Hi my name is Melody (PhD) and welcome to my channel dedicated to science and biochemistry. My goal is to make science easy and I hope you find the content interesting and inspiring.
great and informative video. Have you done thermal shift assay using qPCR it would be nice if you share a tutorial for TSA to find out ligand affinity for protein.
Do you store lipid stocks by aliquoting them to prevent melting the entire bottle before weighing? Does this method help maintain the integrity of the lipids? Alternatively, can they simply be stored at -20°C?
@@biochemistmelo Wishing you success in your job! 🤞 🍀 I am also a TEM engineer with a background in Materials Science, but now I am a cryo-EM laboratory director working with biologists ☺️
@farzaadkhaan wow that is amazing. Microscopy became my favorite technique in my PhD, so for my post-doc I hope to find a lab where I can learn cryo-EM.
I would highly advise you wearing gloves at all times and following a more detailed cleaning procedure. From my experience this will ruin you quite some work you are doing here.
I've been doing this for 8 years and always check my liposomes with TEM. I wear gloves when I'm working because if I didn't I would see it on the TEM. You should worry about yourself.
There are several issues with operating the TEM like that. Please be careful. I would carefully follow this other video instead: ru-vid.com/video/%D0%B2%D0%B8%D0%B4%D0%B5%D0%BE-noE_F1o1XmA.html
Hello I'm a high school senior and I was wondering if possible to set up a biochemist informational interview regarding your career and further insight into it?
I understand you'll probably be unable to do that I'll put the questions down. -Name of person, -Company they work for, -Years of experience, -Contact, -if available, business card. Questions: 1. How did you end up in this career? 2.How do you handle stress/mental health? 3. What qualifications are needed for your job?Do you have any pointers or suggestions for me when pursuing this career? 4. Do you have any regrets about this career choice?Why or why not? What are some negative things about this career choice?
Can you use this one protein samples which contains lots of protein? Or only for a single purified protein? Like for example I’m using a serum sample after depleting albumin and igGs. After performing Western blot I found that the bands were much larger than target protein size. I also tested it with different antibodies but the results were always much larger (almost double). So I thought I may need to take another look at concentration measurements cause perhaps the sample is too concentrated so here I am.
@@biochemistmelo I want to determine the difference of a specific protein level between patients and healthy controls in serum samples using western blot.
Hi, thank you for this excelent video! I have some questions (probably silly ones). Situation: a experiment where i used a buffer as blank and the software was set to execute baseline correction. Does it means the further spectra obtained from samples will be already subtracted from buffer? Do i still need to click on baseline to manually adjust it after smoothing? If so, how i do the adjusting?
That is part of the rehydration step. I don't recommend skipping it. You can see a clear difference in how the lipid looks. In the first lyophilization step it looks very dense but after the second, it will look very fluffy and you know that it's been rehydrated.
Hi, 1. For the elution, do we need to try a few gradients to see which concentration of imidazole is ideal for elution? 2. Does manual purification possess any major disadvantages compared to using the machine (FPLC)? 3. Can we purify ion exchange and size exclusion chromatography manually without the help of the machine (AKTA purifier/ FPLC)?
Hi dear, 1. The concentration of imidazole for the wash and the elution solutions are pretty standard and you don't have to test it. The wash solution has a little to take off impurities and the elution has a lot to completely elute your protein off the column. 2&3. columns for size exclusion and HPLC and such come already pre-packed and you don't want to mess with their packings(if you do it messes with the seperation) so they must be used with a machine that can pump liquid through them. The packing doesn't really matter for nickel affinity so you can just pipette the resin onto a column and use gravity.
Thanks for the video! after complete drying of the lipid layers and adding buffer, my solution is not milky and is not suspended very well, what should I do ? I am afraid of blocking the extruder by the lipids
@Avayekaveh I never worked with pure cholesterol, only 15-30% in phospholipid mixes and mix them in the beginning in chloroform then evaporate, freeze-dry, hydrate, freeze-dry, add buffer and extrude. If you have to mix it with a lipid down the road, it is easier to do it earlier.
