NetNotes
Edited by Thomas E. Phillips University of Missouri
phillipst@missouri.edu
Selected postings from the Microscopy Listserver from March 1, 2011 to May 1, 2011. Complete listings and subscription information can be obtained at
http://www.microscopy.com. Postings may have been edited to conserve space or for clarity.
Specimen Preparation: sputter coater target distance I just got a new sputter coater and need help determining the
sample to target distance. Instructions don’t have a recommendation, just a note that says the specimen height is adjustable. Any advice? Jon Krupp
jkrupp@deltacollege.edu Wed Mar 23 A generic guide to setting the specimen position in a sputter
coater is 5 cm target to specimen surface. T is means you are in the plasma but not too close to the target. Steve Chapman protrain@
emcourses.com Wed Mar 23 For your information we teach the following procedure for
setting up a sputter coater. Each test, other than the fi rst, uses a piece of normal copy paper cut down to about 2 in. × 3 in. to simulate the outgassing that would be typical of a specimen. (1) Using a piece of paper that covers the stage area, run the coater for several minutes at 20 mA. Hold the paper down with a stub placed at the centre. T e coat will demonstrate the coverage of the plasma and pick out areas that are well coated or not so well coated. T e coating should be pretty perfect with a new coater but patchy with a coater that has a contaminated target. If there are areas with less of a coat you need to map your stage for selecting the best position for superior coating in the future? (2) Place a test paper on the stage held down by a stub. T e next step depends upon the voltage of the coater (a) using a low voltage coater (400 V to 600 V) set the current at 10 mA and coat for 30 seconds. (b) with a high voltage coater (>800 V) set the current at 20mA and run for 30 seconds. Note the current and time on the back of the test paper. (3) Repeat the experiment in steps of 30 seconds until the paper starts to exhibit a very slight gold sheen. Note the information on the back of each picture. (4) Select the condition for higher magnifi cation work that shows the fi rst grey coloration. (5) Low magnifi cation operation will probably require the procedure set out in 3. Sputter coating is oſt en treated as a no brainer, but as those attending our course in Missouri later in the year will see, we dedicate an hour to correctly judge and set up a sputter coater for the fi rst time. It is that important if you wish to obtain information that is not infl uenced by the coating; just as is the selection of the correct coating material. For top class SEM results I prefer to use the full facility of the SEM, balancing specimen position, kV and probe current rather than coating. However sometimes you just have to fall back on that route, particularly for novice operators where it is so much easier to slap on a coat to keep life simple. Steve Chapman
protrain@emcourses.com T u Mar 24
Specimen Preparation: carbon rods in evaporators Our lab has a JEOL Vacuum Evaporator JEE-400 all a long
but it’s not frequently used. Recently i am trying to make use of this evaporator to establish a consistent carbon coating of certain thickness. Unfortunately there no one with expertise in our lab and I would to ask for some advice particularly on the sharpness of the carbon rod. Below
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is my question: How sharp must the carbon rod be for the contact? T e JEOL manual illustrate a needle sharp tip but the JEOL carbon sharpener seem only able to thin the rod to a diameter of about 1 mm. Would that actually means that aſt er using the JEOL carbon sharpener, I should fi nd other ways to create a sharp tip? I appreciate very much for your kind advice! Yee Yan Tay
one_twinklestar@yahoo.com.sg Mon Mar 28 My own experience with a Cressington evaporator is that too
fi ne a tip leads to unreliable evaporation—I spent a good deal of time getting the tip pencil sharp only to discover that it would spark up briefl y then break before suffi cient carbon had evaporated. A 1mm tip works well, I get control of the thickness by donning a pair of welding goggles and watching the arc light up, for general purpose grids I allow about a 2 second pulse from the point at which the arc reaches maximum brightness. Ian Portman
i.j.portman@warwick.ac.uk Tue Mar 29 We used to form two wedges by moving the rod across a grinding
paper. T e wedge angle was approximately 30–45 degrees. We also had a rotation vacuum transfer rod, which allowed us to press one wedge against the other stationary one. Josef Zweck
josef.zweck@
physik.uni-regensburg.de Tue Mar 29 T e ideal carbon rod combination is a spigot and a fl at which is
exactly what your JEOL sharpener is providing for you. T e spigot system is better than a point system as it provides a possibility of prolonged coating due to the constant surface area of the contact point. A point system very soon erodes to a surface area that is too large for the evaporation to continue. Set the spigot to strike a fl at surface with the sprung pressure being applied to the spigot rod. Remember the quality of a carbon coat is directly related to the vacuum level when setting up the system; give it time! Steve Chapman
protrain@emcourses.com Tue Mar 29
Specimen Preparation: membrane blebbing I remember seeing some discussion (many years ago?), on the
server about artifacts in microscopy of biological specimens in general. Specifi cally I think I read about membrane blebbing caused by aldehyde-based fi xing. Does anyone have any actual references about this? I have seen what might be this eff ect, but it’s hard to be sure. Peter Eaton
petereaton@hotmail.com Sat Apr 30 T e only paper I’m readily familiar with (I’m sure there are
others), is this one: G.J. Hyde, S. Lancelle, P.K. Hepler, A.R. Hardham (1991) Freeze substitution reveals a new model for sporangial cleavage in Phytophthora, a result with implications for cytokinesis in other eukaryotes. Journal of Cell Science 100, 735–746;
http://jcs.biologists. org/content/100/4/735.abstract. T ere are a number of citing articles on a similar theme. Rosemary White
rosemary.white@csiro.au Mon May 2
T is is a principle of science that any interaction with the specimen modifi es its nature, however we all strive to limit the
doi:10.1017/S1551929511000484
www.microscopy-today.com • 2011 July
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