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This lab procedure describes a method for preparing permanent microscope slide mounts
using Cargille's MeltMount. We use this method when preparing particle samples which are to be retained indefinitely, such as
for reference slides or where the samples may be required in environmental or mold litigation. - (Photographs are available -To Be Added or on request).
Required materials for permanent mount microscope slide preparation using Cargille Meltmount
Cargille Meltmount - Cargille Laboratories www.cargille.com or from secondary suppliers such as www.2spi.com permanent slide mounting media (replacement for Canada Balsam and Aroclor) ND=1.539 @25degC.
Hot plate, Slides, Colored paper, Needle, dropper, or implement for mountant transfer to slide.
Detailed instructions available from Cargille regarding slide preparation using Meltmount. Remember to work in a well-ventilated area, to be careful of hot surfaces and chemicals, and to wear
polyethylene gloves to avoid getting solvents or mountant on skin surfaces.
For in-lab use Cargille describes a procedure for sample preparation in which the sample is placed on the slide, the cover slip installed, and meltmount is introduced to the corners of the cover slip using a dropper.
Cargille also describes a method of pre-coating slides with mountant for subsequent use in the field.
Because the container of Meltmount, as received from Cargille takes some time to heat completely for in-lab use, and because I work with a
variety of field sample materials which can be slow following this routine, I have adapted Cargille's "Pressure method," intended for field use, to a combination of lab and field work as I describe below.
Cut out colored paper to approximately 2" x 4" and place a piece on the hot plate. The paper will be used to provide added contrast to enable easy view of the area of the slide which has been coated with mountant. Draw on the paper an outline of one of your slides and then mark within
that outline the area where you will ultimately want to place a cover slip on the slide.
This permits production of uniform slides and speed
work at the microscope by placing each sample in the same area on the microscope stage. Prepare several of these colored slips so that you can replace
them when soiled or wet.
Heat the mountant to the proper working temperature. Heat surface to
60-70 C - the meltmount will be liquid but not smoking if it's at the easiest working temperature. I heat the meltmount in its container to a lower temperature
Place the colored, marked paper on the hot plate.
Place the slide on the colored paper over the outlined area.
Pick up a droplet of Cargille permanent slide mountant with your implement of choice. I use a dental tool to
pick up a droplet of mountant, let it solidify on the end of the tool (takes only a second), then I use the melted drop on the tool to "paint"
the target area on the slide as described below. Too much mountant may produce a messy slide with a lot of excess mountant to be cleaned off
later. Too little mountant may not adequately handle large particles, leaving a tilted cover slip.
When the slide has been "painted" with mountant move it to another area of the hot plate where it should remain until the mountant has been heated to a level, uniform state. Meanwhile you can be working on coating subsequent slides.
When the mountant is level on the slide remove the slide to a cool, not cold, surface and let the slide cool to room temperature.
Label the slide with blank labels at this stage to make it easy later to quickly see which side has the mountant.
Store prepared microscope slides flat, mountant upright. I use a plastic slide box.
To use previously prepared Cargille Meltmount slides in the field or lab.
Transfer the sample to the area of mountant either by scraping the sample on to the slide, transferring by tape, or other method.
Heat the slide briefly over a match or alcohol lamp.
Install the cover slip.
Permit the slide to stabilize: Cargille recommends pressing the slide between waxed paper and weighted
by a book overnight. When I used an alcohol lamp to heat the slide before installing the cover slip this was not necessary.
Clean off any excess mountant with a razor blade [after the slide has cooled or stabilized] and/or an appropriate
solvent - Cargille recommends xylene or toluene. I had some success also using hexane followed by isopropyl alcohol for final cleanup. [Caution: hexane is a toxic carcinogen.]
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Kansas State University, department of plant pathology, extension plant pathology web page on wheat rust fungus: see http://www.oznet.ksu.edu/path-ext/factSheets/Wheat/Wheat%20Leaf%20Rust.asp
"A Brief Guide to Mold, Moisture, and Your Home",
U.S. Environmental Protection Agency US EPA - includes basic advice for building owners, occupants, and mold cleanup operations. See http://www.epa.gov/mold/moldguide.htm
US EPA - Mold Remediation in Schools and Commercial Building [Copy on file at /sickhouse/EPA_Mold_Remediation_in_Schools.pdf ] - US EPA
US EPA - Una Breva Guia a Moho - Hongo [Copy on file as /sickhouse/EPA_Moho_Guia_sp.pdf - en Espanol
Allergies, Allergens, Allergy Testing in Buildings - References & Products
Allergen Tests in Buildings advice about how to test, what to look for, in evaluating the level of dog, cat, or other animal allergens in a building
"IgG Food Allergy Testing by ELISA/EIA, What do they really tell us?" Sheryl B. Miller, MT (ASCP), PhD, Clinical Laboratory Director, Bastyr University Natural Health Clinic - ELISA testing accuracy: Here is an example of Miller's critique of ELISA
http://www.betterhealthusa.com/public/282.cfm - Townsend Letter for Doctors and Patients
The critique included in that article raises compelling questions about IgG testing assays, which prompts our interest in actually screening for the presence of high levels of particles that could carry allergens - dog dander or cat dander in the case at hand.
