1. We receive the specimen (punch biopsy of skin)fresh in a saline
moistened gauze sponge.

2. We place the specimen in a vial of fresh, cold (4C) 1MNaCl and leave
it in this solution for 48 hours.

3.  We then examine the specimen with a dissecting microscope and using
a very delicate pair of fine tooth forceps, gently lift one edge of the
specimen to assure that a portion of the epidermis is completely
detached on one side of the specimen.  Do not completely detach the
whole epidermis.

4.  Mount the specimen in OCT compound and freeze like a routine direct
immunoflourescence biopsy specimen.

5. Cut cryostat sections as usual for direct immuno and process from
that point on as a routine direct immuno specimen.

6.  An additional cryostat section can be cut and sent to
immunohistochemistry for staining with Type 4 collagen to assure that
the lamina densa is in the floor of the split skin substrate.  We do
this routinely but I have never seen a problem with the plane of
separation.

In our experience this method has worked very well when attempting to
distinguish bullous pemphigoid from epidemolysis bullosa acquista.

Bill Tyler
Department of Pathology
Geisinger Medical Center
Danville, PA
wtyler@psghs.edu