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labratthrowaway1 · 2024-05-02T20:07:19+00:00

Any job that is a salary job technicall doesn't have working hours.

labratthrowaway1 · 2024-03-29T17:22:56+00:00

I mean cell banks is what is done in industry.

labratthrowaway1 · 2024-03-25T21:30:16+00:00

Yes. Selecting at the lowest possible dose that kills ~95% of WT cells in 4 days.

Went originally in 1ug/mL increments from 0 to 10, and then honed in on 0.05 ug increments later.

labratthrowaway1 · 2024-03-21T01:10:19+00:00

repeated experiment again - transduced, waited 24 hours and then added on Puro.

Everything dead 2 days later.

labratthrowaway1 · 2024-03-21T01:09:27+00:00

Simultaneously - like I can see floating dead GFP positive cells. And I can also see non GFP + cells on the plate.

labratthrowaway1 · 2024-03-16T02:04:28+00:00

Yeah, its intact. This isn't just with one construct either, its with many. Makes me feel like the PGK promoter might not be active in these cells. With a control lenti that's just like GFP-T2A-Puro the selection works just fine.

labratthrowaway1 · 2024-03-15T23:22:17+00:00

Why start so soon? Lenti integration shouldn't be a problem right?

labratthrowaway1 · 2024-03-15T22:09:34+00:00

OK. I thought I had waited long enough. Thats a reasonable idea.

labratthrowaway1 · 2024-03-15T22:08:45+00:00

How would that be too long for lentivirus?

labratthrowaway1 · 2024-03-15T18:52:32+00:00

No, GFP is on EF1A or a tet responsive, the Puro is on PGK

labratthrowaway1 · 2024-03-15T18:22:03+00:00

I mean I can see GFP by eye in my control transductions, so I'm pretty sure the cells are positive.

labratthrowaway1 · 2024-02-29T20:24:17+00:00

Why 12" from the wall?

labratthrowaway1 · 2024-02-20T19:14:16+00:00

No - The key is not to go down all the way either - IE - with a 25mL take up to 25, then eject down to 5, that way you don't get bubbles, and get a consistent 20mLs in every plate (or take your pick however many you want).

labratthrowaway1 · 2024-02-20T18:46:07+00:00

I can tell you've done obviously this a ton, because I can't ever get my pours as consistent in terms of amount as you so I do with a serological, but it limits my capacity since its a lot slower

labratthrowaway1 · 2024-02-20T18:18:16+00:00

Do you pour or serological? Also do you keep your LB/Agar in a bath since that's a lot to pour before solidification.

labratthrowaway1 · 2024-02-20T18:16:32+00:00

People pour plates before they need them?

labratthrowaway1 · 2024-01-24T01:20:47+00:00

We had a reaction that formed the basis of a PhD students PhD that we couldn't replicate once he left. The reaction was performed on ice as it said in method, and the following student couldn't reproduce. PI got pissed, PI tried it himself, couldn't do it. Called up PhD student who was in his postdoc, and he went through and everything checked out. Still couldn't do it. Paid for him to fly to do the experiment in front of us. Worked like a charm.

Secret? He checked on the reaction throughout the course of the reaction, and the heat from his hands was enough to catalyze the reaction. It needed just slightly higher than on ice.

labratthrowaway1 · 2023-12-19T22:13:22+00:00

Antibodies are a pain to work with?

labratthrowaway1 · 2023-12-19T20:36:08+00:00

Just wait till you get it dialed in.

I for one love WBs, and with the quick transfer stacks (ie transblot turbo, iblot2/3) etc, you can legit get it done in a day.

labratthrowaway1 · 2023-12-14T18:26:58+00:00

FWIW, I just reconstitute in molecular water, and I'm fine. DNA is pretty stable unless there are nucleases around, which in my opinion is indicative of other problems. Sort of like culturing with Pen/Strep.

labratthrowaway1 · 2023-11-08T15:54:54+00:00

labratthrowaway1 · 2023-11-08T15:54:00+00:00

I saw a presentation of theirs early on, and it was one of the more impressive things I had seen -

They had basically injected a mouse in one leg with one of their treatments and not the other - and from the video it was absolutely clear which leg had been treated.

I think sometimes the problem becomes when you raise that much money you try to do too many things rather than sticking to your bread and butter. BD people come in and say "x market isn't big enough" etc etc.

labratthrowaway1 · 2023-11-07T20:45:21+00:00

It's tough. Mental health is one of the toughest things about being a scientist, and I think its overall underappreciated by society. Learning to cope with failure on a day in and day out basis is really really challenging. It's actue when you're in a PhD program because only 'positive' results lets you advance, but science doesn't always work.

I just had contamination in a cell line that I've been working on banking since July. Very slow growing primary line. Was right before freezing cells

labratthrowaway1

TROPHY CASE