Hello Labrats! Has anyone tried isolating proteins, while isolating RNA, using the Zymo Research "Quick-RNA Miniprep Kit"? In principle it is an acetone precipitation but after centrifugation I cannot resuspend the white protein pellet in RIPA buffer, no matter what I try.
I have tried pipetting up and down, vigorous vortexing, incubating the tubes at 70°C for 30 mins and also a slightly different protocol provided by Thermo Fisher, all to no avail. The pellet breaks up in small pieces that don't get resuspended.
Does anyone have any idea what I could try?
I am confused between denaturation and dehydration...
What happens if you leave them for more than 24 hours?
Derived from sheep's wool. pH value: 5.0-5.8. Contains 20-23% of protein. Molecular weight: 1,100-3,300 Da. Preserved with butylene glycol, phenoxyethanol, and ethylhexlglycerin. Clear amber liquid. Characteristic odor. Soluble in water.
Many people online say they just add it to conditioner, but it precipitates out.
Needs a water phase solution to add it to.
I'm not going to worry about pH just yet, I've got the concentration maths done, I've got measurements on how to get it to 0.5%, 2%, and 5%.
I know that it mixes with glycerin, but I'm trying to think of what other things I can use to add to water to thicken this formulation out.
Figured maybe I'd try some gelling agents, I know I've got some agar and gelatin in the kitchen, I've also got some sodium gluconate, but since that is a chelating agent, I'm going to try that last.
I've only got about 50ml hydrolyzed keratin left, and it takes a few weeks to get here in the mail, I don't want to waste it.
Hello fellow lab rats!
I have recently run into a problem in lab and I just can’t seem to find a good explanation for it. I’m trying to conjugate my protein to Streptavidin beads. These proteins contain one unnatural amino acid that has a reactive azide group, so I can then react them with DBCO-biotin in order to biotinylate them. The thing is... I see A LOT of precipitation when I mix the protein with the DBCO-biotin.
I’ve been working with this protein for years now and it is quite stable. I normally react it with a DBCO-PEG molecule without issues or precipitation, so I don’t understand why a DBCO-biotin would result in so much precipitation. Also I work with mutant variants of the protein, so the unnatural amino acid is in different locations and this pattern is seen no matter which mutant I use (granted, some precipitate more than other).
It seems that what’s precipitating is the biotinylated protein, since I’ve seen a decrease in my % of biotinylated protein over time. Even though my final yields are low it’s still enough to do what I need to do, but I’m just super intrigued about this behavior. Am I increasing the effective protein concentration too much by adding the biotin? Am I blocking hydrophilic regions? Adding the DBCO-PEG would theoretically have the same effect though, so that doesn’t seem to be the case... any suggestion or ideas?
I learned the hard way not to clean cell culture media spills directly with ethanol first...Any suggestions for the best way to clean it? It's not terrible, but I've read that the caked on protein can make it harder to keep the surface sterile. Thanks!
After posting in /r/chemistry i was told i should ask here for additional advice, here is the story till now:
I have been doing the same protein purification for the past few months, only with the murine for of the protein. After eluting the protein from my Ni-NTA column (whole process at 4C in the cold room) using a Tris-HCl (pH 8) and imidazole-based buffer in 10ml fractions, i put the fractions on ice and left them in the cold room for the past two days. Today i wanted to continue and see that the fractions where i predicted the protein to be had big pellets. Is this reversible? How? I need to dialyse the stuff for the next two days. What i did right now was dilute the fractions (added 5ml of elution buffer) and left them to shake at room temperature.
Day 2
I performed the dialysis with the first buffer now (4C), the precipitates are still there but they somehow changed in form. Will do the dialysis with the next buffer tomorrow and see what i get. Do you think dialysis at room temperature might be better than in the cold room?
Edit: I wanted to try the next dialysis step with L-Arginine because i have been told/read that it stabilizes proteins or refolds them. After about 2 hours in this new buffer (1.5M L-Arginine - somehow expensive that stuff O.O) it looks like the precipitates are gone. Before I start jumping around i will wait for tomorrow, i will keep you posted
Edit2: After over-night dialysis with a buffer containing L-Arginine, around 80% of the precipitates vanished. Do you have any idea what I could try to get rid of the rest? or should i just centrifuge the stuff, take the supernatant and be happy with what i have? :)
I have not been able to find any information relating to actual levels of carrageenan remaining dissolved in beer if it is used to floc proteins. My wife was told by one of her doctors to avoid carageenan due some chronic inflammation that she has had. I would be fine doing cold crashes to clarify our homebrew, but it made me curious.
Any food/beer scientists out there who could comment on this? Any recommendations for alternate clarifying agents?
