Cost-Effective Production of Biologically Active Leukemia Inhibitory Factor for Mouse Embryonic Stem Cell Culture

Area of Science:

• Stem Cell Biology
• Biochemistry
• Molecular Biology

Background:

• Leukemia inhibitory factor (LIF) is crucial for maintaining mouse embryonic stem cell (mESC) pluripotency and self-renewal.
• High costs of commercial LIF limit research accessibility and progress in stem cell applications.
• Developing affordable, functional LIF alternatives is essential for advancing stem cell research.

Purpose of the Study:

• To establish an efficient and cost-effective method for producing biologically active recombinant mouse LIF.
• To demonstrate the functionality and efficacy of homemade LIF (HM-LIF) in maintaining mESC pluripotency.
• To provide a reliable, low-cost alternative to commercial LIF for mESC culture.

Main Methods:

• Recombinant expression of non-glycosylated mouse LIF in Escherichia coli using an N-terminal glutathione S-transferase (GST) tag.
• Purification of tagged LIF via glutathione-agarose affinity chromatography.
• Cleavage of the GST tag using HRV 3C protease to yield active HM-LIF.

Main Results:

• Successfully produced biologically active, non-glycosylated mouse LIF in E. coli.
• HM-LIF effectively maintained mESCs in an undifferentiated state, comparable to commercial LIF.
• mESCs cultured with HM-LIF retained pluripotency and demonstrated differentiation potential into glutamatergic neurons.

Conclusions:

• The developed protocol offers a significantly cost-effective and reliable method for producing functional LIF.
• HM-LIF is a viable alternative to expensive commercial LIF, enhancing accessibility for stem cell research.
• This approach supports continued progress in stem cell biology and therapeutic development by reducing research costs.