Evaluation of a high-content screening fluorescence-based assay analyzing the pharmacological modulation of lipid homeostasis in human macrophages

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Abstract
Background For understanding cholesterol and phospholipid efflux pathways there is a need for cellular fluorescence‐based high‐content screens (HCS) to investigated the cholesterol and phospholipid content in human macrophages. Methods Making use of fluorescence imaging based on HCS we have developed a tool to evaluate new agents that can act as inducers of cholesterol efflux. The fluorescence assay is based on the different staining patterns of cholesterol‐loaded (E‐LDL) and deloaded (HDL3) differentiated monocytes by the saturated, fluorescene lipid probe (1,2‐dimyristoyl‐sn‐glycero‐3‐phospho‐ethanolamine)‐tetramethyl‐rhodamin. Results Morphologic examination and statistical evaluation of the staining pattern such as gray value, threshold area, shape factor and the spot size distribution provides evidence for a significant pattern change when cholesterol enriched and cholesterol depleted differentiated monocytes were imaged. © 2006 International Society for Analytical Cytology