Abstract
Fluorescent matrix degradation assay is a popular and widely used assay in the field of invadopodium biology (Artym et al., 2009). Matrix remodeling and degradation can be observed under both physiological and pathological conditions. Cancer cells extensively remodel and degrade the underlying matrix by employing actin-rich protrusive structures called invadosomes. Similar structures are formed by the protozoan parasite Entamoeba histolytica (E. histolytica), upon coming in contact with fibronectin, a major component of the host (extracellular matrix) ECM. Here, we describe a similar assay to measure matrix degradation by Entamoeba histolytica.
Materials and Reagents
Equipment
Procedure
Coating of glass coverslips with fluorescent matrix
Sample preparation for the assay
Preparing samples for fluorescence microscopy
Notes
Recipes
Acknowledgments
We would like to thank Prof. Alok Bhattacharya (Jawaharlal Nehru University, New Delhi, India) and Dr. Sandipan Ganguly (National Institute of Cholera and Enteric Diseases, Kolkatta, India) for providing us with the Entamoeba histolytica HM1: IMSS strain. We are grateful to Professor Stefan Linder (University of Hamburg, Hamburg, Germany) for helpful discussion and suggestions. We are also thankful to Drs. Vira V. Artym, Kenneth M. Yamada and Susette C. Mueller as the protocol for E. histolytica has been adapted from their published protocol for studying cancer cell invasion in Methods in Molecular Biology. We sincerely thank Ms. Lekha Vinod Shah, final year Dual BS-MS degree program student at IISER Bhopal, Department of Biological Sciences, who shot and edited the video for the protocol.We are thankful to the Central Instrumentation Facility at IISER Bhopal, India. This work was funded by intramural funds from IISERB and funds from Max Planck Gesellschaft (MPG), Germany and Department of Science and Technology (DST), India. ME was funded by Senior Research Fellowship provided by University Grant Commission, Government of India.
References
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