SIMULTANEOUS IN-SITU DETECTION OF MESSENGER-RNA AND APOPTOTIC CELLS BY COMBINED HYBRIDIZATION AND TUNEL

We established a new method to allow simultaneous in situ detection of mRNA expression and apoptotic DNA fragmentation in paraffin-embedded tissue sections. We used human thymic tissue to perform in sim hybridi zation with a digoxigenin-labeled CD95 (APO-1/Fas) ligand (CD95L)-spec ific probe followed by TdT-mediated biotin dUTP nick end-labeling (TUN EL) of apoptotic DNA fragments. Bound probes were visualized by an imm unogold-silver enhancement technique and fragmented DNA was detected w ith a streptavidin-peroxidase system. This double labeling technique p roduced a distinct, dark cytoplasmic staining of CD95L mRNA-expressing cells and an intense red nuclear precipitate in apoptotic cells or bo dies. This technique will be a useful tool for microtopographical anal ysis of apoptosis-related gene expression.