10:30 – 1:30A
- checking in situ stains: bio and WGA stains don’t seem to have worked? (ubx-bio, Antp-bio, and en-bio slides all no stain)
- should have put shp in 555 as well as 750 to confirm stain. 🙁
- hb finally has signal at 1:50 conc in 15 min PFA GA fixed embryos. Bad background in hb-channel around embryo periphery: try traditional FA fix for 10 min.
- No WGA. Maybe Xylenes treatment removes membrane? Dostatni’s method according to the citations seems to not use Xylenes. Pare uses a 10 min acetone permialization. “clearing yolk” shouldn’t be necessary for my studies anyway.
- Got unlabeled WGA for Colenso. Diluted to 1mg/mL in PBS.
- should we use PBS azide for antibodies? — currently just used for preservation of labeled cell samples.
- testing DNA dyes: now in 30% resin at 3P. 50:50, 4P.
- Finished Resin embedding, samples at 70C. 1:00A.
- Homeotic mutants: BSC 2235: Antp[Ns] — hets show antenna to legs
- hb stain works in 15 min fix embryos (yay!) substantial background on basement membranes. Maybe due to fix procedure? or goat secondary? basement membrane stain is much stronger than nuclear stain. some background still in non-hb nuclei.
- Try pre-abosrbing hb. Try 10 min 8% FA fix.
- H3K4 stain is definetely pan-nuclear — apparent circular/nuclear membrane feature in last stain probably an artifact. H3K4 detectable in both very late and cc14 embryos — try cutting new embryo.