An automated method for the quantification of immunostained human Langerhans cells

Allergic contact dermatitis is a frequent and increasing health problem. Fo r ethical reasons, the current animal tests used to screen for contact sens itizers should be replaced by in vitro alternatives. Contact sensitizers ha ve been shown to accelerate Langerhans cell (LC) migration from human organ otypic skin explant cultures (hOSECs) more rapidly than non-sensitizers and it has been proposed that the hOSEC model could be used to screen for sens itizers. However, chemically induced decreases in epidermal LC numbers need to be accurately quantified if the alterations in epidermal LC numbers are to form the basis of an alternative system for screening contact sensitize rs in vitro. As manual counting of LCs is labour intensive and subject to i ntra- and inter-personal variation we developed an image analysis routine, using the Leica QWin image analysis software, to quantify LCs in situ using immunohistochemically stained skin sections. LCs can be identified using a ntibodies against the membrane molecule CD1a or the Lag antibody, which rec ognises cytoplasmic Birbeck granules. Quantification of epidermal LC number using the image analysis software had a much lower inter-person variation than when the same specimens were counted manually, using both the anti-lag and CD1a antibodies. The software-aided quantification of epidermal LCs pr ovides an accurate method for measuring chemically-induced changes in LC nu mbers. (C) 2001 Elsevier Science B.V. All rights reserved.