Scanning Electron Microscopy Project Portfolio

Michael LeClair12/11/2013

Submitted for MCR 785 Scanning Electron Microscopy

Fall 2013N.C. Brown Center for Ultrastructure Studies

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Part 1

A portfolio of micrographs demonstrating the following techniques:

• Critical Point Drying or TMS drying• Depth of Field• Backscatter• Low voltage image of uncoated sample• High Magnification (>50,000 )• Stereo Pair• Cryofracture• Aesthetic

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TMS drying dryingFig. 1: SE micrograph of hyacinth anther. Fixed with 2.5% glutaraldehyde in PBS and post fixed with osmium tetroxide. Dehydrated with ethanol series and TMS. Affixed to aluminum stub with carbon tape and sputter coated with platinum.

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Depth of FieldFig. 2. SE micrograph of carpenter ant. Fixed with 2.5% glutaraldehyde in PBS, dehydrated in ethanol and dried with TMS before affixing to an aluminum stub with carbon tape and sputter coating with platinum.

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BackscatterFig. 3: SE micrograph of cross section of a U.S. penny. Spot size 12 chevrons, BS (left) accompanied by SEI of the same location (right). Penny was cut using a Leatherman™ multitool, and cleaned with ethanol and allowed to air dry before affixing to an aluminum stub with carbon paint, uncoated.

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Low voltage of uncoated sampleFig. 4. SE micrograph of fiber in home siding tile. Tile was broken using pliers to give a close to flat surface, and affixed to an aluminum stub with carbon paint.

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High MagnificationFig. 5.: SE micrograph of moth wing scale. Moth was fixed whole with 2.5% glutaraldehyde in PBS and dehydrated with ethanol and TMS before removing wings. Wing was affixed to aluminum stub with carbon paint and sputter coated with platinum.

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Stereo PairStereo image of Opuntia cactus. Cactus samples isolated from tissue culture, and fixed with 2.5% glutaraldehyde in PBS before coating with carbon in a high vacuum evaporator. Images taken at 15kV, working distance of 46mm, at 75x in SEI with a tilt of 8˚ difference between the images. Images combined using Photoshop™ software.

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CryofractureSE micrograph of Christmas cactus ovary. Frozen with liquid nitrogen and fractured using a razor blade before fixing with 2.5% glutaraldehyde and dehydrating with ethanol and TMS. Fixed to an aluminum stub with carbon paint and sputter coated with platinum.

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AestheticSE micrograph of carpenter ant. Fixed with 2.5% glutaraldehyde in PBS and dehydrated with ethanol and TMS before affixing to an aluminum stub with carbon paint. Sputter coated with platinum.

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Part 2Biological Samples

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Moth wingMoth was fixed whole in 2.5% glutaraldehyde in PBS before dehydrating in ethanol and TMS. Wings were removed and affixed to an aluminum stub with carbon paint before sputter coating with platinum.

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White Pine pollenPollen was placed directly on a thin layer of carbon paint on an aluminum stub before sputter coating with platinum. No fixation was used.

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Porcupine quillQuill was affixed to an aluminum stub with carbon paint and sputter coated with platinum, with no fixation prior to mounting.

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Mum stem epidermisChrysanthemum sample was cut into small sections and fixed in 2.5% glutaraldehyde in PBS before dehydrating with ethanol and TMS. Affixed to aluminum stub with carbon tape and sputter coated with platinum.

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Mum disk flowerChrysanthemum flower was selected for its small size and ray flowers were removed before fixing in 2.5% glutaraldehyde and dehydrating with ethanol and TMS. Flower was then affixed to an aluminum stub with carbon paint and sputter coated with platinum

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Wasp ocelliYellowjacket wasp was found dead and was already dried. No fixation was used, and sample was cut into separate segments and affixed to an aluminum stub with carbon paint before being sputter coated with platinum.

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PollenPollen grain from a sample containing anthers from many species of plants. Pollen was fixed in 2.5% glutaraldehyde in PBS and post fixed with osmium tetroxide. Particulate matter on surface of pollen grain is most likely precipitated osmium from leaving in fixative too long. Fixed pollen was dehydrated with ethanol and TMS to dry from fixation, sprinkled on carbon paint coating an aluminum stub, and sputter coated with platinum.

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Wasp paperPaper from a small paper wasp nest. Cut with scissors and affixed with carbon paint to an aluminum stub and sputter coated with platinum.

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Part 2Non-Biological Samples

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Electronics chipChip was removed from a outdated cellphone and was affixed, uncoated, to an aluminum stub using carbon paint.

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GlassA small glass pipette was carefully crushed and the fragments were sprinkled onto carbon paint on an aluminum stub. Due to the non-conductive nature of glass, the specimen was then sputter coated with platinum before imaging.

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Hypodermic needleThe tip of hypodermic needle was removed using a multitool. It was then affixed, uncoated, to an aluminum stub with carbon tape.

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Wire meshWire mesh of unknown composition was cut with scissors and affixed to an aluminum stub with carbon tape, uncoated.

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SnowflakePolymerized with superglue, affixed to aluminum stub with carbon tape. Uncoated. Images at 10.0kV.

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Sea saltSea salt was sprinkled on carbon tape on an aluminum stub. Grains were then sputter coated with platinum to increase conductivity.

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Fuse filamentFilament was removed from a standard bus fuse using a pair of pliers and affixed to an aluminum stub using carbon tape. Uncoated.

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Razor bladeLeading edge of a (new) single bladed razor blade. Blade was broken with pliers to provide a flat edge and stood on edge on an aluminum stub by sticking to carbon tape. Carbon paint was then placed on each side to keep it standing on edge. Uncoated.