Hello everyone,

I am trying to stain macrophages in skin wound healing tissue, and it's been very tricky to establish a proper protocol. I tried markers F4/80 and MOMA-2, and Cy3 as fluorophore (secondary AB). I am using murine skin, freshly (unfixed) frozen, in OCT, and cryosectioning at 7 micrometer, mounting on superfrost plus slides, keeping at -80 until staining.

I've encountered always certain amount of background in my slides. I have tried cold acetone for 10 min and formaldehyde 3.7% (1:10 FA 37% in PBS) at RT for 1h. FA gave me better results, but still not optimal.

Would you have any suggestion to improve the results/protocol?

Thank you in advance!!

Marcela Mafra