Manduca begins development shortly after fertilization. It is desirable to isolate myoblasts, cells that have differentiated to become muscle cells, but have not yet differentiated into muscle. Timing is critical and knowledge of development an important guide. At the proper time, embryo cells can be obtained from eggs by homogenization, separation from the yolk and plating the isolated cells on tissue culture medium in a petri dish. The myoblasts adhere to the media on the plate better than other cell types. Myoblasts will migrate from the original cell masses and fuse. Under proper hormonal conditions, they contract spontaneously in 5 days. Growth can be promoted or suppressed using combinations of 20 hydroxyecysone and juvenile hormone analogs. Cell cultures can be preserved by freezing and stimulated to grow when thawed. Development of basic cell culture methods is an important step in the final goal of producing a biologically derived tissue that is useful to bioengieering applications.
*Isolation and Maintenance-Free Culture of Contractile Myotubes from Manduca sexta Embryos. Baryshyan AL, Woods W, Trimmer BA, Kaplan DL. 2012. PLoS ONE 7(2): e31598.