Re: Imaging Sourdough batter **vendor reply**

[Kilgore, Jason A.]

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An tasty question!

While I have never worked with that sort of sample, I can at least suggest that, instead of DAPI, a more permeant nucleic acid stain such as Hoechst 33342 or SYTO 9 may be better options.  But that will label all DNA (such as from the wheat), not just the yeast.

Other than sections and optical counting, they could also perform a differential gradient centrifugation to separate the yeast from the other batter components. Then maybe perform an O.D. based quantitation.

Jason


Jason A. Kilgore
Technical Application Scientist
Molecular Probes / EVOS Tech Support
Thermo Fisher Scientific
 
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-----Original Message-----
From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Joel Sheffield
Sent: Tuesday, March 20, 2018 12:48 PM
To: [hidden email]
Subject: Imaging Sourdough batter

CAUTION: This email originated from outside of the organization. Do not click links or open attachments unless you recognize the sender and know the content is safe.


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Colleagues,

Here's something to chew on.

This is not strictly a confocal question, but applies to widefield as well.  Some folks here are interested in analysis of the time course of the maturation of  sourdough, and have asked us to help them prepare samples of the batter for analysis of, say, yeast content.  Have any of you had any experience with this sort of material?  We have thought about making frozen sections of the batter at different times after initiation, and staining with DAPI. I'm curious about fixation (note that there are some interesting pH issues here), and quantitation methods.  Any suggestions will be appreciated.

Joel


Joel B. Sheffield, Ph.D
Department of Biology
Temple University
Philadelphia, PA 19122
Voice: 215 204 8839
e-mail: [hidden email]
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[Benedetto, Alexandre]

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How about TEM after high-pressure freezing? Sticky dough is perfect for this, it should be free from fixation artefacts, you would definitely get the temporal and spatial resolution to look at fine processes.
Best wishes,

Alex

> On 20 Mar 2018, at 20:09, Kilgore, Jason A. <[hidden email]> wrote:
>
> *****
> To join, leave or search the confocal microscopy listserv, go to:
> http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> Post images on http://www.imgur.com and include the link in your posting.
> *****
>
>
> An tasty question!
>
> While I have never worked with that sort of sample, I can at least suggest that, instead of DAPI, a more permeant nucleic acid stain such as Hoechst 33342 or SYTO 9 may be better options.  But that will label all DNA (such as from the wheat), not just the yeast.
>
> Other than sections and optical counting, they could also perform a differential gradient centrifugation to separate the yeast from the other batter components. Then maybe perform an O.D. based quantitation.
>
> Jason
>
>
> Jason A. Kilgore
> Technical Application Scientist
> Molecular Probes / EVOS Tech Support
> Thermo Fisher Scientific
>  
> 1-800-955-6288 then option 4, then option 3, then option 2.
> Or dial direct at [phone]
> [hidden email]
>  
> This communication is intended solely for the individual/entity to whom it is addressed. It may contain confidential or legally privileged information.  Any unauthorized disclosure or copying is prohibited and may be unlawful. If you have received this communication in error, please notify the sender immediately and delete it from your system.
>  
>  
>
>
> -----Original Message-----
> From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Joel Sheffield
> Sent: Tuesday, March 20, 2018 12:48 PM
> To: [hidden email]
> Subject: Imaging Sourdough batter
>
> CAUTION: This email originated from outside of the organization. Do not click links or open attachments unless you recognize the sender and know the content is safe.
>
>
> *****
> To join, leave or search the confocal microscopy listserv, go to:
> https://urldefense.proofpoint.com/v2/url?u=http-3A__lists.umn.edu_cgi-2Dbin_wa-3FA0-3Dconfocalmicroscopy&d=DwIBaQ&c=q6k2DsTcEGCcCb_WtVSz6hhIl8hvYssy7sH8ZwfbbKU&r=MVp-2yJ1A-yQtCbZ-DE9xd0W6E7srQQpV-yioYjTLyQ&m=k-Zx02tX7trsPJ1Penysg5iJChR7COlkdSxjClwfNHE&s=BEO5EsTWp_Ny4EvDpjIWQX-O997jFrrPdAdGzNiUfZA&e=
> Post images on https://urldefense.proofpoint.com/v2/url?u=http-3A__www.imgur.com&d=DwIBaQ&c=q6k2DsTcEGCcCb_WtVSz6hhIl8hvYssy7sH8ZwfbbKU&r=MVp-2yJ1A-yQtCbZ-DE9xd0W6E7srQQpV-yioYjTLyQ&m=k-Zx02tX7trsPJ1Penysg5iJChR7COlkdSxjClwfNHE&s=8pixJss-tzRi0l3BIi7ZUjhHQewWCzzPon3qjag3sO0&e= and include the link in your posting.
> *****
>
> Colleagues,
>
> Here's something to chew on.
>
> This is not strictly a confocal question, but applies to widefield as well.  Some folks here are interested in analysis of the time course of the maturation of  sourdough, and have asked us to help them prepare samples of the batter for analysis of, say, yeast content.  Have any of you had any experience with this sort of material?  We have thought about making frozen sections of the batter at different times after initiation, and staining with DAPI. I'm curious about fixation (note that there are some interesting pH issues here), and quantitation methods.  Any suggestions will be appreciated.
>
> Joel
>
>
> Joel B. Sheffield, Ph.D
> Department of Biology
> Temple University
> Philadelphia, PA 19122
> Voice: 215 204 8839
> e-mail: [hidden email]
> URL:  *https://urldefense.proofpoint.com/v2/url?u=https-3A__bio.cst.temple.edu_-7Ejbs_&d=DwIBaQ&c=q6k2DsTcEGCcCb_WtVSz6hhIl8hvYssy7sH8ZwfbbKU&r=MVp-2yJ1A-yQtCbZ-DE9xd0W6E7srQQpV-yioYjTLyQ&m=k-Zx02tX7trsPJ1Penysg5iJChR7COlkdSxjClwfNHE&s=v2s5ujeTsuV_7HlEC1O2OUfAaPtyZ7wzYAvfIhldH5U&e=
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