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Alkaline phosphatase release assay to determine cytotoxicity for Listeria species
1Food Microbiology and Immunochemistry Laboratory, Department of Life Sciences, Alabama A & M University 35762, USA. aamaxb01@asnaam.aamu.edu
Abstract:
A simple cytotoxicity assay for Listeria species was developed by assaying alkaline phosphatase (AP) release from an infected hybrid B lymphocyte (Ped-2E9) line. Eight of eight L. monocytogenes and six of 11 L. ivanovii strains induced significantly high AP release from Ped-2E9 cells compared to five other L. ivanovii strains and other Listeria spp. In contrast, all L. monocytogenes and L. ivanovii test strains showed high release of lactate dehydrogenase (LDH) activity from Ped-2E9 cells. The molecular mass of AP was estimated to be about 128-165 kDa, suggesting severe membrane damage in Ped-2E9 cells due to Listeria infection. The data presented here indicate that AP assay could be used over LDH assay to detect Listeria-induced cell cytotoxicity.
Insights
A new alkaline phosphatase (AP) assay effectively detects cytotoxicity from Listeria infections. This method offers a more sensitive alternative to the lactate dehydrogenase (LDH) assay for identifying Listeria-induced cell damage.
Area of Science:
- Microbiology
- Cell Biology
- Biochemistry
Background:
- Listeria species are significant human pathogens.
- Assessing Listeria-induced cell damage is crucial for diagnostics.
- Existing cytotoxicity assays may have limitations.
Purpose of the Study:
- To develop and validate a simple cytotoxicity assay for Listeria species.
- To compare the efficacy of alkaline phosphatase (AP) release assay with lactate dehydrogenase (LDH) assay for detecting Listeria-induced cell damage.
Main Methods:
- Utilized a hybrid B lymphocyte (Ped-2E9) cell line.
- Infected cells with various Listeria species, including L. monocytogenes and L. ivanovii.
- Measured the release of alkaline phosphatase (AP) and lactate dehydrogenase (LDH) from infected cells.
Main Results:
- Eight of eight L. monocytogenes and six of 11 L. ivanovii strains induced significantly high AP release.
- All tested L. monocytogenes and L. ivanovii strains showed high LDH release.
- AP release correlated with severe membrane damage, indicated by its molecular mass (128-165 kDa).
Conclusions:
- The AP assay is a sensitive method for detecting cytotoxicity induced by Listeria species.
- AP assay can be a superior alternative to LDH assay for Listeria-induced cell damage detection.
- This assay provides a valuable tool for microbiological and toxicological studies.