We established an advanced protocol for in vitro propagation of Aloe vera via comparison of basal media, sucrose contents, growth hormone combinations, and additional supplementation with various polyamines. The maximal number and growth of shoots after 5 weeks was obtained using MS media including $30g;L^{-1}$ sucrose supplemented with $1.0mg;L^{-1}$ BA and $0.1mg;L^{-1}$ NAA. To improve shoot production, various concentrations of putrescine, spermidine, and spermine were added under optimal growth hormone conditions (MS media supplemented with $30g;L^{-1}$ sucrose, $1.0mg;L^{-1}$ BA, and $0.1mg;L^{-1}$ NAA). Maximal shoot number and growth after 5 weeks were achieved with supplementation of $50mg;L^{-1}$ spermidine. Regenerated plants were successfully acclimatized in soil with 100% efficiency. Cytogenetic inspection revealed that the regenerated plants maintained intact chromosomes identical to those of plants grown in field conditions. This protocol provides a valuable alternative for mass production of elite Aloe vera.