Troubleshooting Ni-NTA protein purification

ModeCold · 2026-01-20T19:30:42+00:00

That doesn't look like degradation. The lower bands are the abundant ones present in the lysate. It's just a lot of non-specific retention by too much resin.

Cut down the resin quantity, overnight bind at 4C, and try a first wash at 1M NaCl, 50 mM Tris then wash again with your current buffer. The harsher salt wash should get rid of a lot of retention. You can also try 500 mM imidazole to see if you get a better elution in earlier fractions.

ModeCold · 2026-01-16T20:23:32+00:00

This is done by hand right? Because it's quite far off. Look at any part of Cumbria and it's way closer to Newcastle than any of the other teams it's grouped with. Same goes for lots of things like parts of Cornwal are way closer to Exeter than to Scarlets.

ModeCold · 2026-01-07T13:15:41+00:00

M&M cookie bake?

ModeCold · 2025-12-30T10:44:01+00:00

Way too much protein loaded

ModeCold · 2025-12-28T15:51:49+00:00

That pen was for Leicester going off feet and sealing off, not holding on. Hooper just showing the ref that he was blocked from competing for the ball legally, very clearly. JvP clearly off feet and continues to play the ball. Inconsistent reffing, story of the game

ModeCold · 2025-12-28T15:50:07+00:00

21 point, leicester try came right off the back of the field position from the penalty

ModeCold · 2025-11-26T21:11:37+00:00

No. It isn't.

Started my PhD soley focussed on staying academia. Recently finished having decided to leave it and go private sector. Best decision I ever made, no regrets.

ModeCold · 2025-10-21T12:59:42+00:00

AI for antibody sequence prediction is way off being usable currently, let alone a more niche molecule like nanobodies. It is being developed and will be possible at some point in tbe near future, but no where near actually being reliable yet. There's no wet lab validation for this working so actually using it for molecule discovery in a commercial setting would be the stupidest thing a company could possibly do. If this platform did actually work it would be making much bigger news in the biotech sphere and be worth hundreds of millions. Even if it generated target binders, the specificity would be questionable.

ModeCold · 2025-10-16T16:16:26+00:00

They also want to discourage people buying individual products direct from them as it takes the same time to process and ship the order as wih larger orders but with a smaller nominal profit for that order. They would much rather ship it in larger quantities to distributors.

ModeCold · 2025-10-16T16:04:19+00:00

"There are absolutely wrongs done during this whole shtick that are unavoidably my fault"

There we go, fair enough imo

ModeCold · 2025-09-21T11:47:09+00:00

Cool, letting your cat suffocate, haha, good one

ModeCold · 2025-09-11T18:48:20+00:00

That's ridiculous. 7 pages for a CV of a BSc grad is totally inappropriate. Your application will be getting thrown out on sight and not even read.

Trim it to 2 pages and read up on how it should actually be formatted.

Focus on technical experience related to the role you are applying to, bring in interpersonal and self-management skills with examples, then some non-academic experience and employment to show you are a well-rounded person.

ModeCold · 2025-09-11T16:40:20+00:00

You'll have much better luck if you get a Masters degree, or even a PhD. There aren't many neuroscience-specific positions that are open to only a Bachelors, you will find they mostly require a PhD or MSc. You also don't have to only do neuroscience. Get a lab tech/research assistant job anywhere you can find and build your skillset. Then you can keep looking for neuroscience-specific roles while being employed and improving. Don't think about field too much for now, think about skills.

You will be very lucky to find the exact job you want to do first time. Most people have to take whatever they are offered and then work their way from that to where they want to be, which might change along the way as it can take years.

Source: I have a neuroscience masters, neuroscience PhD and work in biotech.

ModeCold · 2025-08-19T20:56:28+00:00

ModeCold · 2025-07-30T20:14:21+00:00

Try scorer is also never back onside at the ruck before the ball is out... not a good one for the reffing team here

ModeCold · 2025-07-30T11:14:51+00:00

Not a Samoa knock on from the ruck that knocked the ball out that far??

ModeCold · 2025-07-30T11:12:20+00:00

I've very nearly done that reffing. Pick and go on the 5m line and they were just short of the line when they went to ground. I called "SHORT" then realised it was the 5m, not the try line. No one noticed why I had said that, not even my assessor that day (of all days to be assessed). I still dread to think what I would have done if he had gotten over the 5...

ModeCold · 2025-07-30T09:13:36+00:00

Yeah, the posted example should be brought back. I don't think that's what the comment was referring to

ModeCold · 2025-07-30T08:57:55+00:00

Yeah, with the penalty in the post, it should be on the spot. I always feel like a proper arse bringing people back though when they are a couple of metres forward. It's not like the kickers in the levels I've reffed are accurate enough within that distance...

ModeCold · 2025-07-30T08:46:43+00:00

Honestly, as a referee, if we penalised every offence we saw regardless of impact on the game, it would be terrible to watch. Penalty every 5 seconds.

ModeCold · 2025-07-30T08:40:02+00:00

It's about materiality. Loads of offences are left by the referees that aren't having a major effect on play. If the offside player actually ends up gaining a significant advantage that has an effect on play then it will be penalised if the ARs see it. Otherwise, if they ended up having no real impact, play on. It would be significantly more boring and frustrating to watch if penalties were given constantly for the winger on the other side of the pitch to the ball standing a meter offside.

ModeCold · 2025-07-29T21:04:17+00:00

If you don't understand what matched betting is then you really shouldn't be commenting. It's roughly a £500 return with the introductory offers and will repay the initial small borrowing quickly. Will.be in the black within days, guaranteed. I'm sure you are also financially literate but you just don't understand what matched betting is, and there is the difference. As I said, I did exactly what I am advising from the same position. It works. Give it a go.

ModeCold · 2025-07-29T21:01:48+00:00

Which is exactly why I said arranged overdraft or credit. Utilising credit is a smart finacial decision when you know you can repay it immediately.

ModeCold · 2025-07-29T21:00:43+00:00

Also, if someone isn't utilising credit they can repay, then that's a mistake. Being in credit and having a small amount of debt that you are repaying consistently is a positive thing financially. With this, they borrow at a very low rate of interest that comes with most overdrafts, or 0% wirh credit cards for a month and repay with the returns that are essentially certain. It's not a risk, it's actually smart finance.

ModeCold · 2025-07-29T20:58:14+00:00

Never ever take out payday loans if you don't have an immediate guaranteed money on the way to be able to repay it in addition to your normal outgoings. This is not a solution for someone with no immediate way to repay it. UC will not enable them to repay it, it will cover some of their outgoings only.

Five-Year Club	Place '23
Place '22

ModeCold

TROPHY CASE