I don't want to step on anybody's toes here, but the amount of non-dad jokes here in this subreddit really annoys me. First of all, dad jokes CAN be NSFW, it clearly says so in the sub rules. Secondly, it doesn't automatically make it a dad joke if it's from a conversation between you and your child. Most importantly, the jokes that your CHILDREN tell YOU are not dad jokes. The point of a dad joke is that it's so cheesy only a dad who's trying to be funny would make such a joke. That's it. They are stupid plays on words, lame puns and so on. There has to be a clever pun or wordplay for it to be considered a dad joke.
Again, to all the fellow dads, I apologise if I'm sounding too harsh. But I just needed to get it off my chest.
Alot of great jokes get posted here! However just because you have a joke, doesn't mean it's a dad joke.
THIS IS NOT ABOUT NSFW, THIS IS ABOUT LONG JOKES, BLONDE JOKES, SEXUAL JOKES, KNOCK KNOCK JOKES, POLITICAL JOKES, ETC BEING POSTED IN A DAD JOKE SUB
Try telling these sexual jokes that get posted here, to your kid and see how your spouse likes it.. if that goes well, Try telling one of your friends kid about your sex life being like Coca cola, first it was normal, than light and now zero , and see if the parents are OK with you telling their kid the "dad joke"
I'm not even referencing the NSFW, I'm saying Dad jokes are corny, and sometimes painful, not sexual
So check out r/jokes for all types of jokes
r/unclejokes for dirty jokes
r/3amjokes for real weird and alot of OC
r/cleandadjokes If your really sick of seeing not dad jokes in r/dadjokes
Punchline !
Edit: this is not a post about NSFW , This is about jokes, knock knock jokes, blonde jokes, political jokes etc being posted in a dad joke sub
Edit 2: don't touch the thermostat
This is something I see come up from time to time in various groups so I wrote up something on the subject. I hope some of you find this helpful, and if there are any issues with what I’ve posted feel free to let me know.
Introduction: Managing Type 1 Diabetes in an emergency or survival situation is a significant challenge, but it is not impossible. In this document, we will go over some of the preparations you can make to give yourself the best chance of survival.
Test Meters: A fundamental problem with storing diabetic testing strips over long periods of time is that they expire and become unreliable or simply do not work at all. There are different products on the market called CGMs or Continuous Glucose Monitors; the most useful of these for our purposes is the Freestyle Libre, which is essentially a CGM and works similarly but only delivers readings when scanned and not continuously. These systems generally depend on electronics to obtain readings, so having a method of powering them (such as a generator as discussed below) is important. Unfortunately, these devices have the same shortcoming that standard meters do; it should be noted that the Libre uses sensors that must be replaced after 2 weeks, and expire in 6 months but studies have shown that they remain effective 18 months past this date, which means that although it requires stockpiling, it is still an alternate option to standard test strips.
All that being said, it is still a good idea to also have a standard monitor and test strips in your stock in case of emergencies, even if you have a CGM. Ensure that you stock the brand with the test strips with the longest expiration dates. I know, for example, Accu-Chek have an 18 month expiration date from manufacture, but am unsure how these rate in terms of longevity among all competitors. This can always change over time, so make sure to do research to see which currently available test strips last the longest before expiring. I have conducted limited testing on Accu-Check strips that were 2 years expired and they were still accurate compared to in-date control strips. This may not be representative of all Accu-Check strips but I felt it worth sharing.
Stockpiling: Stockpiling Insulin can be accomplished relatively easily. Currently, in all states other than Indiana, one can get Insulin over-the-counter at Wal*Mart locations for approximately $25 per vial. This offers a convenient and easy way to create a sizable stockpile of insulin, but
... keep reading on reddit ➡Do your worst!
How the hell am I suppose to know when it’s raining in Sweden?
Ants don’t even have the concept fathers, let alone a good dad joke. Keep r/ants out of my r/dadjokes.
But no, seriously. I understand rule 7 is great to have intelligent discussion, but sometimes it feels like 1 in 10 posts here is someone getting upset about the jokes on this sub. Let the mods deal with it, they regulate the sub.
They were cooked in Greece.
I'm surprised it hasn't decade.
Now that I listen to albums, I hardly ever leave the house.
Don't you know a good pun is its own reword?
Two muffins are in an oven, one muffin looks at the other and says "is it just me, or is it hot in here?"
Then the other muffin says "AHH, TALKING MUFFIN!!!"
For context I'm a Refuse Driver (Garbage man) & today I was on food waste. After I'd tipped I was checking the wagon for any defects when I spotted a lone pea balanced on the lifts.
I said "hey look, an escaPEA"
No one near me but it didn't half make me laugh for a good hour or so!
Edit: I can't believe how much this has blown up. Thank you everyone I've had a blast reading through the replies 😂
It really does, I swear!
And now I’m cannelloni
Because she wanted to see the task manager.
