Neural stem cells (NSCs) with the capacity of extensive self-renewal and multi
lineagedifferentiation have attracted more and more attention in research as NSCs will play an importantrole in the nerve disease treatment and nerve injury repair. The shortage of NSCs, both theirsources and their numbers, however, is the biggest challenge for their c
linic app
lication, andhence, in vitro culture and expansion of NSCs is vitally important to rea
lize their potentials.In this work, mouse-derived NSCs were cultured in three-dimensional calcium alginate beads(Ca-Alg-Bs). Gel
ling conditions, cell density, and cell harvest were determined by the explorationof formation and dissociation parameters for Ca-Alg-Bs. Additionally, the recovered and thesubsequent induced cells were identified by immunofluorescence staining of Nestin,
-tubu
lin,and GFAP. The results show that the 2-mm diameter Ca-Alg-Bs, prepared with 1.5% sodiumalginate solution and 3.5% CaCl
2 solution and with gel
ling for 10 min, is suitable for the NSCsculture. The seeding density of 0.8 × 10
5 cells·mL
-1 for the encapsulation of NSCs resulted inthe most expansion, and the NSCs almost doubled during the experiment. The average cellrecovery rate is over 88.5%, with the Ca-Alg-Bs dissolving in 55 mM sodium citrate solutionfor 10 min. The recovered cells cultured in the Ca-Alg-Bs still expressed Nestin and had thecapacity of multi
lineage differentiation into neurons and g
lial cells and, thus, remained to beNSCs. These results demonstrate that NSC expansion within Ca-Alg-Bs is feasible and providesfurther possibi
lities for NSC expansion in bioreactors of the scale of c
linical relevance.