In forming organoids, we and others (Mariani et al., 2012; Pasca et al., 2015) do not include Matrigel as supportive matrix, but there are reports suggesting the protective effects of Matrigel in organoid culture (Kadoshima et al., 2013; Qian et al., 2016). We found that Marigel embedding does not improve the formation of hMGEOs and hCOs in our system. However, we do not exclude potential benefit of adding dissolved Matrigel. We found adding FGF2 or EGF is not necessary in our culture system, especially given SHH signaling promotes progenitor proliferation (Lien et al., 2006). Nevertheless, we did notice supplementing FGF2 (20 ng/ml) could enhance proliferation of progenitors and consequently the size of organoids (data not shown). Adding FGF2 could be a potential option for cell lines showing lower proliferation rate without exogenous growth factors.
