Drs Veninga, Morse and Wieringa reply:

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Drs Veninga, Morse and Wieringa reply: Sir, The comments made by Dr Sundberg are very much appreciated, and we thank him for providing copies of the references. It is our opinion, however, that the references cited by Dr Sundberg convincingly support the findings we reported on pigmented spleens seen in 150/0of our C57BL mice (Veninga et al., 1989), rather than refute them. For the sake of clarity, the references cited in our reply will be those provided by Dr Sundberg. The observed pigmentation in our study was proved to be primarily haemosiderin and not melanin because: (a) Macroscopically, the pigment colours up to twothirds of the spleen uniformly, giving the affected part a black appearance. It is not present in localized spots as has been described for melanin (Weissmann, 1967). It is of interest to note that the black part is maintained in spleens of animals infected with 106 lymphosarcoma ceIls,causing an exponentially growing splenic tumour which loosens the tissue in one week. It enabled us to obtain isolated macrophages filled with pigment from 2 such spleens. (b) Microscopically, the pigment is yellow-brown which is characteristic for haemosiderin (Frith & Ward, 1988). (c) The pigment specificaIly stains with prussian blue which proves haemosiderin and absolutely excludes melanin (Frith & Ward, 1988; Sundberg, 1989), with the exception of a small number of very dark granules which are possibly the melanin Dr Sundberg refers to. (d) Unlike melanin, the pigment is present in macrophages (Frith & Ward, 1988; Veninga et at., 1989), although some loose granules were observed outside the cells. These elements have not the characteristic elongated appearance of melanin spots (Frith & Ward, 1988; Sundberg, 1989). (e) The EM picture in our article shows pigment granules surrounded by a membrane, typical for haemosiderin. We agree that such membrane is less clear for a second set of granules in the same cell. Nevertheless, their arrangement is very similar and therefore suggestive of a coherent structure. Both granule clusters are enclosed in a macrophage, the typical cell type containing haemosiderin (Frith & Ward, 1988). We realize that our article obviously was not completely clear. As indicated, most of the mice we worked with were 3 months of age; after publication of the article we also discovered pigmented spleens in 2-month-old animals, showing that this is not strictly an age-onset condition. Although we would be the last to deny the possibility that a few melanin particles (such as the particles not staining with prussian blue) were present in the spleens of our mice, we prefer our original interpretation (Veninga et al., 1989). In conclusion, we maintain that the pigmented apex of the spleens of 15% of our young adult C57BL mice contain concentrations principally of haemosiderin, not melanin a disorder characteristic for haemosiderosis. Tjeerd Veninga Howard Morse Andre Wieringa