Hippeastrum (Amaryllidaceae) is very popular worldwide, both as a pot plant and grown for cut flowers. Bulbs of hippeastrum are produced from seeds, offsets, and bulblets obtained by twin-scaling. The in vitro propagation of this genus has also been developed. Studies were done on H. hybridum ‘Double Roma’ and on the interspecific hybrid H. × chmielii Chm. clone no. 18. The effect of plant growth regulators and the plant biostimulator Goteo on efficacy of hippeastrum micropropagation (regeneration, multiplication, bulb formation, and the acclimatization) was evaluated. Also, the effect of biostimulators Goteo and Calibra and fertilizer Florovit on the first bulb yield ex vitro were researched. Supplementation of MS medium with α-naphthaleneacetic acid significantly increased the number of leaves and roots produced by bulblets, and doubled the weight of bulblets regenerated in vitro in H. × chmielii. Addition of Goteo to the medium resulted in formation of largest number of roots, heaviest bulblets, and highest bulblet mass increase during the in vitro multiplication of H. × chmielii. The positive effect of Goteo on increase in number of regenerated bulblets as well as on increase in root number and plantlet weight were observed during the bulb formation stage in both genotypes. In H. × chmielii, treatment of bulblets ex vitro with combination of biostimulator Goteo and fertilization with Florovit resulted in longer leaves and more roots when compared with other treatments. In both hippeastrum genotypes the application of Goteo together with Florovit positively affected the weight of bulblets after the first season of vegetation. Each bulb of H. × chmielii produced daughter bulbs, what was not observed in ‘Double Roma’.