Immunohistochemical demonstration of alpha-1-antitrypsin in liver tissue. A methodological investigation

The influence of the tissue preparation on the immunohistochemical demonstration of alpha-1-antitrypsin (AAT) in liver tissue was evaluated using autopsy and biopsy material with and without AAT globules. On comparing frozen sections of unfixed material with paraffin sections of formalin fixed material a slightly better preservation of the immunoreactivity of AAT was observed in frozen sections. On comparing different fixatives, fixation in 10% neutral buffered formalin. Lillie's AAF or 96% ethanol/1% acetic acid caused a clearly better preservation of demonstrable AAT than fixation in Clarke's or Bouin's fixatives. The fixation time had only minor influence when using fixation times within a week. Extremely long fixation for several months caused, however, a clear reduction in demonstrable AAT. Pretreatment with proteolytic enzymes of deparaffinized sections of formalin fixed tissue caused an increase in demonstrable AAT, especially in tissues fixed for extremely long periods of time. On comparing three different immunohistochemical techniques: indirect immunoperoxidase, peroxidase/anti-peroxidase (PAP), and indirect immunofluorescnece, no convicting difference in sensitivity was observed between the three techniques. It is concluded that for the immunohistochemical demonstration of AAT in liver tissue the employment of indirect immunoperoxidase staining on formalin fixed, paraffin embedded material is recommendable as it combines a sensitive staining technique with a satisfactory preservation of immunoreactivity and tissue morphology.