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  • Bruno Vellutini 16:37 on 2015/08/25 Permalink
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    Membranipora simple staining 

    Just a simple staining of mixed stages of Membranipora using DAPI and Phalloidin 647 for morphology.

    • 3x 30min 0.2% PTx washes.
    • 1x PTx overnight at 4°C.

    26/08/15

    • Staining with Phalloidin and DAPI for 2h.
    • Let embryos in PBS+DAPI 1:1000 at 4°C.

    It failed… weird.

     
  • Bruno Vellutini 12:09 on 2015/08/25 Permalink
    Tags: ,   

    Membranipora in situ + mapk, a re-try 

    Since I saved some de-glycerolled embryos I’ll repeat the immuno staining with a longer developing time.

    gata b nk2.1
    • Washed 1h in PBT (4x).
    • Washed 3h in 5% NGS (3x).
    • Added anti-mapk antibody 1:100.
    • Let overnight at 4°C.

    26/08/15

    • Washed primary antibody with 3x PBT 5min + 3x 30min.
    • 5% NGS 3x 30 min.
    • Added both anti-mouse-pod/hrp at 1:250 for 4h at room temp.
    • Washed 4x 5min with PBT and let overnight at 4°C.

    27/08/15

    • Wash 1x PBT.
    • Wash 3x TNT buffer.
    • Develop with cy3 for 5min.
    • Wash with PBS (no detergent solution).
    • Embed in TDE.
     
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