Re: Tetraspeck beads for PSFs **vendor reply**

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>I think you may simply keep dry >beads on a coverslip

The bead may be 100 nm, but you want to measure the full PSF of about 4 micron, so proper mounting is important.

Andreas

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> On 4 Aug 2018, at 13:36, MODEL, MICHAEL <[hidden email]> wrote:
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> I think you may simply keep dry beads on a coverslip and there is no need in a mounting medium. No spherical aberration would accumulate over a 0.1 um depth
>
>
> Mike Model
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> From: Confocal Microscopy List <[hidden email]> on behalf of Andreas Bruckbauer <[hidden email]>
> Sent: Saturday, August 4, 2018 4:01 AM
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> Subject: Re: Tetraspeck beads for PSFs **vendor reply**
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> Thanks Jean for raising this issue, I have a similar problem, the dye diffuses out of the beads and leaves a large spot around them. It can happen very quickly when the slides get to warm and the current heatwave is not helping. Putting them in the fridge is also not an option because of drift once put back on the microscope.
> This is with mounting medium from Moleculr Probes for beads.
> No solution yet, I don’t think it used to be like this, have you changed the beads?
>
> Aren’t 100 nm beads too big for PSF measurements on the Elyra? 40 nm should be better.
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> Best wishes
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> Andreas
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>> On 3 Aug 2018, at 22:05, Kilgore, Jason A. <[hidden email]> wrote:
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>> ** Vendor reply **
>>
>> Polystyrene microspheres, such as the TetraSpecks, will swell in the presence of certain solvents.  So you'll want to use an aqueous-based mounting media, preferably one that cures (such as ProLong products or Fluoromount-G) instead of  using an organic solvent-based one, like Cytoseal-60.  When dye-labeled microspheres like these swell, the dye is free to diffuse out of the polystyrene matrix.
>>
>> Cheers,
>>
>> Jason
>>
>>
>> Jason A. Kilgore
>> Technical Application Scientist
>> Molecular Probes / EVOS Tech Support
>> Thermo Fisher Scientific
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>> 1-800-955-6288 then option 4, then option 3, then option 2.
>> Or dial direct at [phone]
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>> -----Original Message-----
>> From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Jean Ross
>> Sent: Friday, August 03, 2018 12:49 PM
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>> Subject: Tetraspeck beads for PSFs
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>> Hi Everyone,
>>
>> I have been preparing 0.1um Tetraspeck bead slides to use when measuring point spread functions on our Zeiss Elyra PS1 super resolution microscope
>> and Zeiss LSM880 confocal with Airyscan.   The problem is that over time
>> the beads appear to swell to about 3 times in size and lose much of their fluorescence.  I have tried many different mounting medias (hardening and
>> nonhardening) all with the same result.  Has anyone else experienced this problem?  Any suggestions to solve the problem would be appreciated.
>>
>> Thanks,
>> Jean
>>
>> --
>> Jean Ross
>> Delaware Biotechnology Institute, BioImaging Center University of Delaware
>> 15 Innovation Way
>> Suite 117
>> Newark, DE  19711
>> Phone:  [phone]
>> Fax:  [phone]