[OpenSPIM] Follow-up: [Open-SPIM] Is anyone using digital scanning to reduce banding?

Tue Jul 5 08:29:47 CDT 2016

Hi Tim,

I would be very interested in an example "before and after" image once you  
have them!

Regards,
Monika

W dniu .07.2016 o 15:18 Feinstein, Timothy N <tnf8 at pitt.edu> pisze:

> Hi folks,
> Many thanks for your help.  After much testing I have concluded that  
> anti-striping along the lines of Huisken & Stainier (2007) is extremely  
> >important for imaging large structures like a zebrafish embryo.  This  
> adds significant complexity and cost (~$2,000) to the basic openSPIM  
> >design but in my opinion it makes the instrument dramatically more  
> useful for segmentation and quantiative imaging.  Importantly the fact  
> >that I could make it suggests that it is within the engineering  
> abilities of most people who can build an openSPIM.  We have not started  
> using it >yet as our lasers are also out for an upgrade; I will update  
> the list when I have had a chance to test it.  
> I had to fill in some details from the 2007 paper on my own, so it is  
> likely that more elegant/less kludgy designs exist.  Still, this guide I  
> wrote >for our group may be useful for those thinking of adding  
> anti-striping in the future.  
> https://www.dropbox.com/s/6r1z13pggat7v76/Anti-striping%20mirror.docx?dl=0
>
> All the best,
>
> Tim
>
> Timothy Feinstein, Ph.D.Research Scientist
> University of Pittsburgh Department of Developmental Biology
>
>
>> From: <openspim-bounces at openspim.org> on behalf of Timothy Feinstein  
>> <tnf8 at pitt.edu>
> Date: Tuesday, May 3, 2016 at 9:27 AM
> To: Jan Huisken <huisken at mpi-cbg.de>
> Cc: "openspim at openspim.org" <openspim at openspim.org>
> Subject: Re: [OpenSPIM] Is anyone using digital scanning to reduce  
> banding?
>
>
> Correction, I meant 0.05% tricaine.  
>
> Best,
>
>
>> T
>
>
> From: Jan Huisken [mailto:huisken at mpi-cbg.de]Sent: Friday, April 29,  
> 2016 3:01 AM
> To: Feinstein, Timothy N
> Cc: openspim at openspim.org
> Subject: Re: [OpenSPIM] Is anyone using digital scanning to reduce  
> banding?
>
>
> Dear Tim,
>
>
> I think you are mixing up two issue.
>
>
> 1. Getting rid of stripes.
>
>
> Yes, you can do a little bit in post-processing but the stripes can be  
> quite complex and are not necessarily straight. Oftentimes you >see a  
> lot of lensing and I do not think you can easily remove it with image  
> processing.
>
>
> a) Double-sided illumination helps. Depending on the sample you may want  
> to do simultaneous or sequential double-sided >illumination. There are  
> various ways of merging the two images.
>
> b) A resonant mSPIM mirror is cheap and easy to integrate. You will get  
> rid of most stripes, but again this depends on the sample. >The details  
> for a) and b) are in our Opt. Lett. paper (Huisken & Stainier, 2007).  
> Basically, you are pivoting the light sheet around the >center of your  
> field of view, e.g. with 1kHz. The stripes are “washed out” during the  
> exposure time of your camera. We use this on >most of our systems and it  
> helps a lot to reduce stripes.
>
> c) Multi-view fusion can also help to some extent.
>
>
> 2. DSLM vs. SPIM
>
>
> Yes, DSLM should also help reducing stripes but I have no experience  
> with that. Basically, your light sheet is not coherent anymore >and the  
> sheet does not “interfere with itself”. However, the setup is more  
> expensive and requires you to power and control another >element: the  
> galvo scanner to sweep the beam up and down. You need a proper scan  
> mirror and not just the cheap mSPIM mirror. >The additional benefit is  
> that the resulting light sheet is more uniform than the cropped Gaussian  
> light sheet. However, you need to >illuminate each line with higher  
> intensity which may result in saturation and worse dynamic range.  
> Obviously you need to wait for the >scan and need to synchronize your  
> readout. Depending whether you have a global shutter or rolling shutter  
> camera you need to take >some precaution.
>
>
> Best
>
> Jan
>
>
>
>
> —
>
> Jan Huisken
>
> Head of Max Planck Research Group
>
>
> Max Planck Institute of Molecular Cell Biology and Genetics (MPI-CBG)
>
>
>>
>> On Apr 27, 2016, at 4:13 PM, Feinstein, Timothy N <tnf8 at PITT.EDU> wrote:
>>
>>
>> Hello all,
>> I found the Selzer group's solution (Keller et al. 2013) quite  
>> interesting
>> and would love to know if anyone had a positive experience doing that.
>> The banding from my basic openSPIM is definitely a problem for
>> segmentation and analysis.
>>
>> As far as I can tell fixing the banding will mostly involve buying a  
>> scan
>> mirror.  I don't have a quote yet for the Cambridge VM500 scanning  
>> mirror
>> that the Selzer group used but its ThorLabs equivalent (#GVS101?) costs
>> about $1k.  That seems reasonable and not that hard for regular folks to
>> implement.  
>> In related news I just noticed that the TeraStitcher plugin for Vaa3D  
>> now
>> has an option to fight banding in light sheet data.  I have not tested  
>> it
>> yet but potentially that feature could be useful.  I will just post my
>> usual warning for folks trying TeraStitcher for the first time: it
>> stitches really well but it could be a hair more user-friendly.
>>
>> Best,
>>
>> Tim
>>
>> Timothy Feinstein, Ph.D.
>> Research Scientist
>> University of Pittsburgh Department of Developmental Biology
>>
>>
>> _______________________________________________
>> OpenSPIM mailing list
>> OpenSPIM at openspim.org
>> http://openspim.org/mailman/listinfo/openspim
>
>

-- 
Dr Monika Pawłowska
Nencki Institute
02-093 Warsaw
Pasteura 3
Poland
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