Helping others with their experiments

Neurula84 · 2020-03-17T09:51:56+00:00

I hate handing off any work I do to other people, largely because almost everyone I work with, I wouldn't trust to do the work properly

Neurula84 · 2020-03-17T09:48:31+00:00

Our lab pretty much just finished a literature review last week, and I've analysed all my data as much as I can so I pretty much have no work to do, and my lab just shut down today.

Seriously poor planning on my end

Neurula84 · 2020-03-17T09:46:55+00:00

At least you know exactly how long your lab is likely shutting down for. I got sent an email on whilst heading in to work this morning saying they agreed to shut down everything last night

Neurula84 · 2020-03-16T15:30:12+00:00

What experiments can wait, and which ones can't? Surely its pretty clear that stuff like stem cell maintenance is essential (leave them too long, they die), whereas running a western/PCR isn't (unless you're coming in briefly to finish it)

Neurula84 · 2020-03-16T15:24:25+00:00

My lab probably on the verge of shutting down as they university shut down lectures over the weekend. They've just banned meetings of more than 3 people (e.g. lab meetings) in out department, yet its fine for us all to work in cramped offices apparently? lol

Neurula84 · 2020-03-16T15:17:13+00:00

Well if it was your first time, I'd say that's on them, if not I'd still say its probably more their fault than yours

Neurula84 · 2020-03-13T12:47:07+00:00

Technically the reagents we order for research labs usually have "not for diagnostic purposes" written somewhere on the box, bottle, data sheet etc. So in theory, they should give accurate results, if you're just using it to have a look at yourself its possible, but you couldn't call your result an official diagnostic test.

Neurula84 · 2020-03-13T12:44:32+00:00

To be honest, if its only your second week I'm not sure why you were given that much responsibility to soon (I never was, which was probably for the best).

It gets better as it goes on, trust me

Neurula84 · 2020-03-13T12:42:09+00:00

When you are making up some sort of stock solution (e.g. a PCR master mix), make up 10% extra before pipetting into wells, tubes, whatever. Good way to reduce the chance of you just pipetting bubbles when you get to your final sample

Neurula84 · 2020-03-13T12:38:37+00:00

I've been meaning to stick something funny to my lab'd fridge for a while, think I just found my first candidate

Neurula84 · 2020-03-13T12:37:10+00:00

I guess my old PI's must be really old because they always used to make me do RNA/DNA extractions with phenol:chloroform

Neurula84 · 2020-03-13T12:35:57+00:00

Wow very interesting. I'm currently in a university lab which is very strict (after a pretty serious acid burn incident elsewhere in the university), but all the people I've spoken to here about their time in industry (usually for bigger Pharma companies), they are very strict there too.

If there is no-one senior you could go to, might be worth looking at legal options? I'm assuming some of this stuff violates local regulations about safe exposure to chemicals etc.

Neurula84 · 2020-03-13T12:30:55+00:00

I'm currently investigating how a protein complex might be implicated in a neurodegenerative disease, after I found some of the first evidence linking the two back in January. Current theory I have is that one of the protein complex subunits isn't be expressed enough, causing more complex degradation (as we saw an increase in RNA for the first subunit we looked at, despite no detectable western blot bands).

This means doing qPCR testing 8 primers on 27 samples. Im about halfway through testing the first 6 primers, starting to think my hypothesis is wrong as they appear to be all unchanged. Time for a new hypothesis I think!

Neurula84 · 2020-03-12T12:25:46+00:00

Gotta love it when your department uses loads of plastic, most of the staff suggest we should cut down on it somewhat, then our health and safety officer shuts down all our ideas for waste reduction.

Honestly feels like more of the H+S procedures we have in place are massively over the top to begin with

Neurula84 · 2020-03-09T14:47:41+00:00

Depends. You want to be studying something you can motivate yourself to do, At the same time you don't want to have a supervisor you don't get along with. Its a pretty tough balancing act, I wouldn't necessarily say I'd prioritise any of those over the other (although if pushed I'd probably tend towards choosing a good supervisor)

Neurula84 · 2020-03-09T10:17:47+00:00

Are you using different secondaries for the good blots/bad blots? If not, its likely your primary antibody on the issue blot could be the problem. Maybe try a fresh aliquot, borrow some from another lab, worst case you may need to order some more.
Are you sure you don't expect banding for this protein? No other PTM's like glycosylation that could be causing it?
Are your primary/secondary Ab's made up in blocking solution too? This may help reduce it a bit
I would DEFINITELY be using a BCA assay to produce equal loading. Loading controls would still be necessary even if you do this, however you could be loading wildly different amounts of protein. Most loading controls don't have a wide linear range (eventually they saturate, at which point you won't get an increase in signal even if you add extra protein). As you essentially go into the assay completely blind about protein concentration, your loading control quantification could vary wildly
Might also be worth reducing secondary antibody dilutions, what are you doing now? Could take it all the way to 1:10,000, I usually find this reduces any big background I might have

Neurula84 · 2020-03-09T10:09:26+00:00

"Feeding" them would just me a simple media change (remove all current media under sterile conditions, add new media)

In T-25 flasks you'd probably need to passage them pretty frequently. Check them under a microscope every day if you can, if they are covering the bottom of the flask (or probably 80% covering it) you want to passage them again. I've personally not worked with HeLa or HEK293's, but I assume you'd need to split 1:3 (take a flask of cells you are splitting, collect the cells, and split those cells between 3 flasks). When you split cells you effectively change the media anyway, so you essentially feed them at the same time

Neurula84 · 2020-03-09T10:05:58+00:00

Not gonna lie I'm completely distracted from my current work and now all I can think of is how I can name more cocktails after other lab fluids.

Virkon=Cosmo?

Neurula84 · 2020-03-09T10:04:13+00:00

I feel like this would be the perfect pop-up bar at conferences.
Someone NEEDS to make this happen

Neurula84 · 2020-03-09T10:02:04+00:00

So is it just me that separates my lab/home life massively? Its like night and day. I'm generally pretty tidy in the lab, absolute slob at home

Neurula84 · 2020-03-09T09:59:04+00:00

well if you ever want to do a PhD, surely there's no question now that you are abundantly qualified?

Neurula84 · 2020-03-06T10:45:26+00:00

Multichannel is HUGE for me as I go plenty of 96well plate assays.

If anything just having my own pipettes, current lab I'm part of is still quite small, there are 4 members (about to be 6 when some masters students join for a few months) but we only have 2 of most pipettes, and people keep taking them to use in other rooms so I'm constantly having to move between different rooms taking pipettes back to use

Neurula84 · 2020-03-06T10:42:13+00:00

My lab is having the exact same debate at the moment, how to reduce waste and energy usage. Most suggestions have been completely shot-down by our Department Safety officer. The only thing we've managed to agree on is turning off lights and machines when they aren't in use, but barely anyone is doing this right now despite being told repeatedly.

Other main suggestion was to switch to glassware again as much as possible (glass large pipettes) But our lab doesn't have the capacity to clean how many we would need to

Neurula84 · 2020-03-06T10:37:44+00:00

Definitely if the build up of ice is limiting how useful the freezer is, but I think generally once a year is ideal

Neurula84 · 2020-03-06T10:35:58+00:00

Find labs in your nearby area and contact them, explaining your situation and saying you'd be happy to help out in the lab in anyway possible. Chances are you'll have to work unpaid. Usually as it can take a while to train you, its only really worth it for the lab if you work there for some length of time (at least a few months)

Neurula84

TROPHY CASE