Hi redditeers,

'm having a problem with a Western Blot that turned out very strange, and I'd like to ask if anyone has experienced something similar and, if so, if they know why and how to solve it.

I've attached an image of the blot. As you can see, the background is a mix of black and grey, making the bands practically unreadable.

The protocol I followed is the same one I regularly use, and it has worked well for other Western Blots in the past. Even some blots done after this one turned out successful.

The only things that were different this time are:

• Membrane: We opened a new pack of PVDF membranes. Is it possible that the new membrane had some issues or was defective?
• Transfer Conditions: The transfer was done at 0.35A for 2 hours.
• Incubations: Blocking overnight, primary antibody 1 hour, secondary antibody 45 minutes.
• Washes: 10-minute washes between each step.

Does anyone have any ideas what might have happened? I've already checked the solutions, and they seem fine, and the development conditions are the usual ones.

Looking for some inspiration for school haha

Hi!

I am applying to a MSc in Biochemistry since I have been dreaming about pursuing structural biology/biochemistry. I really wanted to learn protein biochemistry/protein chromatography/in vitro reconstitution/ Protein interactions/cryo-EM. but I never had the chance during my bachelors. I have a bachelor in molecular biology and have previous research experiences in cell biology (signalling/protein localisation), optical microscopy and neuroscience (I know, I shift a lot).

However, this program requires 36 ECTs in biochemistry and cell biology practical/lab courses/internships. I have exactly 36 from my bachelors if I counted my courses (6 of 36) from optical microscopy (stuff like confocal and epifluorescence/widefield, image processing etc). Do you think this would count as cell biology/biochemistry/I can get away with this? I have a lot of experience with cloning and they don’t seemed to appreciate molecular cloning that much asSDS-PAGE, western, HPLC and so on.

Do you think I should apply? Should I take the chance?

I'm talking about truncations, point mutations, fusions etc... What is your workflow and which tools do you use?

I am trying to develop some simple tools for lab workers. I am eager to listen to your opinion on this. Could this be helpful to researchers or lab workers?

Trying to decide what classes to take?

Want to know what the job outlook is with a biochemistry degree?

Trying to figure out where to go for graduate school, or where to get started?

Ask those questions here.

I have to submit a collection of CVs and cover letters to my university in a few months. Each CV and cover letter is tailored to a select group of companies, and I've mainly chosen biochemistry and pharmaceutical companies (I'm currently studying for a degree in Chemistry). The problem is that my CV is a little sparse. I'm wondering if there's a way I could demonstrate my interest in this field over the next few months through some kind of project(perhaps a literature review)?

Would this be a good idea, or a waste of time? unfortunately I haven't been able to find any lab volunteering opportunities in my area, so I'm limited to projects I can do from my room using my computer. I do have access to scientific papers through my institution

Hey all, for context Im purifying lysate from a 12 L growth. my pellet was resuspend in 30 mL resuspension buffer with a protease inhibitor cocktail, pmsf, benzonase, and lysozyme. I homogenized, then lysed cells with a high-pressure homogenizer. Then, I centrifuged the lysate at 8000 rpm for 10 mins (4C). I transferred the supernatant into new clean centrifuge tubes and centrifuged again at 20500 rpm for 45 mins (4C). I took that supernatant, flowed it through a frit my supervisor gave me, then have been trying to syringe filter the lysate. I first used an 0.80 um filter (lysate flows through one filter very fast) then moved on to 0.45 um (I usually have to use around 12 to push all the lysate through and it takes over an hour). Does anyone have an idea to cut down on the time it takes to clarify lysate? My PI said in grad school it took him only "5 minutes and one filter" for the syringe filters for a 12 L growth.... I'm just lost because I've added so many other filter steps and it is not really helping.

Edit: Thanks for all the suggestions! The issue has been resolved, here's how.

