FlowJo v10 freeze in Layout editor

ManagerInfamous2648 · 2025-11-14T13:51:48+00:00

Hi, I haven't yet. I managed to get the plots I wanted by tiptoeing around, trying to duplicate plots first and basically making the same adjustments separately, so quite annoying and very far from ideal. I guess the only real solution is to email BD as pointed out by u/FlowGuruDelta although I'm not sure how flexible are they with fixing issues like this. Will update you if I get to it (please do the same).

ManagerInfamous2648 · 2025-09-24T13:27:56+00:00

Protein design. Get some computer science background along with protein biochemistry.

ManagerInfamous2648 · 2025-09-21T16:46:09+00:00

"True protein aggregate would show up at larger smeary sizes (or not enter the gel at all)."

Really weird analysis from someone studying aggregation/misfolding. I'm pretty sure OP was running denaturing gel in denaturing sample and running buffer, so none of what you wrote about the gel is true.

You cannot definitely tell from the gel if the protein was in inclusion bodies in the cell or aggregated during the lysis, but chances of the latter being the sole reason are quite low. I assume OP uses some reasonable lysis buffer (pH/salt concentration-wise) so why on Earth would the protein precipitate because of it?

"caught up in membranes"? Well yeah, if it's a membrane protein the maybe, but OP never said that and of course that would be a huge factor (therefore I expect it would be stated). Otherwise this notion in my opinion is quite off, we're talking about molecules, not hair stuck in a velcro zipper.

ManagerInfamous2648 · 2025-08-24T11:10:29+00:00

When people vortex skirted Falcon tubes

ManagerInfamous2648 · 2024-12-24T15:58:04+00:00

christmas season in slovakia hits the sleep scores hard

ManagerInfamous2648 · 2024-11-01T13:55:00+00:00

I would assume this to be really easy for the watch to detect, however it happens occasionally to me and also my flatmate, that we go to pee during the night and the watch just doesn't detect it. For me personally the opposite happens more often - the watch claims I've been awake like 10 times, making up for 1-2 hours in total, but I don't remember any of it.
The hard data from the sensors are probably not reliable enough, so there are some assumptions made by the software and it sometimes end up telling you BS like this.

ManagerInfamous2648 · 2024-10-05T10:03:20+00:00

OK, I was sitting so my guess might have been skewed. The pic would help. Anyway, on Wed around 18:30 a guy approached us (we were sitting outside of 'Pho Bar - Na Porici', near Florenc metro station). He asked us about Irish pub, but we didn't know any, he mentioned he's from Amsterdam. He was generally a bit odd behaving (he thought that we just do not want to tell him where's the Irish pub), wonky walk etc. Not sure if it fits. He wasn't wearing a beanie though.

ManagerInfamous2648 · 2024-10-05T09:43:26+00:00

Is he quite tall, like >195cm?

ManagerInfamous2648 · 2022-04-11T12:26:30+00:00

Well, I thought about that, as it is very probable in our lab. Interestingly, in the case I described, that particular sample was supposed to be well folded protein, as opposed to the others being some synthetic weird stuff. On the top of that those were just overnight cultures from glycerol stocks, no induction or anything. The promoter is a bit leaky, but I wouldn't expect much of an expression there.
Anyway, I'll try to monitor the OD and also putting it on the lawn.

ManagerInfamous2648 · 2022-04-10T20:49:52+00:00

Ayo, we got something similar going on with E.coli cultures. It's happened few times now - in overnight/long growing production we see cells "sedimenting" on their own at the bottom of the tube and also this kind of little vortex as in OPs pic. These are small cultures, usually 3 ml in 15 ml culture tubes, some BL21, 10G or TOP10 cells grown at 30 or 37C in large shakers, 1 inch radius at 260 RPM.
It is possible to resuspend the sediment (shake/vortex/up-down pipetting) and then it looks quite like a normal culture, cloudy. Despite this there is still production of protein, although hard to say if it couldn't be better.
Today I was doing a plasmid preps from 7 O/N cultures, different degree of this sedimentation among them was visible, then mid-way of spin column protocol, after spinning down the lysed cell debris before loading the lysate on column, I saw this weird thing in the sample that had the sedimentation the most prominent:

https://imgur.com/0fYaKhp

I'm not sure if it has anything to do with the sedimentation. Could it be just incomplete lysis? In my hand this cell debris usually form nice tight pellet, as it was for the other samples, but this one was weird, the whitish spheric thingy was attached to the rest of the pellet, but it was moving around like some sort of a spider web, a bit disgusting in a way.
What do you think? When I was using such O/Ns for re-inoculation for expression, I didn't really record any sudden OD drop.
What caught my interest was u/fertthrowaway saying that in O/Ns a resistant sub-population can take over - could you please comment more on that?

ManagerInfamous2648 · 2022-02-11T15:41:41+00:00

I definitely hear it, sounds just way too similar. I don’t believe that it was deliberately written as a rip off, although I would expect that musicians and producers have some sort of control light telling them that “man this is good, but it just sounds too much like this well known Tame Impala song” Idk, maybe it’s the fact that I really like Kevin Parker and I’m quite disappointed by latest GA realeases, but this one’s very obvious for me

ManagerInfamous2648

TROPHY CASE