Evaluating Cytotoxicity and Membrane Integrity in Lung Slices via Lactate Dehydrogenase Assay

Abstract

Source: Neuhaus, V., et al. Assessment of the Cytotoxic and Immunomodulatory Effects of Substances in Human Precision-cut Lung Slices. J. Vis. Exp. (2018).

This video demonstrates the lactate dehydrogenase, LDH assay to measure the cytotoxic effect of a test substance and its effect on the membrane integrity in human precision-cut lung slices. The assay provides a quantitative measure of cell death or damage in the sample.

Protocol

All procedures involving sample collection have been performed in accordance with the institute's IRB guidelines

1. General Preparation of Human PCLS and Subsequent Exposure to Chemicals

NOTE: Two persons are required to fill a lung. An up-to-date vaccination record for hepatitis A and B is recommended. Patients are routinely screened for HCV and HIV prior to lung transplantation. If an active infection with Mycobacterium tuberculosis is diagnosed or suspected, the lung should be rejected. Nevertheless, all fresh human lung tissue and samples derived from ....

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Nunc MicroWell 96-Well	Thermo Scientific (Schwerte, Germany)	260836
Image rendering software	Bitplane AG (Zürich, Switzerland)	IMARIS 7.6
LSM Image Browser	Zeiss (Jena, Germany)
Multiwell-Reader	Tecan Group Ltd. (Männedorf, Switzerland)	Tecan infinite F200Pro	Plate Reader
Cytotoxicity Detection Kit (LDH)	Roche (Basel, Switzerland)	11644793001
Cell Proliferation Reagent WST-1	Roche (Basel, Switzerland)	11644807001
Washing buffer	Merck (Darmstadt Germany)	524653	0.05% Tween 20 in PBS
Detergent	Sigma (Saint Louis, USA)	X100-100ML	Triton X-100