Single-Molecule Array (Simoa) Technology as a Highly Sensitive Screening Tool for Protein–Protein Interactions

Transcription factor (TF) interactions are generally considered to be weak and transient, which enables the dynamic assembly and disassembly of transcription complexes. While advantageous for modulating cellular physiology, requiring rapid adaptation, these characteristics make it challenging to identify and study TF interactions using conventional methodologies. To overcome this issue, we have adapted an ultrasensitive single-molecule array (Simoa) assay, typically used for biomarker detection, to screen for and identify TF-TF interactions. Leveraging Simoa-based digital ELISA, we have screened the human transcription factor library for interaction partners of ARX (aristaless-related homeobox), a transcription factor associated with a wide spectrum of neurodevelopmental disorders. Our successful application of Simoa technology to a high-throughput screening for protein interaction partners opens up new opportunities to study transcription factor-transcription factor interactions, especially those that have been elusive to detect.