The vacuole occupies a large portion of plant cell volume, it is especially true to fruit tissues. Berry flesh cell vacuole serves as storage organelle for water, sugars, acids, secondary metabolites and others, which largely determining berry quality (Fontes et al., 2011a, b; Shiratake and Martinoia, 2007, Conde et al., 2006). However, the molecular basis of these compartmentation processes is still poorly understood. As in many species, the major bottle neck to study these aspects in grapevine is to obtain highly purified vacuoles with a good yield (Fontes et al., 2010). Up to date, several vacuole or tonoplast proteome researches were applied on a few plants mainly on Arabidopsis thaliana, vacuoles or tonoplast were derived from mesophyll cells (Carter et al., 2004, Endler et al., 2006, Schulze, et al., 2012) or cell culture (Jaquinod et al 2006, Shimaoka et al 2004), cauliflower buds (Schmidt et al., 2007) and sugar beet taproots (Jung et al., 2015). Though the grape berry protoplasts and intact vacuoles were successfully isolated from Cabernet Sauvignon berry suspension-cultured cells (Fontes et al., 2010), the vacuoles isolated from grape berry or different development and ripening stages of grape berry mesocarp tissues were not achieved.