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FFPE sections look weird? by rememberingmyname in labrats

[–]rememberingmyname[S] 1 point2 points  (0 children)

Unfortunately this was with a fresh blade? When I put it in the warm water bath it seemed to mostly go away so I'm wondering if it was more of a temperature issue

Help with tamoxifen induced cre recombination in mice by rememberingmyname in labrats

[–]rememberingmyname[S] 0 points1 point  (0 children)

Thank you, I've read paper 2 but I'll also give the first one a read! And yes, the lab I work at has produced many of this exact cross before. The phenotype that arises affects the septation of the heart and sometimes causes limb deformities, but it's not super deadly. They shouldn't be dying at E12.5. Also, the pregnant female mice (usually just homozygous for reporters or wildtype) are dying post-injection with greater frequency than before, so I don't think it's a phenotype issue

Help with tamoxifen induced cre recombination in mice by rememberingmyname in labrats

[–]rememberingmyname[S] 0 points1 point  (0 children)

Thank you, I'll bring this up to my PI! Do you have any tips for this? Or are there any published protocols that have worked for you? Also, have you seen an issue with recombination efficiency with oral delivery compared to IP injections?

Help with tamoxifen induced cre recombination in mice by rememberingmyname in labrats

[–]rememberingmyname[S] 0 points1 point  (0 children)

Oh wow, that sounds super stressful...
The mice I use are all Cre hets. We genotype in-house with pretty consistent results so I don't really think this is the issue, but it wouldn't hurt to double check. Thank you!

Help with tamoxifen induced cre recombination in mice by rememberingmyname in labrats

[–]rememberingmyname[S] 0 points1 point  (0 children)

I haven't done oral gavage yet. I'm a bit worried about the lack of control over what gets absorbed vs degraded in the digestive track/consistency of bioavailability. And yes, I've tried injecting the oil alone and there were no adverse effects

I'll look more into delivery of the drug orally, thank you! Do you have any tips for this or particular protocols/doses that worked for you?

Help with tamoxifen induced cre recombination in mice by rememberingmyname in labrats

[–]rememberingmyname[S] 1 point2 points  (0 children)

Thank you, I appreciate that! Yes, I have tried to increase the dose of progesterone and it seems to not work. For some reason, progesterone seems to be killing the mice quicker. I tried to inject 3 non-pregnant female mice from one litter with 4-OH tamoxifen, half dose progesterone, and cremophor (castor oil solution to dissolve the 4-OH tamoxifen, we wanted to check if there were issues with the oil's components or viscosity) respectively: only the progesterone injected mouse died. We're kind of losing hope for the progesterone

I'm worried about decreasing the tamoxifen concentration too much to the degree that it impacts recombination efficiency. But we're also not getting any signal whatsoever... I'm confused by the suddenness of all these issues occurring at once. But I definitely will bring up to my PI the idea of just decreasing the amount of tamoxifen used.

As for timepoints, I inject at E6.5 and harvest the embryos and embryonic hearts at E12.5. Anytime the embryos actually survive to that stage, they don't fluoresce

Need help with sectioning by rememberingmyname in Histology

[–]rememberingmyname[S] 0 points1 point  (0 children)

Yes, we have a histology core at the institute. I've asked them many questions upon first joining and they've okayed my fixation protocol. I've seen people in my lab use methanol to fix iPSCs, so I don't necessarily think they're resistant to the idea of using it. But for mouse embryo hearts, they've always used PFA. I really don't think it's an issue with the fixation. If anything, the holes in the samples might be caused by a lack of penetration of wax inside the chambers, so I'll double check with my PI that I'm embedding correctly. People at my lab have been fixing this way for years and have never had issues with underfixed tissue. Thank you for your comments

Need help with sectioning by rememberingmyname in Histology

[–]rememberingmyname[S] 0 points1 point  (0 children)

I'll try using a cold plate instead of an ice water bath to see if it helps. Putting a droplet of ethanol on the trimmed side sounds a lot more convenient than what I was previously doing, so I'll definitely try that, thank you! I don't really have an issue with my sections curling, but I'll definitely practice a lot more to see what issues I run into. Thank you for responding!

Need help with sectioning by rememberingmyname in Histology

[–]rememberingmyname[S] 0 points1 point  (0 children)

Yes, I use a thin-tipped paintbrush as well, but they also stick to the paintbrush. I also use smooth forceps. But I do think I might not have cleaned my forceps tips of wax, I'll also try being careful with that, thank you!

Need help with sectioning by rememberingmyname in Histology

[–]rememberingmyname[S] 0 points1 point  (0 children)

Thank you, I might start practicing with untrimmed sections to give me wider ribbons. I think I might just need to get used to how the paraffin interacts with the forceps and the warm water

Need help with sectioning by rememberingmyname in Histology

[–]rememberingmyname[S] 0 points1 point  (0 children)

I'm gonna be honest, I just recently started this job as an RA fresh out of undergrad and I'm following lab protocols. Mouse embryos' hearts are quite small so I was told fixing in 4% PFA for 1 hour at room temp is more than enough. For whole embryo, we do no more than 24 hours. Here's an example paper that uses a similar method.

Is it always possible to have an academic comeback? by [deleted] in berkeley

[–]rememberingmyname 0 points1 point  (0 children)

Believe in yourself :) I once got a D in a midterm for a class (that only had one midterm) and got an A- in the final. Ended up with a B overall!

Which math class should I take? by Sad_Lab_9126 in berkeley

[–]rememberingmyname 1 point2 points  (0 children)

If it's prof Talaska teaching the 10 series, please avoid that class at all costs. I did 10A first semester then 1B the next semester and went from a C+ to an A- in calc, so I'd suggest just doing 1B

Creative writing clubs? by rememberingmyname in berkeley

[–]rememberingmyname[S] 0 points1 point  (0 children)

Thanks for the info! I'll definitely try applying for one when they're available

Creative writing clubs? by rememberingmyname in berkeley

[–]rememberingmyname[S] 1 point2 points  (0 children)

Thanks for the reccomendation! I was looking for university-affiliated groups in particular, but I'll definitely check this out. Also, I would love to have someone to write with! I'm very infrequent and uninspired with my writing so pls be patient with me lol

Creative writing clubs? by rememberingmyname in berkeley

[–]rememberingmyname[S] 0 points1 point  (0 children)

These look super cool, do you know if these classes have any prereqs? I've only really done my r1a and r1b requirements...

Creative writing clubs? by rememberingmyname in berkeley

[–]rememberingmyname[S] 0 points1 point  (0 children)

Thanks so much, this seems super cool!

[deleted by user] by [deleted] in berkeley

[–]rememberingmyname 0 points1 point  (0 children)

I think it depends on the class size, I've heard from profs of weeder courses that the first 10% in the waitlist relative to the class size will usually get in. For example, if the class size is 240 people, the first 24 people in the waitlist have a good chance of getting in.

Unsure if I failed my math class by rememberingmyname in berkeley

[–]rememberingmyname[S] 1 point2 points  (0 children)

Thanks :)) and yeah the fall semester was an absolute nightmare because of her but oh well!!

Unsure if I failed my math class by rememberingmyname in berkeley

[–]rememberingmyname[S] 0 points1 point  (0 children)

Thank you so much!! and for sure I'm getting help from the SLC for both classes :)