Application of a modified lactate dehydrogenase assay to evaluate the viability of cells cultured on 3D scaffolds when commonly used assays fail.

Abstract

The development of new compounds or materials for medicine suggests a study of their cytotoxicity. The effect of a material on cell viability can be evaluated by several methods based on DNA content, DNA synthesis, plasma membrane integrity, cellular enzyme activity, cellular reducing potential, and ATP level. Sometimes it is impossible to apply widely used commercially available reagents, e.g., when cells are cultured on materials, that interfere with the chemicals used or resulting in the course of enzymatic reaction. Here, we offer a method for monitoring the viability of cells proliferating on different supports in vitro. The method is based on the measurement of lactate dehydrogenase (LDH) activity in cellular lysates. After cells were lysed in 1% Nonidet P40 and supernatants were transferred into fresh tubes, LDH activity was measured in the supernatants using colorimetric method. The usefulness of the test was studied using human cervical adenocarcinoma HeLa cells and human gingival fibroblasts cultivated on different materials, including activated carbon-loaded scaffolds. The comparison with widely used AlamarBlue® assay confirms the LDH-based method as an appropriate alternative for measuring the living cell number in vitro in a quick, simple, and cost-effective manner when widespread methods for the evaluation of cell viability could not be used.