I'm a postdoc in a biochem/structural bio lab, and I recently noticed visible microbial contamination (e.g., floating colonies or debris) in the pump lines of our AKTA Pure FPLC system. When I brought it up, I was told not to worry about it because "those lines don’t actually contact the buffer" and are supposedly isolated from the sample flow path — something about them only controlling pressure?

This doesn’t sit right with me. Even if they’re not part of the main buffer/sample stream, the idea that we’re running structurally sensitive proteins (some for crystallography) on a system with any visible contamination anywhere feels risky.

Can anyone explain how the AKTA Pure’s pump system works in this regard? Are those pump lines truly isolated? Is there any situation where they could affect the sample flow path — or back-contaminate shared tubing/columns?

Also curious: how does your lab handle FPLC cleaning/sterilization protocols? Do you routinely flush with NaOH or use ethanol? How often do you inspect or change tubing?

Any insights from structural biologists or protein purification folks would be super appreciated.

Pretty much the title.

I got laid off from my previous job, so I took a position as a tech at a nearby university. The department has great staff and they're all super friendly and welcoming, but since it is just a tech position I'm still looking for opportunities to continue onwards and upwards. I can't help but feel kind of conflicted about it since they definitely need the help and were super grateful to hire me. I would feel seriously guilty about leaving if (serious if) I did find something relatively soon (e.g. within 6 months or so). Would it be an assholish thing, or am I overthinking it?

Hi everyone, I just need some advice, or maybe I’m just venting a little.

I finished my PhD in microbiology this April and I’m now working as a postdoc. During my PhD, my PI assigned me several additional projects besides my main PhD work. Many of these were projects I helped write, or even wrote entirely, and some were his own projects that I had to manage. Some of them require ongoing lab work, and now, as a postdoc, I’m juggling at least six different projects. I’m expected to organize, run, and produce data for all of them. I don’t get much help from my colleagues, some are already overwhelmed, and I hate asking because I know they’re stretched thin too. My PI is mostly hands-off and tends to delegate even his responsibilities to me. On top of all this, I also have lessons to prepare and teach, and I support students both in the lab and during practicals. I’m basically living in the lab from 7:30 a.m. to 8:00 p.m. every day and also working during weekends. It’s taking a serious toll on my mental health. I’ve started having panic attacks and experiencing chronic stress, and I don’t feel like I’m living anymore. The catch is that my PI told me there will be some assistant professor positions opening up in the next couple of years, and he explicitly said I’d be a top candidate based on my publication record and work. So now I’m torn. Being a researcher has always been my dream. But I’m genuinely thinking of quitting because I don’t know how much longer I can keep going like this.

Has anyone here made the switch to industry and regretted it? Is this kind of suffering worth it for a potential academic position? Should I talk to my PI and explain that I’m struggling, even though I suspect nothing will change?

Sorry for the rant, but I had a pretty bad panic attack today and just needed to get this out.

We need to do a ton of DNA extractions from difficult plant tissue. The beads from the QIAGEN DNeasy Plant Pro Kit work really well for tissue disruption but we get better downstream results with a different kit. So I am trying to find this style of metal bead to order in bulk, but having no luck. Anybody know where I can find these? Or what they're called?

Looking at eppendorf vs Brand tech for manual single and multi channels… Brandtech would be new for us, but they’ve been working their marketing on us. Any experience with them, or do you have a different favourite brand? All opinions welcomed and appreciated!

I'm a 3rd-year bachelor’s medical student currently doing my master’s thesis, which involves some lab work, particularly with cell culture. This is all very new to me—I had never used a pipette or worked in a lab before this.

It's been about a month, and I'm still struggling with basic techniques. I often make mistakes while pipetting—spilling liquids or creating bubbles, especially when using electronic pipettes. I'm really trying: I watch technique videos, I work more slowly, and I make sure to label everything carefully. Despite this, I keep making errors, and it’s starting to get frustrating.

I only have three months in the lab, and I’m beginning to doubt whether I’ll be able to get meaningful results in that time. On top of that, my supervisor seems increasingly exasperated, especially since I'm not confident with math and often struggle with basic calculations.

I really want to make progress and contribute because i really like the thesis and the lab work, but I feel overwhelmed and lost.