http://www.tldp.com/issue/174/IgG%20Food%20Allergy.html contains similar criticism in another venue but interestingly by the same author, Sheryl Miller. Sheryl Miller, MT (ASCP), PhD, is an Immunologist and Associate Professor of Basic and Medical Sciences at Bastyr University in Bothell, Washington. She is also the Laboratory Director of the Bastyr Natural Health Clinic Laboratory.
Allergens: Testing for the level of exposure to animal allergens is discussed at http://www.animalhealthchannel.com/animalallergy/diagnosis.shtml (lab animal exposure study is interesting because it involves a higher exposure level in some cases
Allergens: WebMD discusses allergy tests for humans at webmd.com/allergies/allergy-tests
Animal Allergens: Dog, Cat, and Other Animal Dander - Cleanup & Prevention Information for Asthmatics and regarding Indoor Air Quality.
Recognizing Allergens: What various indoor allergens look like - identification photos to help identify pollen, dust mites, animal dander, toxic or allergenic mold - Common Mold and other Allergens, Irritants, Remedies & Advice
Rodent control issues, including dander, fecal, and urine contamination of Buildings and Building insulation are discussed at our
Associations: Sick House, Sick Building, SBS - Air Quality, Government, Private Associations and Information Resources
Atlas of Clinical Fungi, 2nd Ed., GS deHoog, J Guarro, J Gene, & MJ Figueras, Centraalbureau voor Schimmelcultures, Universitat Rovira I Virgili, 2000, ISBN 90-70351-43-9 (you can buy this book at Amazon) - The Atlas of Clinical Fungi is also available on CD ROM
"A Brief Guide to Mold, Moisture, and Your Home", U.S. Environmental Protection Agency US EPA - includes basic advice for building owners, occupants, and mold cleanup operations. See http://www.epa.gov/mold/moldguide.htm
"Disease Prevention in Home Vegetable Gardens,"
Department of Plant Microbiology and Pathology,
Department of Horticulture, University of Missouri Extension - extension.missouri.edu/publications/DisplayPub.aspx?P=G6202
Fifth Kingdom, Bryce Kendrick, ISBN13: 9781585100224, is available from the InspectAPedia online bookstore - we recommend the CD-ROM version of this book. This 3rd/edition is a compact but comprehensive encyclopedia of all things mycological. Every aspect of the fungi, from aflatoxin to zppspores, with an accessible blend of verve and wit. The 24 chapters are filled with up-to-date information of classification, yeast, lichens, spore dispersal, allergies, ecology, genetics, plant pathology, predatory fungi, biological control, mutualistic symbioses with animals and plants, fungi as food, food spoilage and mycotoxins.
Ozone Warnings - Use of Ozone as a "mold" remedy is ineffective and may be dangerous.
Rot concerns in buildings-some building mold such as Meruliporia incrassata "Poria" risks serious rot and hidden structural damage
US EPA: Una Breva Guia a Moho - Hongo [Copy on file as /sickhouse/EPA_Moho_Guia_sp.pdf - en Espanol
OTHER IAQ ISSUES: How To Find and Address Other Indoor Air or Indoor Environment Contaminants Besides Mold
Mold or allergens may not be the only or even the main indoor environmental contaminant. Don't let media attention to mold
cause so much enviro-scare fear that other, possibly more urgent hazards go un-addressed.
Rodents, Mice, Squirrel Control - I find high levels of mouse and rodent dander, fecal dust, and urine-contaminated dust in some buildings,
and high levels of these materials in building insulation in those locations. If you have a mouse problem, particularly if mice and their waste (fecals or urine) are contaminating
the building HVAC or building insulation, may need both steps to clean up or remove infected materials and steps to stop an ongoing
rodent problem. If squirrels are a problem, the cleanup needs to include closing off entry openings into the building. Get some
help from a licensed pest control expert.