Droplet Microarray Based Screening Identifies Proteins for Maintaining Pluripotency of hiPSCs

Area of Science:

• Stem Cell Biology
• Biomaterials Science
• High-Throughput Screening

Background:

• Human induced pluripotent stem cells (hiPSCs) are vital for disease modeling and personalized medicine.
• Animal-derived materials used in hiPSC culture limit their medical applications.
• Defining effective, animal-free substrate coatings is crucial for advancing hiPSC technology.

Purpose of the Study:

• To develop and utilize a miniaturized droplet microarray (DMA) platform for screening protein combinations as hiPSC culture coatings.
• To identify novel protein coatings that maintain hiPSC pluripotency.
• To compare the efficiency of the DMA platform with conventional high-throughput screening (HTS) methods.

Main Methods:

• A miniaturized droplet microarray (DMA) platform was employed to screen 11 proteins in various binary and ternary combinations.
• The ability of these protein coatings to maintain hiPSC pluripotency, assessed by NANOG expression, was evaluated.
• The DMA platform's efficiency in terms of protein and cell usage compared to traditional 96-well plate HTS was quantified.

Main Results:

• Ten protein group coatings significantly enhanced NANOG expression in hiPSCs compared to Matrigel.
• Two identified coatings supported long-term hiPSC pluripotency and differentiation into three germ layers.
• The DMA platform reduced protein and cell consumption by approximately 860-fold and 25-fold, respectively, compared to 96-well plate HTS.

Conclusions:

• Novel protein coatings were identified that effectively maintain hiPSC pluripotency and support differentiation.
• The miniaturized droplet microarray (DMA) platform offers a highly efficient and cost-effective method for screening scarce or expensive materials.
• These findings are essential for advancing research and applications utilizing human induced pluripotent stem cells.