Essential 6 Medium—Differentiate PSCs into Various Lineages

Essential 6 Medium is designed for defined, feeder-free pluripotent stem cell culture under specific transition or experimental conditions. It is best suited for:

• Feeder-free culture systems: Supports human pluripotent stem cells (hPSCs) in fully defined, feeder-free conditions when used with appropriate matrices
• Transition to xeno-free workflows: Excellent for gradually adapting cells from serum- or feeder-dependent systems to more defined or xeno-free culture environments
• Differentiation initiation: Commonly used as a basal medium for directed differentiation protocols due to its simplified formulation compared to Essential 8
• Transient culture conditions: Suitable for short-term culture steps where removal of specific growth factors (e.g., FGF2, TGF-β) is required
• Customizable supplementation: Helps provide a minimal, defined base that allows researchers to add lineage-specific factors for controlled differentiation studies
• Research requiring reduced signaling input: Enables tighter experimental control by omitting key pluripotency-maintaining components present in Essential 8 Medium

Essential 6 Medium is used when pluripotency-maintaining growth factors need to be removed in a controlled, defined system. It is ideal for:

• Initiating embryoid body formation
• Directed differentiation into specific lineages
• Early-stage lineage commitment studies
• Reprogramming workflows where removal of TGFβ improves efficiency

For routine maintenance and expansion of undifferentiated PSCs, Essential 8 Medium is recommended.

Unlike other media used in PSC culture, Essential 6 Medium does not contain bFGF or TGFβ. Therefore, Essential 6 Medium can be used as a base for the differentiation of various cell types in the endodermal, mesodermal, and ectodermal lineages (Figures 1–3).

Figure 3. Motor neuron (MN) differentiation using Essential 6 Medium. Combinations of growth factors and small molecules were added sequentially to PSCs in Essential 6 Medium according to the protocol in Lippmann ES, et al. Stem Cell Reports (2015) in order to generate various types of motor neurons. (A) Expression of GFP (green) under the motor neuron-specific Hb9 promoter in the HUES3-HB9::GFP hESC line indicates differentiation into the motor neuron lineage. (B) RT-PCR analysis was performed and data were normalized at time 0 of CHIR treatment. The data reveal different Hox expression profiles indicative of regionalized motor neuron progenitors (pMN). (Data generated by Randolph Ashton, PhD and Nisha Iyer, PhD, University of Wisconsin, Madison.)