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  • Bruno Vellutini 18:52 on 2014/06/14 Permalink
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    Live membrane staining in Membranipora 


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    I decided to try the live membrane staining FM® 4-64FX, fixable analog of FM® 4-64 membrane stain with Membranipora embryos on the confocal. I dissolved the dessecated FM into 100 µL of DMSO and then diluted 2.5 µL in 500 µL of FSW (final concentration of 5µg/mL). I then added embryos to a slide and completed the volume with the staining solution and sealed as for the 4D. Using the 561 laser I get a really nice signal and I did a few xyzt stacks to test. They look nice!

     
  • Bruno Vellutini 18:47 on 2014/06/14 Permalink
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    Membranipora U0126 experiment with different concentrations 


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    With the nice Saturday spawning I managed to setup a new U0126 inhibitor experiment. I made 2 replicates of the following setup:

    FSW DMSO 5 µM 7.5 µM
    1 µM 2.5 µM 10 µM 12.5 µM

    Embryos were activated at 14h and incubated at 17h in the facility incubator at 15 °C!

    I’ll fix the embryos on Monday after 48h.

    16/06/14

    Replicate one everything was dead except for FSW control, very weird. The second replicate seemed fine, although there should be a lot of noise. I’ll repeat  the same today and fix 24h after.

    I’ll fix with 48h.

     
  • Bruno Vellutini 18:42 on 2014/06/14 Permalink
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    Fresh new Membranipora 4D 


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    Started a Membranipora 4D from the Saturday spawning. First try was not well embedded, but the second is ok. Started around 16h. Eggs activated at 13h.

    Filename: BCV_Mm_2014_06_14 (old Mmem_b14_06_14)

    16/06/14

    It finished well, but the top is not visible in the middle of the recording and the embryo moves outside the picture borders in the end.

    It does gastrulate, but I lost the embryo when trying to get it out the slide…

    It is a lateral view, antero or postero lateral. Not sure yet.

     
  • Bruno Vellutini 18:40 on 2014/06/14 Permalink
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    After 2 days of no eggs I think I got a good colony. There is still a lot of poop, but the embryos look good.

     
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