reSolution_LNT_No1_en - Leica Microsystems

BIOLOGY
10 reSOLUTION
Conclusion
OTE and Pt Blue were assessed as substitutes for UA in
section post-staining for electron microscopy with regard
to contrasting effi ciency, toxicity, handling, and price. Manual
as well as automatic staining procedures with the Leica
EM AC20 were tested. No en bloc staining was performed.
Furthermore, resins other than Agar 100 or different section
thicknesses may lead to different results.
Both the rather weak contrast obtained with OTE as well
as the contamination observed on the specimen were not
convincing with manual staining and the Leica EM AC20. Pt
Blue delivered clearly better results with both approaches.
However, slight differences in contrasting properties as
compared to UA have to be taken into account for interpretation
of micrographs (Yamaguchi et al., 2010). The quality
of results that can be obtained from automatic staining is
comparable with the manual procedure for both OTE and
Platinum Blue.
Regarding toxicity, both reagents tested as substitutes of
UA have the advantage of not being radioactive. Furthermore,
working with health-damaging inhalable powder can
be avoided. Being a food product it can be presumed that
OTE is non-hazardous. In contrast, Pt Blue is toxic but as it
is delivered as a stock solution further risky handling can
be reduced to a minimum. The utilization of an automatic
staining device such as the Leica EM AC20 can help to further
minimize contact with toxic reagents.
Staining with UA and Pt Blue is comparable in time
and labor, whereas contrasting with OTE is more time
consuming without delivering as satisfactory results.
As with the Leica EM AC20, it is possible to stain
Instruments related to this sample preparation:
Leica EM AC20
Automatic Constrasting Instrument for Ultrathin Sections
Leica EM TRIM2
Specimen Trimming Device for TEM, SEM, LM
up to 20 grids at once and all steps are carried out
automatically, this becomes less of a disadvantage of OTE.
From the economic point of view, OTE was by far the cheapest
product. The price per grid at the used concentration of
0.2% was signifi cantly lower than for Pt Blue (at a dilution
of 1:100) and the 2.0% UA solution.
Summing up, electron microscopists in need of a replacement
for UA for post-staining of sections have the choice
between one reagent at a very cheap price and minimal
risk with moderate results – Oolong tea extract – and
another one, that delivers very convincing results, but at
higher cost and not without safety risks – Platinum Blue.
The experiments here have shown that this is applicable
for both manual as well as automated staining with the
Leica EM AC20.
Acknowledgements
The authors would like to thank Yanli Tong (Leica Microsystems,
Shanghai) for assistance with acquiring reagents and
Jean Trichereau (IMBA, Vienna) for providing samples. The
work of N.F., M.B. and G.P.R. was supported by the City of
Vienna/Zentrum fuer Innovation und Technologie through
the Spot of Excellence grant “Center of Molecular and Cellular
Nanostructure.”
Contact
Dr. Günter Resch
Head of Electron Microscopy, Campus Science Support
Facilities GmbH, Vienna, Austria
guenter.resch@csf.ac.at
Leica EM UC7
Ultramicrotome for Perfect Sectioning at Room Temperature and Cryo