what I heard:

1, using trypsin will lose some macrophages' markers, not good for your typing, etc..

2. there are some other reagents (not enzyme) for detaching the macrophages.

Dear Juan Diego,

Why do you prefer collagen coated plate? My suggestion is about regular cell culture plate for this. Collagen is a target for M2 Macrophages than M1. And, It may be change M differentiation. I attached a manuscript about that.

I wish good luck for your studies.

Ishak

Dear Juan Diego

I agree with the previous answers. I worked in my PhD with primary macrophages and my advice to you

1- in order to grow macrophages use the normal petri dishes ( not coated) and add new media on day 3 and replace the whole media on day 5 by new media.

2- when harvesting macrophages on day 7 try to aspirate the old media and add 5ml cold media and leave for 2 minutes in the fridge and then use the scraper very gently to collect the cells in the bottom of one side of the flask. after you finish all your plates collect the cell suspension in a 50 ml tubes and centrifuge for 5 minutes in 1000 rpm( this step will wash your cells and gives you just the good cells). after that you can resuspend the cells and they will be ready for phenotyping and also for re-seeding.

I hope this helps

Good luck

Shilan

Juan Diego Sanchez your monocytes are actually have property to attach to surface so, you need not you use collagen or any other (PLL) for culture. concentration of MCSF is good (varies from 50-100 ug/ml).

I would suggest to not do enzymatic treatment for detachment, instead prefer to scrape the cells in cold PBS.