http://confocal-microscopy-list.275.s1.nabble.com/organoid-imaging-tp7590835p7590856.html
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> Dear Mary Ellen,
>
> staining of organoids within matrigel is a challange and results in bad
> signal to noise ratios... but if your organoids are expressing GFP or
> even better long wavelength fluorophores, imaging can be performed
> within the matrigel matrix. A dilution of matrigel with medium works
> also well and dilution of up to 1:5 are applicable.
>
> To increase the amount of organoids close to the coverslip, you can
> place the dish on an ice pack while you seed the
> matrigel-medium-organoid suspensions and keep it on ice for 5 minutes.
> Within cold matrigel the organoids sink by gravity and you´ll have more
> organoids within the working distance of the lens... I recommend
> spinning disc microscopy, due to low photo-toxic stress. Matrigel and
> 2Photon might be challenging, due to the second harmonics of the
> collagen matrix...
>
> Best regards,
>
> Philipp
>
>
> On 28/04/2020 16:21, Mary Ellen Pease wrote:
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> > Hi all,
> >
> > I have a user who would like to image live organoids via
> confocal/multiphoton without jeopardizing the sample. Has anyone figured
> out a material in which the organoid could be placed that would eliminate
> movement/drift during imaging, without damaging it so the sample could go
> back into culture afterwards? So far, the papers I've looked at on this
> topic cryoembed them at a certain time point for immunohistochemistry. My
> user's lab has samples which are expressing GFP and RFP, and would like to
> look at them in vivo. Someone in our department suggested putting them in a
> mix of matrigel and culture media, which could slow them down temporarily,
> but in my experience, matrigel attenuates the signal. We have a multiphoton
> system on an inverted Zeiss platform (LSM 710). we can image using an
> objective inverter and dipping lenses, or from the bottom through a
> coverglass dish. Since we are working remotely at this point in time, I was
> hoping to get some direction from this group.
> >
> > Best,
> >
> > Mary Ellen Pease
> > Manager, Wilmer Microscopy and Imaging Core Facility (MICF)
> > Johns Hopkins SOM, Baltimore MD
>
> --
> Philipp Tripal
> Research Associate
> Optical Imaging Centre Erlangen
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