Ploidy level in grapevines varies, especially since in vitro techniques are employed in the breeding process and after plants are treated with either chemicals or radiation. Detection of ploidy level in grapevines by microscopic chromosome counting is complicated by their high number and the small size of chromosomes. Flow cytometry provides an accurate and rapid method in determining the ploidy level in plant tissue by measuring the nuclear DNA content in living cells and thus is a very useful tool in plant breeding or genetic studies. The objective of this research was to analyze the ploidy level of a selected group of muscadine vines that were different from normal diploid vines in morphology. These grapes were derived from either chemical treatment of known varieties or from controlled/open pollinations. Among the 26 grapevines investigated, 8 were found to be diploids, 11 were tetraploids, and 7 were chimeric aneuploids. Results of this study indicate that flow cytometry is a quick, reliable tool for determining ploidy levels of grapevines.