Adaptation of a super-sensitive epitope detection technique for the immunoelectron microscopy of titin filaments in vertebrate striated muscle

A super-sensitive epitope-detection technique based on gold-silver intensif ication was adapted for pre-embedding immunolabelling of titin filaments in vertebrate striated muscle, Indirect immunoelectron microscopy of titin fi laments was performed with monoclonal titin antibodies as primary antibodie s and F-ab anti-mouse IgG conjugated with 1.4 nm gold particles as secondar y antibodies. The secondary antibodies penetrated easily into the tissue ow ing to their reduced size and the very small gold particles. After the labe lling procedure, the tissue was fixed in glutaraldehyde, Since the gold par ticles were not visible by conventional transmission electron microscopy, t hey were intensified with a silver developing system. Although the particle size varied nonlinearly with the developing time, very fine grain size was achievable. The technique provided super-sensitive detection with excellen t contrast and demonstrated epitopes with both strong and weal; affinities.