Muscle trophic factor: I. Assay of a muscle trophic factor by measurement of muscle cell nuclei

A method for assaying a skeletal muscle cell trophic factor in chicken serum is described. The number of myoblast and myotube nuclei was considered to reflect the rate of myoblast multiplication and was thus adopted as an index for the activity of the trophic factor. In a 35-mm Falcon plastic Petri dish, 2.5 x 10(40) to 10(5) myoblasts were cultured. The culture medium consisted of 85% Eagle's minimum essential medium and 15% horse serum. The assay medium was composed of 2.4 ml of this culture medium and 0.1 ml of partially purified trophic factor or appropriately diluted chicken serum. Four dishes were prepared for each sample under identical conditions. On day 4 of incubation, cells were fixed, stained and observed under a microscope, and the number of nuclei in 20 fields was counted. The mean number of nuclei obtained from the four dishes was used to denote a single point. A three-point common-zero method of slope-ratio assay was used to calculate activity. The activity of the trophic factor was expressed with reference to the effect of the control serum on myoblast multiplication.