How to isolate and culture murine epidermal langerhans cells - (Dec/21/2005 )

could anyone tell me how to isolate murine epidermal langerhans cells & culture them .

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I've isolated langerhans cells from human epidermis. I cut the tissue into 3mmx3mm chunks and incubated them overnight with dispase which will liberate the epithelium from underlying stroma (you can peel it off with some forceps). I then did a short trypsin digest (5 mins only) and then put the tissue pulp through a Biojector which breaks the tissue into single cells. If you strain the suspension through a 70uM cell strainer this removes any remaining lumps. To isolate the LC, I added an anti CD1a mouse antihuman antibody and then collected the CD1a +ve cells using goat anti-mouse magnetic beads (you could maybe try goat anti mouse CD1a and then use anti goat beads). If you culture the cells, the beads should fall off as the CD1a is recycled. Maybe try X-VIVO medium.
Hope that helps.

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thanks maudkin.

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