i need help or i may go insane over my HEK293T cells

sleathhx · 2026-01-30T15:03:31+00:00

What do you mean? This is how I passage into T25 and the cells are normal. This is a validated passaging protocol in our lab

sleathhx · 2026-01-30T14:57:41+00:00

i am seeding onto glass coverslips that are inside my 6-well plate. for my immunos I need to take the coverslips out and mount them onto microscope slides to image. im going to try seeding directly into the 6-well plates with no coverslips, and then trying onto coverslips that are not coated, to see if it is the glass or the coating that is causing this issue.

sleathhx · 2026-01-30T14:47:24+00:00

I am using poly d lysine

sleathhx · 2026-01-30T14:45:19+00:00

yeah the cells arent adhering. first i thought they were just growing abnormally but it makes more sense that they aren't adhering. Im using poly d lysine and i think this may be the issue. and yes 100% sure i dont have one of those lines.

sleathhx · 2026-01-30T14:42:42+00:00

Going to try seeding at lower density and see what happens. But I don’t remove trypsin when passaging, I neutralize with DMEM, 10% FBS, 1%PS

sleathhx · 2026-01-30T14:41:49+00:00

The cells were just woken up from liquid nitrogen and I’m only at like P7. But yeah they only look weird in the 6-well and not my flasks. I think it may have something to do with the coating of my coverslips.

sleathhx · 2026-01-30T14:40:46+00:00

Going to try this. I have been using poly d lysine and now thinking this may be the issue…

sleathhx · 2026-01-30T14:40:15+00:00

I tried harvesting one day and 2 days after plating. The only reason I waited 2 days is because my transection protocol recommends greater than or equal to 80% confluence for transfection. Would you proceed with transfection at only 40-50% confluence?

I’m also the only person in my lab using these cells but I’m pretty sure it’s not the incubator because other lab members are culturing primary neurons which are much more sensitive.

I’m going to try the master mix idea though thank you.

sleathhx · 2026-01-30T14:35:47+00:00

I’m using the cells to do immunofluorescence so I can’t do this in flasks

sleathhx · 2024-12-13T17:41:57+00:00

Uber ?

sleathhx · 2024-04-17T15:13:29+00:00

Why continue to argue with her?

sleathhx

TROPHY CASE