[3dem] Is there an asymmetric biological sample out there that reveals handedness at CET resolution?

Friedrich Foerster frido at salilab.org
Tue Aug 7 12:30:09 PDT 2012


hi christian,

the handedness problem arises because tilt series alignment cannot
distinguish a tilt axis of x and x+180 degrees. one option is indeed
to acquire tomograms of something asymmetric like ribosomes.
alternatively, you do the same thing people do in random conical
tilting to determine the tilt direction: take micrographs of carbon
and tilt your sample. the ctf rings right and left of your tilt axis
will tell you which side is top and which is bottom.

cheers

friedrich

On Tue, Aug 7, 2012 at 8:33 PM, Mike Strauss
<mikestrauss13 at crystal.harvard.edu> wrote:
> Hi Christian,
>
> you could use a helical particle and look at the rise and rotation of the subunits.  Perhaps microtubules, or a helical virus.
>
> If you want to see if a sample is loaded in the reverse direction, you could reconstruct a plastic section with fiducially on a known side (i.e. always on top), and then see where they end up in the program.
>
> good luck.
>
> mike
>
>
> On Aug 7, 2012, at 2:27 PM, Christian Geiss <geiss at biophysik.org> wrote:
>
>> Hello,
>>
>> I am a Phd student, do cryo electron tomography and have a problem determining the handedness of my sample, because viewers for tomograms like Amira, Matlab-based scripts can invert the handedness depending if you look at your tomogram from top to bottom or vice versa. Since those programs behave to some extent like black boxes, the easiest would be to have a nice asymmetric biological sample where you know the handedness and that the latter can be judged quite easily from a reconstruction, meaning without further processing like subtomogram averaging.
>>
>> So far, I can only think of bacterial ribosomes since they are asymmetric and quite big, but for cryoET the resolution would be not reasonable to resolve the handedness at all. Thus I could need in the ideal case e.g. something like a large complex that links each other to repetitive units with a known handedness.
>>
>> I know its a tough question, but maybe someone can give creative input here? I would highly appreciate it!
>>
>> Best regards,
>>
>> C. Geiss
>>
>>
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> Mike Strauss
> Harvard Medical School - Dept. BCMP
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>
>
>
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-- 
Dr. Friedrich Foerster
Max-Planck Institut fuer Biochemie
Am Klopferspitz 18
D-82152 Martinsried

Tel: +49 89 8578 2632
Fax: +49 89 8578 2641

www.biochem.mpg.de/foerster


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