Sample guide XFM
- Microscopy Cluster (Unlicensed)
In general, place samples closely together on the sample mounts. We typically scan a whole sample mount quickly to get an overview, so having samples closely packed will save significant scan time. This is particularly relevant to XANES standards.
Please see below info for preparation of various sample types.
Most approaches to specimen preparation for metals mapping in animal tissues rely on metals immobilisation. This can occur through chemical fixation or cryo-fixation followed by dehydration or freeze substitution.
A recent investigation [1] has indicated that formaldehyde fixing can result in metals redistribution in certain types of tissue sections, and that a short rinse in PBS can also disturb metals distribution. In that article, cryofixation and drying is used as the reference standard for preserving metals distributions.
Unfortunately it is not always possible to plunge freeze a specimen, especially when the specimen is so large that cooling rates are not sufficient to prevent ice-crystal formation [2]. However, where plunge freezing and freeze-drying (or freeze substitution) is an option, we at least recommend that a comparison of the two techniques be performed as a part of the beamtime request.
We highly recommend the use of silicon nitride (or silicon carbide) windows for mounting specimens. Adherent cells can be grown directly onto silicon nitride windows, and these are compatible with a variety of imaging and spectroscopic investigations [3]. We have a limited supply of these windows at the beamline; so please feel free to ask for some if you require less than 10 or so windows for your investigation. However, if you need more than this or plan repeated trips you should consider these to be a part of your cost for the beamline access. Such windows can be obtained from one of a number of companies including Silson. Please feel free to consult us prior to making your purchase.
Place specimens on the membrane-side aperture of a silicon nitride window, not in the 'well'. A cross-section view of a silicon nitride window is shown below to demonstrate.
We DO NOT recommend the use of plastic coverslips for mounting biological samples. Coverslips contain many metal impurities such as cobalt. Coverslips are also too thick and create excess scatter.
[1] Hackett et al., Analyst 136, 2941 (2011).
[2] Studer et al., Journal of Microscopy 203, 285 (2001).
[3] Carter et al., Molecular Biosystems 6, 1316 (2010).
At XFM we typically use Perspex sample holders to attach samples. We have various styles of sample mount. In the image below, silicon nitride windows are attached to the Perspex mount with double-sided adhesive tape. The aperture of each nitride window here is 4mm wide. The samples are facing us in the photo.
The Perspex mounts have slots or apertures to permit the transmitted beam to pass through.
We use a magnetic kinematic mount to attach the Perspex holder to our raster scanning stages (shown on the bottom left). This is quick and sample registration is usually kept to within 20 microns if the Perspex mount is removed then reattached.
We advise to leave a 5mm gap between samples and the far left and right ends of the mount to ensure we can reach the samples.
Shown below is a detail of leaves mounted within thin Ultralene film. The width of the rectangular aperture is approximately 41 mm. There are three apertures in total for this Perspex mount.
Shown below are many rice sections heroically mounted with Kapton tape! The three apertures are each 41 mm wide by 100 mm high. Tightly spaced samples save scan time.
Shown below is the wide variety of sample mounts we have on offer. Note the vertically aligned 150mm ruler in the middle row for a sense of scale. Four petrographic thin sections are shown mounted beside the ruler.