u/Calm_realistic

HI. This is gated among single live cells of human PBMC (patients). 1st image PBMCs are thawed, stained and passed on a cytometer. 2nd, same staining after 24h culture (1M/ml if RPMI 10% FBS 1% P/S). Culture is done in 12 well plates. Cells are recovered and then the bottom is scraped to recover attached cells.

I have one population after thawing, but which is clearly composed of high CD33 and high CD14 cells. After culture, I have 2 distinct populations. The high ones and the one with 1D14 very low/negative. Both populations express CD11c and HLA-DR, are negative for CD3, CD56, CD19 , CD20.

When activated with IFNg, only the CD14high CD33 up regulate PD-L1, the CD14 low cells have keep the basal level (almost no expression).

I don't know much about myeloid cells. What is the population that becomes CD14 very low? Can it be an artefact?

Thank you for all the help:

https://preview.redd.it/zqvystx5cu9h1.jpg?width=681&format=pjpg&auto=webp&s=c64c0226646c7843bc6c5e3e8d28cfb548be67a5

Hi, I would like to make some, but I need some advice

1. If I understood well, I fill. jar with 2/3 garlic, cover it with honey. Garlic should be submerged, or turned upside down one a day.

2. For how long should the burping be done? I am leaving on vacation in 2 weeks. Should I start after I come back?

3. Getting a pH measuring something (strips are apparently not perfect because the color of honey can give falser results) is a good idea?

I am right for these points?

Thank you

Hello,

I need to dissociate spheroids with MDA and HepG2 cell lines (300um in diameter, 1000 thousand cell on monday and I am recovering them on tuesday). I am going to use accutase, but what is missing in my protocol is how to pellet my spheroids after I recover them from cultured wells for washing with PBS before adding accutase.

What do you put them in? Do you just leave them to pellet or you centrifuge them? what speed ?

I am also a bit confused with accutase protocol. In one reserachgate sub, people were saying not to put cells at 37°C, i was told to put in the incubator according to the data sheet and pipet every 10 minutes to dissociate them.

Thank you