Chapter 15
Cell Monolayer Translocation Assay
Elsio A. Wunder Jr.
Abstract
An essential property associated with leptospiral virulence is the pathogen’s ability to translocate across host
cells, enabling Leptospira to evade the host immune response, disseminate, and establish infection. Cell
monolayer translocation assay allows for the quantification of Leptospira strain’s competence to cross cell
barriers while measuring the integrity of the polarized eukaryotic cell monolayer during this process.
Key words Leptospira, Polarized monolayers, Transwell
® , Translocation, Eukaryotic cells, Electrical
resistance, Leptospiral pathogenesis
1 Introduction
One of the most important features of leptospiral pathogenesis is
the aptitude of Leptospira to expeditiously penetrate mammalian
cell barriers and disseminate to multiple organs throughout the
host [1, 2]. Previous studies have emphasized the ability of pathogenic Leptospira to translocate across polarized mammalian cell
monolayers [2, 3]. Together with adhesion to extracellular surfaces
(see Chapter 16), rapid translocation of host cell barriers is used by
pathogenic Leptospira as means of evading the bodily immune
response and establishing infection, essential properties associated
with bacterial virulence [4, 5].
Polarized cell monolayer translocation assay with transepithelial/transendothelial electrical resistance (TEER) measurement was
first developed to study endothelial macromolecular transport
[6]. Since then, this method has been widely used to assess the
ability of a diverse range of pathogens to translocate across a cell
barrier [7–10], including Leptospira [11, 12] and other spirochetes
[13, 14]. This in vitro assay to assess bacterial translocation is based
on the culture of eukaryotic cells on a permeable membrane
contained within a plastic insert and placed on a plastic well, establishing an upper and lower compartment (Fig. 1). Culture medium
is added to each compartment, promoting cell growth and
Nobuo Koizumi and Mathieu Picardeau (eds.), Leptospira spp.: Methods and Protocols, Methods in Molecular Biology, vol. 2134,
https://doi.org/10.1007/978-1-0716-0459-5_15, © Springer Science+Business Media, LLC, part of Springer Nature 2020
161
Cell Monolayer Translocation Assay
Elsio A. Wunder Jr.
Abstract
An essential property associated with leptospiral virulence is the pathogen’s ability to translocate across host
cells, enabling Leptospira to evade the host immune response, disseminate, and establish infection. Cell
monolayer translocation assay allows for the quantification of Leptospira strain’s competence to cross cell
barriers while measuring the integrity of the polarized eukaryotic cell monolayer during this process.
Key words Leptospira, Polarized monolayers, Transwell
® , Translocation, Eukaryotic cells, Electrical
resistance, Leptospiral pathogenesis
1 Introduction
One of the most important features of leptospiral pathogenesis is
the aptitude of Leptospira to expeditiously penetrate mammalian
cell barriers and disseminate to multiple organs throughout the
host [1, 2]. Previous studies have emphasized the ability of pathogenic Leptospira to translocate across polarized mammalian cell
monolayers [2, 3]. Together with adhesion to extracellular surfaces
(see Chapter 16), rapid translocation of host cell barriers is used by
pathogenic Leptospira as means of evading the bodily immune
response and establishing infection, essential properties associated
with bacterial virulence [4, 5].
Polarized cell monolayer translocation assay with transepithelial/transendothelial electrical resistance (TEER) measurement was
first developed to study endothelial macromolecular transport
[6]. Since then, this method has been widely used to assess the
ability of a diverse range of pathogens to translocate across a cell
barrier [7–10], including Leptospira [11, 12] and other spirochetes
[13, 14]. This in vitro assay to assess bacterial translocation is based
on the culture of eukaryotic cells on a permeable membrane
contained within a plastic insert and placed on a plastic well, establishing an upper and lower compartment (Fig. 1). Culture medium
is added to each compartment, promoting cell growth and
Nobuo Koizumi and Mathieu Picardeau (eds.), Leptospira spp.: Methods and Protocols, Methods in Molecular Biology, vol. 2134,
https://doi.org/10.1007/978-1-0716-0459-5_15, © Springer Science+Business Media, LLC, part of Springer Nature 2020
161