Development of a Spectral Flow Cytometry-based assay to analyze drug induced hepatoxicity
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Contributing OfficeCenter for Drug Evaluation and Research
Abstract
Autofluorescence is an intrinsic characteristic of many cells, including that of primary human hepatocytes (PHHs). This has long been an issue in traditional flow cytometry because autofluorescence interferes with spectral signatures, resulting in difficulties when detecting certain fluorescent antibodies at low levels. Traditional compensation methods may correct for some overlap of fluorochrome emission peaks but their use in reducing background noise caused by autofluorescence are limited and can potentially exclude valuable data points. For these reasons an effective protocol for the analysis of PHHs via flow cytometry has not yet been developed. However, with the emergence of spectral flow cytometry, autofluorescence can be minimized without the cost of decreased sensitivity. Our aim is to utilize the Cytek® Aurora Spectral Flow Cytometer to develop an effective method for analyzing PHHs. Preliminary data indicates that we can overcome the autofluorescence of PHHs, though methods for cell detachment and staining are still being finetuned. This spectral flow-based assay has the potential to detect liver toxicity with high sensitivity and using this technique has allowed us the ability to distinguish between necrotic and apoptotic PHHs.