Improvement of automation on a High-Throughput TRPML-1 calcium 6 FLIPR Tetra assay for Identifying TRPML1 agonist

ABSTRACT For Poster

TRPML1 is a Ca2+ channel on lysosomes and a key regulator of lysosomal processes including exocytosis, membrane trafficking and autophagy. Deficits in TRPML1 channel activity are linked to various dysfunctions in intracellular protein and lipid trafficking. TRPML1 is a potentially druggable mechanism by which to modulate the TFEB pathway. The TRPML-1 assay is a calcium 6 FLIPR Tetra assay identifying compounds that activate human TRPML-1.  The assay utilizes a HEK cell line inducibly expressing the human TRPML-1 receptor. After incubation with compounds, upon target activation, a direct measurement of intracellular fluorescence due to increased calcium concentration occurs. We developed an advanced tip washing protocol with an 80% DMSO wash to overcome compound carry over challenges in the TRPML1 High-Throughput Screen. We will share results highlighting that compound carry over was resolved after washing the tips using 80% DMSO. Assay performance, data quality and accuracy improvements will also be reviewed. Tas a result of the improved results, the advanced wash method has become our standard operating method for FLIPR-based assays.