
Imaging Center ◽  
Effective Use ◽  
Scanning Electron

A partnership has developed between the Microscope and Graphic Imaging Center (MAGIC) at California State University, Hayward and Ohlone Community College. The purpose of the collaboration is to develop a program to allow community college students to gain experience in preparing and viewing samples using scanning electron microscopy technology. The learning module involves students from the Ohlone College Biology Majors Program and student mentors from CSUH. An additional component is the introduction of under-represented students into a Biology Fellowship Program in which they also participate in the SEM learning module. Participants for these programs are selected on the basis of their interest and how this experience will benefit them as expressed in a one-page written essay. Ten students are selected to participate in the programs.The objectives of the learning module are to: 1) learn specimen preparation techniques and develop skills in SEM technology; 2) gain hands-on experience and develop some laboratory skills necessary for effective use of a SEM in studying biological specimens; 3) share the experience gained with peers at their home institution.

Osmium tetroxide vapor fixation for preserving fungal structures for SEM

1990 ◽  
Vol 48 (3) ◽  
pp. 748-749
Author(s):  
L.R. Tiedt
Keyword(s):  
Room Temperature ◽  
Osmium Tetroxide ◽  
Petri Dish ◽  
Usual Method ◽  
Water Soluble ◽  
Fungal Morphology ◽  
Drying Method ◽  
Critical Point Drying ◽  
Inner Surface ◽  
Scanning Electron

Scanning electron microscopy has provided valuable information in studies of fungal morphology and ontogeny, but undisturbed preservation of sporulating structures is often difficult.In this study of sporodochia development in Fusarium spp. in the section Liseola, the usual method of aqueous fixation, dehydration and the subsequent critical-point drying method, could not be used. Macroconidia forms sporodochia by means of water soluble slime and when aqueous solutions come in contact with the sporodochia, the conidia become scattered. To preserve these morphological structures, the samples had to be vapour fixed with osmium tetroxide. Sample blocks were excised from the sporulating cultures and carefully transferred to the inner surface of a Petri dish cover. The cover with several adherent agar blocks were inverted over a Petri dish bottom containing several drops of 2% osmium tetroxide. The Petri dish was left at room temperature for 24h. The Petri dish cover with the fixed samples was transferred to a clean Petri dish bottom and left to air dry.

Visualization of the Structure of the Rat Oocyte by Scanning Electron Microscopy

1977 ◽  
Vol 35 ◽  
pp. 608-609
Author(s):  
P. Bagavandoss
Keyword(s):  
Electron Microscopy ◽  
Scanning Electron Microscopy ◽  
Osmium Tetroxide ◽  
Transmission Electron ◽  
Electron Microscopes ◽  
Preparation Techniques ◽  
Scanning Electron ◽  
Single Ovary ◽  
Sem Studies

Mammalian oocytes have been well studied with the light and the transmission electron microscopes(1,2). Glutaraldehyde and osmium tetroxide, which offer excellent fixation have certain limitations when used for SEM studies of rat ovaries. These fixatives require very small specimens for penetration, but such small specimens provide only a few intact oocytes for SEM observations. However, for meaningful observation of the changes brought about by various hormone treatments on the oocytes, a maximum number of oocytes must be available frcm a single ovary. Also, aldehyde and OSO4 fixed tissues are not easy to cut into smooth halves. This report cctnmuni- cates preparation techniques that yield useful SEM of the oocyte and associated structures of rat ovaries.Rat ovaries were dissected out and immediately placed in Bouin's fluid for 24 hours at rocm temperature. After fixation each ovary was cut into two halves with a sharp blade and kept in 70% alcohol overnight.

scholarly journals Freezing Techniques: History, Comparisons, and Applications

2008 ◽  
Vol 16 (5) ◽  
pp. 12-17 ◽  
Author(s):  
Bill Graham ◽  
Jotham R. Austin ◽  
Andres Kaech ◽  
John E. Heuser
Keyword(s):  
Electron Microscopy ◽  
Osmium Tetroxide ◽  
Basic Problem ◽  
High Vacuum ◽  
Harsh Environment ◽  
Specimen Preparation ◽  
Biological Processes ◽  
Thin Sectioning ◽  
The Cost ◽  
Preparation Techniques

Specimen preparation techniques have evolved hand in hand with microscopy since the first microscopes. Since the introduction of the first Electron Microscope (EM) in the 1930’s, the basic problem with biological electron microscopy has been to preserve the structure of soft condensed, hydrated matter (e.g. tissues, cells, proteins, etc.) so that they can be viewed in the harsh environment of the electron microscope's high vacuum and ionizing radiation. For this, cells must be “fixed” with chemical cross-linkers, commonly glutaraldehyde, formaldehyde or some combination of both, stained with heavy metals (osmium tetroxide that provides contrast of biological components), dehydrated with an organic solvent, and infiltrated with a resin for eventual thin-sectioning. Only then can it be viewed with the EM. Such treatment with chemical fixatives and stains remains the standard approaches to arrest biological processes in cells or tissues, but at the cost of introducing clearly recognizable artifacts.

Texture and Artifacts in SEM Observations Due to Vapor Coating

1972 ◽  
Vol 30 ◽  
pp. 406-407
Author(s):  
G. G. Paulson ◽  
R. W. Pierce
Keyword(s):  
Electron Microscopy ◽  
Noble Metals ◽  
Specimen Preparation ◽  
Polystyrene Spheres ◽  
Final Thickness ◽  
Quantitative Measurements ◽  
Glass Slides ◽  
Beam Currents ◽  
Preparation Techniques ◽  
Scanning Electron

Specimen preparation techniques for scanning electron microscopy have not necessarily kept pace with the advances in instrumentation, especially in fields where quantitative measurements are important. Even though many operators have found that nonconducting surfaces can often be examined directly with low voltages and small beam currents, most commonly these surfaces are vapor-coated with 100 to 600 A of a noble metal alloy. More often than not, the coating procedures involve a bit of guesswork as to the final thickness and structure of the vapor coating. The purpose of this research was to illustrate the importance of close control on coating thickness and uniformity in order to minimize artifacts and distortion. Several noble metals and alloys were studied. The results for gold coatings are reported.Polystyrene spheres of statistically uniform diameters were dispersed on glass slides. The slides were rotated as they were coated by thermal deposition of Au from a W boat at a pressure of 2 x 10-5 torr in a standard oil diffusion pumped vapor coater.

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