Reprinted from: Publication of this reprint collection is supported by paid advertising SLAS Technology 27 (2022) 44–53 Contents lists available at ScienceDirect SLAS Technology journal homepage: www.elsevier.com/locate/slast Full Length Article Simple assessment of viability in 2D and 3D cell microarrays using single step digital imaging Anna A. Popova a,∗ , Markus Reischl b , Daniel Kazenmaier a , Haijun Cui a , Timo Amberger a , Pavel A. Levkin a,c,∗ a Karlsruhe Institute of Technology, Institute of Biological and Chemical Systems – Functional Molecular Systems (IBCS-FMS), Hermann-von-Helmholtz-Platz 1, 76344 Eggenstein-Leopoldshafen, Germany bKarlsruhe Institute of Technology (KIT), Institute for Automation and Applied Informatics, Hermann-von-Helmholtz-Platz 1, 76344 Eggenstein-Leopoldshafen, Germany c Karlsruhe Institute of Technology (KIT), Institute of Organic Chemistry, Fritz-Haber Weg 6, 76131 Karlsruhe, Germany a r t i c l e i n f o Key words: Droplet microarray Colorimetry Hydrophobicity Viability assays Digital imaging a b s t r a c t Simple and rapid imaging and analysis of 2D and 3D cell culture compatible with miniaturized arrays of nanoliter droplets are essential for high-throughput screening and personalized medicine applications. In this study, we have developed a simple one-step, cost-effective and sensitive colorimetric method for the analysis of cell viability in 2D and 3D cell cultures on a nanoliter droplet microarray. The method utilizes a flatbed document scanner that detects a color change in response to cell metabolism in nanoliter droplets with high sensitivity in a single step without the need for expensive specialized equipment. This new nanoliter-based method is faster and more sensitive than equivalent methods using multi-well plate assays. The method detects quantifiable signal from as few as 10 cells and requires only 5 min. This is 2.5 to 10-fold more sensitive and 12 times faster than the same assay in multi-well plates. The method is simple, affordable, fast and sensitive. It can be used for various applications including high-throughput cell-based and biochemical screenings. Introduction Cell-based high-throughput screening is an essential approach to fundamental scientific research, drug development and personalized medicine. As a state-of-the-art technique, the majority of screenings is performed in multi-well plates in microliter volumes. To accelerate advances in research and drug discovery, it is important to enable screening of large chemical libraries in cost-effective way. In addition, reducing the amount of cell material required for the experiment might be crucial for some applications, for example for screenings of rare or hard to expand cells, like primary or patient-derived cells. Therefore, it is absolutely necessary to miniaturize screening platforms frommicroliter to nanoliter volumes and beyond. In recent years, a number of miniaturized platforms have emerged as alternatives to multi-well plates for use in screening applications. Such platforms are usually not compatible with plate readers or other equipment designed for multi-well plates and alternative analytical techniques are required to facilitate the measurement of different parameters in small culture reservoirs. The platforms based on droplet microfluidic principles, for example, are not compatible with standard spectrophotometers and customized read-out equipment based on absorbance spectroscopy has to be developed for such platforms [1–4]. ∗ Corresponding author E-mail addresses: anna.popova@kit.edu (A.A. Popova), levkin@kit.edu (P.A. Levkin). Digital imaging for estimation of color changes in culture vessels is a cost-effective alternative to complex and expensive spectroscopy [5–7]. Colorimetry is widely used in microfluidic paper-based analytical devices to detect color changes in analytes from different sources, such as food [8,9], environment [8–10], and patient material [11,12]. The change of color is analyzed either by visual detection, or by using digital cameras or scanners [13]. Obtained digital images can be evaluated using computer vision-based analytical procedures and different color spaces (e.g., RGB or grayscale) [13,14]. Colorimetric assays based on changes in the absorbance of a culture media are commonly used in a variety of multi-well cell-based assays, including cell viability and proliferation assays [15,16]. Cell viability is routinely used to determine the cytotoxic effects of tested compounds or materials. Viability assays are based on estimation of cell metabolic activity by measuring biochemical markers such as the presence of the reducing agents NADH and NADPH. These assays involve incubation of viable cells with a specific agent, leading to the a change in absorbance that is proportional to the number of metabolically active cells. The most commonly used colorimetric viability reagents are tetrazolium reduction assays, e.g., WST-8, and the oxidation–reduction indicator, resazurin [15,16]. https://doi.org/10.1016/j.slast.2021.10.017 2472-6303/© 2021 The Authors. Published by Elsevier Inc. on behalf of Society for Laboratory Automation and Screening. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/)
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