Abstract
The presence of atrial natriuretic polypeptide (ANP) was immunohistochemically demonstrated in the formalin-fixed and paraffin-embedded tissues of 25 autopsied normal human hearts using monoclonial antibody. The ANP amounts were immunohistochemically semiquantified and compared with were divided into 5 groups according to the interval of formalin fixation or the length of time between death and fixation. Formalin-fixation intervals were one week in group 1A and 1B; 1 year in group 2; 4 to 5 years in group 3 and 10 to 12 years in group 4. The hearts of group 1A, 2, 3 and 4 were fixed within 5 hours after death. Those of group 1B were fixed 14 to 18 hours at 4°C. After fixation, the left and right atrial appendages (LAA and RAA), the left and right atrial free walls (LA and RA), the left and right ventricular free walls (LV and RV) and the ventricular septum (VS) were transmurally dissected from each heart. They were embedded in paraffin, cut into 4μm sections and immunohistochemically stained by the avidin-biotin-peroxidase complex (ABC) method using monoclonal antibody against α-human ANP. Under a light microscope, they were evaluated semiquantitatively according to the incidence of ANP-positive cells and the intensity of immunostaining. For every heart i group 1A, the tissue concentrations of ANP in the different parts were also measured separately by RIA before fixation. ANP-positive myocytes were noted in the atria of all hearts of all groups, but no in any ventricular myocytes. Both their incidence and grade in the atria were similar among groups 1A, 1B and 2. However, they were less in group 3, and least in group 4 among all groups. For all groups, they decreased in following order: LAA>RAA≒LA>RA; the inner 1/3>the middle 1/3>the outer 1/3 of the atrial walls. The order in LAA, RAA, LA and RA was consistent with the results of RIA measurements on group 1A. In conclusion, the immunoreactivity of ANP in paraffin-embedded sections is well preserved for one year formalin-fixation interval. However, the same difference of tissue distribution in ANP is seen even in more long-term formalin-fixed hearts. Thus, the immunohistochemical method is useful for retrospective studies of ANP, if the formalin-fixation interval is considered.
