In an immunohistochemical study of 59 routinely processed tissue specimens from 48 adult testes with isolated carcinoma-in-situ (CIS) changes and of 66 specimens from adult testes without neoplasia, placental-like alkaline phosphatase (PlAP) was shown to be a reliable marker of CIS cells preceding the development of a testicular tumour. Thus, a positive reaction was encountered in all 36 biopsies treated with formaldehyde, or Bouin's or Stieve's fluid. However, only 11 of 23 specimens fixed with Cleland's fluid were immunoreactive for PlAP. None of the non-malignant components of seminiferous tubules, including the large abnormal spermatogonia, reacted with the antibody against PlAP. Besides the antibody against PlAP, monoclonal antibodies M2A and 43-9F were tested on CIS specimens fixed with the above-mentioned fixatives. In the 17 specimens fixed with Stieve's or Bouin's fluid, a positive reaction was obtained in all sections with all three antibodies tested. However, for each antibody at least two specimens gave a weak staining reaction. When all three immunostainings were performed, in each case at least one of them gave a moderate or strong reaction, thus making CIS cells easily detectable. In the samples fixed with Cleland's fluid, a negative reaction was found in one to three specimens, depending on the antibody used. However, at least one of the three antibodies gave a positive reaction if all three immunostainings were applied. In only one of the formaldehyde-fixed paraffin specimens did CIS cells react with the monoclonal antibody 43-9F, whereas M2A gave no positive reaction at all if this method of fixation was used. Thus, the sensitivity of the immunohistochemical staining procedure in the detection of CIS is dependent on the fixative used and increases when immunostainings with all three markers are performed simultaneously.