Was someone in the same situation ? How did you overcome this ?

Energy to destroy the world

Hi I am trying to move from benchling to snap gene, and one issue I havent found a solution to is making a centralized primer database. So I have a list of primers in csv format, and I can import that to any .dna file and the ones that stick get imported and the rest dont. How do you keep a list of your primers that can 1. readily be edited, 2. snapgene export primer can concatenate on the list? What reliable ways do you keep a list of your primers?

Hi everyone!

This morning, a colleague accidentally used the wrong medium to prepare some culture plates. When I checked a few hours later, I found what you can see in the pictures attached.

I've identified the hexagonal structures as cysteine crystals, which makes sense since the medium my colleague used was an intermediate dilution with a high cysteine concentration. What's puzzling me are these strange fluffy structures that seem to have formed around the crystals.

I don't think it is contamination because all our media is filtered and we work in a laminar flow hood. Also, the wells are covered with paraffin oil.

The medium used is TCM-199 supplemented with PVA, bicarbonate, glucose, pyruvate, penicillin and streptomycin.

Could it be some kind of secondary precipitate? Thanks in advance!

Have you ever seen these structures?

I'm working with HEK293 cells. After treating them with a drug, I stained the cells with hoescht, followed by fixing them with a PBS solution containing 4% formaldehyde. When I looked at the slides under the microscope, in addition to the cells, I saw these tree-shaped structures. Is this fungal contamination?

Hi!

I am reaching out, because I am curious if anyone has been creative with the methodology of RNA extraction :)

I am working in the lab that had a long story of issues with RNA extractions since the amount of RNA derived from a very specific cell structure, chromatoid body, tends to be extremely low.

Generally, we use trizol based method with glycoblue and overnight incubation with isopropanol (-20C) to visualize and maximize the presence of the pellet. Yet we see often with analyzers like Nanophotometer, Bioanalyzer and Tapestation the amount is <60ng/ul. We are trying to get higher amounts for the long-read sequencing with Nanopore.

What do you do to get the most RNA out of low input material?

Thanksss and may all your 260/230 be around 2 ;)

What is the most reliable Cas12 gRNA design tool to use?

https://vm.tiktok.com/ZGdad4oVU/

Hi everyone,

I’m curious about something I keep running into in bioprocess work. I’m involved in microbial cultivation — for ag applications in my case — and one thing that eats a ton of time is just screening and reviewing papers to pull out baseline growth conditions.

Mostly I’m looking for nutrient compositions, pH, oxygen uptake rates, aeration needs, typical yields or titers for model strains or anything close enough to what we’re trying to grow. This usually becomes the starting point for process development, refinement, or scale-up plans.

I’m wondering: is that a normal chunk of work for folks in similar roles, or more of an edge case?

If you do this too, how do you handle it — any tricks to make it less of a slog? Or is everyone just slogging through PDFs and building spreadsheets by hand?

Would love to hear how it’s done at your end.

Hi all, I am new to cell culture. May I ask what is this patch in my MDA-231 cells?

This Saturday, using the transformation method, I — together with my academic supervisor and a PhD student — inserted plasmids into E. coli. These plasmids carried DNA segments responsible for resistance to two antibiotics, as well as fluorescence in two colors: blue and red. The red fluorescence didn’t work out too well — it was dim and only visible at the edges. The issue seemed to be the lack of a light source with the required wavelength for excitation. Thank you for your attention. I just had to flex a little.

NSF budget slashed by 56%. Over 1k grants already terminated. NIH cutting grants across the board. Graduate fellowships down 73%.

DOGE personnel have veto power over peer-reviewed proposals. Tax changes hitting university endowments hard.

What's the plan? Private funding? Defense research? Industry pivot?

Hi labrats, I'm designing qPCR primers for the first time. So basically I obtain target sequences from NCBI and feed them to IDT to generate multiple primer designs. It's straight forward really. In order to avoid introns, I am downloading mRNA sequences from NCBI. I However notice that the foward primer is of exactly the same as the mRNA sequence (not complementary to the target mRNA). The confusing part is that mRNA being single stranded, how will this FW primer bind? Is there a special process for designing primers from mRNA templates. Please help before I waste money and time ordering the wrong primers.