[3dem] Vitrobot Mark 4 issues

Bob Grassucci rg2502 at columbia.edu
Thu Feb 19 14:15:33 PST 2009


Hi All,
We just got our FEI Vitrobot Mark IV and I have been trying to get the 
parameters worked out for reproduceably making good frozen hydrated 
grids.  We had a previous version Mark 2 before we moved tho Columbia 
and were able to optimize it but the Mark IV has been modified so that 
these parameters are no longer valid.  We are using Quantifoil grids 
with thin carbon spread over them and the ice is either too thick or the 
carbon is damaged by plunging.  The parameters  offset have been changed 
to blot force which varies the distance the pads are from each other 
when blotting.  I have tried blotting times from 1 to 8 seconds and blot 
forces from -25 to +20.  FEI suggests that I use their blot papers only 
but when I tried it there was no improvement. We still are able to make 
grids the old fashioned way with our home made device but would like to 
take advantage of this expensive apparatus.  If anyone else has a 
similar set up i.e. Mark IV and using thin carbon, I would appreciate 
hearing your experience with it and if you got it to work what 
parameters you used.  Thanks for your help
Regards,
Bob

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Howard Hughes Medical Institute
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