Object. The multipotency of neural stem cells (NSCs) can be assessed in vitro by detection of stage-specific
markers in response to a suitable differentiation signal. This test is frequently used because it is fast and affordable.
However, it is not clear how the in vitro potential for multilineage differentiation and stem cell marker expression
would reflect the ability of NSCs to engraft into the brain following transplantation. The authors undertook this study
to directly compare the in vitro potency and in vivo migration of human NSCs (hNSCs) expanded under conditions
of gradually increased concentration of fetal bovine serum (FBS) as a maturation factor.

Methods. Human NSCs isolated from fetal brain were propagated in serum free media (SF-hNSCs) and in media
containing 0.1% and 0.2% serum. At Passage 4 in tissue culture the NSCs were harvested and either differentiated in
vitro or transplanted into the lateral ventricle of chicken embryonic brain at the late stage of its development (Hamburger
and Hamilton Stage 26). The in vitro differentiation was evaluated by immunostaining with neural or glial
specific markers, and the in vivo migration was assessed using immunohistology.

Results. The authors found that SF-hNSCs successfully engrafted into the chicken embryonic brain, which correlated
with their ability to differentiate in vitro. NSCs grown at as low as 0.1% concentration of FBS failed to demonstrate
the robust in vivo migration pattern but still preserved the capability to differentiate in vitro. Furthermore,
NSCs generated in media containing a higher concentration of FBS (0.2%) lost both the in vivo engraftment and in
vitro differentiation potential.

Conclusions. The present study suggests that marker expression and in vitro differentiation assays might not
provide adequate information regarding the behavior of NSCs following their transplantation. The in vivo migration
following injection into chicken embryonic brain may provide an important assay of the potency of NSCs.