Biomedical Engineering Reference
In-Depth Information
is advantageous for scaling up. Here, recent technologies in the
scaling up of EB-based differentiation culture of hESCs/iPSCs are
introduced. There are two major conventional methods for the
preparation of EBs. One is a hanging-drop method, and the other
is a suspension culture method. In the case of the hanging-drop
method, the size of EBs can be controlled. Because EB size is one
of the important factors in ES/iPSC differentiation, the hanging-
drop method is advantageous for efficient and reproducible
differentiation into cardiomyocytes. However, the hanging-
drop method requires a labor-intensive procedure and thus is
unfavorable for large-scale culture. On the other hand, in the case of
the suspension culture method, a large number of EBs can be easily
prepared. However, in a usual static suspension culture method, EB
size cannot be controlled at all, which results in lower efficiency and
reproducibility of cardiac differentiation than that by the hanging-
drop method. Therefore, the development of a simple and scalable
method for preparing size-controlled EBs has been undertaken. One
of the simple solutions for this issue is a stirred suspension culture
system. There are some reports showing that the uniformity of EB
size is enhanced to some extent by a stir in a suspension culture
system [25, 26]. However, a stir allows just a loose control of EB size,
and thus additional technologies are required to allow more precise
control of EB size. There are several attempts to address this subject
by utilizing microfabrication technology [27-30]. Here, recent two
reports are introduced. Utilizing the technique of microcontact
printing, Bauwens et al
printed a dot pattern of Matrigel on culture
surfaces and cultured hESCs on the surfaces as circular colonies
of a fixed diameter [29]. Subsequently, the circular colonies were
stripped from the culture surfaces, and then uniform-sized EBs
were successfully prepared in a scalable manner. Ungrin et al.
developed microwell substrates for the scalable preparation of size-
controlled EBs [30]. The microwells on the substrates have a square-
pyramidal shape, which is advantageous to the efficient assembly
of cells. They demonstrated that hESCs seeded onto this microwell
substrate formed finely size-controlled EBs inside the microwells.
The microwell substrates have already been commercially available
as AggreWell plates (Stemcell Technologies Inc).
As a method for the subsequent large-scale culture of prepared
EBs for cardiac differentiation, a stirred suspension culture system
still has the advantage of scalability and online control of culture
.