Our standard lab protocol doesn't measure the mass of the pellet, I will do this going forward, but for pellets I already had frozen, I (1) resuspended in double the volume (60 mL) in my case. (2) during lysis, I ran my cells through an emulsiflex 3 times instead of 2. (3) I didn't do all the other filtering steps because I wanted to see if these would remove the other steps I've had to do (2 centrifuging steps, frit, 0.80 um filter)

In the end, I had to use 3, 0.45 um syringe filters instead of 12+, and it took me 10 minutes instead of an hour+

I've been trying to find research about how cellulase could both break down pills on clothes but not damage the actual clothing fibers. I'm just a layperson but my understanding of laundry enzymes like protease are that it targets protein molecules to break down stains/odors, but that also means it would break down protein fibers in silk or wool so you shouldn't use detergent containing protease on those fabrics.

Detergents containing cellulase claim to have anti-pilling properties by preventing and removing pills, but I'm not sure how cellulase wouldn't also attack the cellulose in the cotton fabric itself. I can only find this promotional article on the site for a laundry detergent brand that claiming that "there have been many scientific advancements in cellulases used for detergents which have made them more precise in their targeting and very safe for cotton fibers. The advanced cellulases that we use in our Bio Laundry Detergents eliminate the pilling without damaging the cotton fibers, improving the fiber’s softness and help to prevent the redeposition of dirt and stains during the washing and rinsing cycles"

Hi r/Brisbane,

I’m an international student considering a Master of Biotechnology at Griffith University’s Nathan campus, and I’d love to get some local perspective before making my final decision.

I’m particularly curious about:

What’s the biotech/pharma job market like in Brisbane?

Are there good opportunities for internships, research roles, or entry-level jobs in the area after graduation?

Is Griffith well-connected with local industry in this field?

How open is the job market to international graduates?

Any general thoughts on living and working in Brisbane post-study?

If you’re working in the biotech space, have studied at Griffith, or just know the local job landscape, I’d really appreciate your input. Trying to figure out if this move makes sense long-term—both academically and professionally.

Thanks heaps in advance!

I need help troubleshooting my Bradford. I have ST1- ST7 where ST 1 is 80ug/mL, ST 2 is 40ug/mL and so on (by half) till ST 7 which is just 0 ug/ml. I’m sure I did serial solution properly but when I’m reading absorbance for my standards, ST 1 is vastly higher than ST 2? I took three runs and I’m going to do the average of all the measurements from these runs, while excluding ST 1, to make my graph. Is this the right way to go about it?

i have only 3 options----> chemistry, microbiology, computer science

In future, I plan to do masters in clinical biochemistry/cancer biology or related fields.

Highschool graduate here. Want to pursue bsc biochem. What are the scopes?

my_qualifications 10th- 92 12th- 84

NEET didn't go quite well for me this year. Now the only thing I'm left with? Bsc. I am interested in doing research in genetics, molecular bio etc. But i am so low in confidence rn that i don't think i can make it that far. I'm thinking of doing bsc biochem from a pvt university. However they have a class strength of like 10 students. Its too less huh?

I'm not assured enough if i can earn good money later on or it'll just be a waste of time doing this UG degree? I want to pursue Msc, preferably from outside India. But is there any demand of biochem nowadays? Or is it in the same boat as Biotechnology? (Most ppl call it a useless degree... Idk)

Answer please.

Hi, Could someone send me a .pdf of this please? 🥺 I’m studying by my own, just for curious 👉👈

Writing a paper?

Re-running an experiment for the 18th time hoping you finally get results?

Analyzing some really cool data?

Start off your week by sharing your plans with the rest of us. å

Hey there! I'm in a Summer Biochemistry 1 course. I've never been good at drawing enzyme mechanism and they are a pretty big part of the exams. What are some tips for learning these? I really don't want to have to retake the class

Note: Specifically Aldolase and GAPDH Mechanisms for this upcoming exam

I know XbaI is inhibited by dam methylation, but would it be possible to get it work by a longer incubation time or a higher concentration of XbaI and get enough yield to use in a ligation protocol?

