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Combined Fluorimetric Caspase-3/7 Assay and Bradford Protein Determination for Assessment of Polycation-Mediated Cytotoxicity

Lookup NU author(s): Professor Moein MoghimiORCiD


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Cationic polyplexes and lipoplexes are widely used as artificial systems for nucleic acid delivery into the cells, but they can also induce cell death. Mechanistic understanding of cell toxicity and biological side effects of these cationic entities is essential for optimization strategies and design of safe and efficient nucleic acid delivery systems. Numerous methods are presently available to detect and delineate cytotoxicity and cell death-mediated signals in cell cultures. Activation of caspases is part of the classical apoptosis program and increased caspase activity is therefore a well-established hallmark of programmed cell death. Additional methods to monitor cell-death related signals must, however, also be carried out to fully define the type of cell toxicity in play. These may include methods that detect plasma membrane damage, loss of mitochondrial membrane potential, phosphatidylserine exposure, and cell morphological changes (e.g., membrane blebbing, nuclear changes, cytoplasmic swelling, cell rounding). Here we describe a 96-well format protocol for detection of caspase-3/7 activity in cell lysates, based on a fluorescent caspase-3 assay, combined with a method to simultaneously determine relative protein contents in the individual wells.

Publication metadata

Author(s): Larsen AK, Hall A, Lundsgart H, Moghimi SM

Publication type: Book Chapter

Publication status: Published

Book Title: Nanotechnology for Nucleic Acid Delivery: Methods and Protocols

Year: 2019

Volume: 1943

Pages: 301-311

Online publication date: 06/03/2019

Acceptance date: 02/04/2018

Series Title: Methods in Molecular Biology

Publisher: Humana Press

Place Published: New York


DOI: 10.1007/978-1-4939-9092-4_19

PubMed id: 30838624

Library holdings: Search Newcastle University Library for this item

ISBN: 9781493990917