A monoclonal antibody, MF20, which has been shown previously to bind the myosin heavy chain of vertebrate striated muscle, has been proven to bind the light meromyosin (LMM) fragment by solid phase radioimmune assay with alpha-chymotryptic digests of purified myosin. Epitope mapping by electron microscopy of rotary-shadowed, myosin-antibody complexes has localized the antibody binding site to LMM at a point approximately 92 nm from the C-terminus of the myosin heavy chain. Since this epitope in native thick filaments is accessible to monoclonal antibodies, we used this antibody as a high affinity ligand to analyze the packing of LMM along the backbone of the thick filament. By immunofluorescence microscopy, MF20 was shown to bind along the entire A-band of chicken pectoralis myofibrils, although the epitope accessibility was greater near the ends than at the center of the A-bands. Thin-section, transmission electron microscopy of myofibrils decorated with MF20 revealed 50 regularly spaced, cross-striations in each half A-band, with a repeat distance of approximately 13 nm. These were numbered consecutively, 1-50, from the A-band to the last stripe, approximately 68 nm from the filament tips. These same striations could be visualized by negative staining of native thick filaments labeled with MF20. All 50 striations were of a consecutive, uninterrupted repeat which approximated the 14-15-nm axial translation of cross-bridges. Each half M-region contained five MF20 striations (approximately 13 nm apart) with a distance between stripes 1 and 1', on each half of the bare zone, of approximately 18 nm. This is compatible with a packing model with full, antiparallel overlap of the myosin rods in the bare zone region. Differences in the spacings measured with negatively stained myofilaments and thin-sectioned myofibrils have been shown to arise from specimen shrinkage in the fixed and embedded preparations. These observations provide strong support for Huxley's original proposal for myosin packing in thick filaments of vertebrate muscle (Huxley, H. E., 1963, J. Mol. Biol., 7:281-308) and, for the first time, directly demonstrate that the 14-15-nm axial translation of LMM in the thick filament backbone corresponds to the cross-bridge repeat detected with x-ray diffraction of living muscle.

Most of the sounds of human speech are produced by vibration of the vocal folds, yet the biomechanics and control of these vibrations are poorly understood. In this study the muscle within the vocal fold, the thyroarytenoid muscle (TA), was examined for the presence and distribution of slow tonic muscle fibers (STF), a rare muscle fiber type with unique contraction properties. Nine human TAs were frozen and serially sectioned in the frontal plane. The presence and distribution pattern of STF in each TA were examined by immunofluorescence microscopy using the monoclonal antibodies (mAb) ALD-19 and ALD-58 which react with the slow tonic myosin heavy chain (MyHC) isoform. In addition, TA muscle samples from adjacent frozen sections were also examined for slow tonic MyHC isoform by electrophoretic immunoblotting. STF were detected in all nine TAs and the presence of slow tonic MyHC isoform was confirmed in the immunoblots. The STF were distributed predominantly in the medial aspect of the TA, a distinct muscle compartment called the vocalis which is the vibrating part of the vocal fold. STF do not contract with a twitch like most muscle fibers, instead, their contractions are prolonged, stable, precisely controlled, and fatigue resistant. The human voice is characterized by a stable sound with a wide frequency spectrum that can be precisely modulated and the STF may contribute to this ability. At present, the evidence suggests that STF are not presented in the vocal folds of other mammals (including other primates), therefore STF may be a unique human specialization for speech.

Monoclonal antibodies (McAbs) against the myosin heavy chain (MHC) of adult chicken pectoralis muscle have been tested for reactivity with pectoralis myosin at selected stages of chick development in vivo and in vitro. Three such McAbs, MF 20 and MF 14, which bind to light meromyosin, and MF 30, which binds to myosin subfragment two (S2), were used to assay the appearance and accumulation of specific MHC epitopes with: (a) indirect, solid phase radioimmune assay (RIA), (b) immunoautoradiography, (c) immunofluorescence microscopy. McAb MF 20 bound strongly and equivalently to MHC at all stages of embryonic development in vivo. In contrast, the MF 30 epitope was barely detectable at 12 d of incubation but its concentration rose rapidly just before hatching. No detectable binding of MF 14 to pectoralis myosin could be measured during myogenesis in vivo until 1 wk after hatching. Immunofluorescence studies revealed that all three epitopes accumulate in the same myocytes of the developing pectoralis muscle. Since all three McAbs bound with high activity to native and denatured forms of myosin, it is unlikely that differential antibody reactivity can be explained by conformational changes in myosin during development in vivo. When myogenesis in vitro was monitored using the same McAbs, MF 20 bound to the MHC at all stages tested while reactivity of MF 30 and MF 14 with myosin from cultured muscle was never observed. Thus, this study demonstrates three different immunochemical states of the MHC during development in vivo of chick pectoralis muscle and the absence of later occurring immunochemical transitions in the MHC of cultured embryonic muscle.