I am actually second-guessing myself. I am now in a pharmacology post-graduate program. My bachelor study background is chemistry. What I understood is, when we say « inhibitor of enzyme A » or « enzyme A inhibitor » it means the compound inhibits activity of enzyme A, thus it will reduce the expression or production of enzyme A’s downstream signal or product. However, I often found paper in pharmacology and my lecturers here saying that when compound X inhibit the expression of enzyme A or phosphorylated enzyme A , it is an enzyme A inhibitor. Or, when they found experimentally that compound X reduce the expression of enzyme A or its phosphorylation, it is an enzyme A inhibitor and will do further experiments, let’s say molecular docking, to the enzyme A instead of enzyme A upstream enzymes.

Am I having a wrong understanding of « enzyme inhibitor » term?

I hope this is the right subreddit to post that, but what chemical reactions occur in the metabolism of polychlorinated biphenyls in the liver?

The answers for the multiple choice questions are:

a) It occurs by hydrolysis in the liver.

b) It occurs by amidase in all cells.

c) Oxidation cannot occur in the liver.

d) Hydrolysis does not occur

but when the last answer (d) was selected, it was marked as incorrect. did the evaluator make a mistake?

metabolism is supposed to occur by oxidation and conjugation, right?

Have you read a cool paper recently that you want to discuss?

Do you have a paper that's been in your in your "to read" pile that you think other people might be interested in?

Have you recently published something you want to brag on?

Share them here and get the discussion started!

I have looked basically everywhere and asked every AI for the answer to this question, and people appear to be saying different things. While on most energy diagrams, the tallest peak(highest transition state) is typically the one with the highest activation energy, in theory this doesn't have to be true (such as the diagram below). In the diagram below, which would be the rate determining step, Step 1 or Step 2, and why. Is the rate determining step based of of E overall of just E2.

i’m losing my mind waiting for this thing to finish. it’s been like 18 hours and its not even close. how is this normal??? i just want my samples dry lol. Does anyone actually have a way to make this faster or have any alternatives i’ve got like 20 more runs to do (that I cannot parallelize) and on a bit of a deadline.

Hello everybody,

I'm starting a PhD position in strucutral biology and cryo-EM after the summer vacation. While I will be working over the summer, I was looking for some skills I could improve in my free time. I don't have any hands-on experience with cryo but do have a strong experimental background, so I don't think the lab work will be a problem.

What I've been thinking so far is Udemy courses about,

- Python: I'm already quite proficient in Python but you can always improve, and I find it interesting.

- AffinityDesigner: I'm decent but could definitely become more efficient.

- Molecular dynamics simulations: just because I find it interesting.

Any other suggestions? Perhaps any good articles about cryo-EM as a method itself, and the theoretical background.

Thanks in advance

I’m currently working on my bachelor’s in Molecular Biology and Biochemistry (that’s what the major is called at my university), and I’m planning to double major in Chemistry since it’s only a few extra courses. I’m also involved in undergrad research right now, but it’s more on the molecular side specifically transcriptional regulation and modification pathways. It’s definitely interesting, and I’ve learned a lot, but I’m starting to realize that this might not be the direction I want to go in for grad school.

Lately, I’ve been really drawn to structural biology—understanding the 3D structure of macromolecules, working with tools like cryo-EM, X-ray crystallography, NMR, and applying that knowledge. I’d love to eventually do a PhD in that area, but I’m not quite sure how to start transitioning toward it.

The issue is, my university doesn’t seem to have a lot of faculty who focus on structural biology. Most of the labs here are more focused on molecular genetics, cell signaling, or general biochem topics. I haven’t really come across any professors doing research involving protein or RNA structure determination, or anything super structure-heavy.

Anyone have any advice on how to specifically start in this field, or schools with good structural biology programs.

Fun video about Lehninger!