What is LDH Release Assay?
LDH (Lactate Dehydrogenase) release assay is a widely-used method to assess cell membrane integrity. When cells are damaged or undergo cell lysis, LDH, an enzyme found in the cytoplasm, is released into the extracellular space. By measuring the activity of LDH in the extracellular medium, researchers can quantify the extent of cell damage or cytotoxicity.
Cell Culture: Cells are cultured in appropriate conditions and treated with various concentrations of nanomaterials.
Treatment: After a specified incubation period, the cell culture medium is collected to measure the extracellular LDH activity.
LDH Detection: The collected medium is mixed with a reaction mixture containing NADH and pyruvate. LDH catalyzes the conversion of pyruvate to lactate, and the decrease in NADH absorbance is measured spectrophotometrically.
Data Analysis: The extent of LDH release is quantified and compared to control groups to determine the cytotoxicity of the nanomaterials.
Simplicity: The assay is straightforward and does not require specialized equipment.
High Sensitivity: The assay is sensitive enough to detect low levels of cell damage.
Quantitative: Provides quantitative data that can be statistically analyzed.
Non-Radioactive: Unlike some other cytotoxicity assays, the LDH release assay does not involve the use of radioactive substances.
Non-Specificity: The assay does not distinguish between different types of cell death, such as apoptosis and necrosis.
Cell Type Dependency: The assay's sensitivity can vary depending on the cell type used.
Interference by Nanomaterials: Certain nanomaterials may interfere with the assay reagents, leading to false results.
How to Overcome the Limitations?
To overcome these limitations, it is recommended to use the LDH release assay in conjunction with other cytotoxicity assays, such as
MTT assay or
flow cytometry. Additionally, proper controls should be included to account for any potential interference by nanomaterials.
Conclusion
The LDH release assay is a valuable tool in nanotechnology for assessing the cytotoxicity of nanomaterials. Its simplicity, sensitivity, and quantitative nature make it a preferred choice for many researchers. However, it is essential to be aware of its limitations and use complementary techniques to obtain comprehensive toxicity profiles of nanomaterials.
