In Vitro Co-culture of Bacterial and Mammalian Cells to Investigate Effects of Potential Probiotics on Intestinal Barrier Function
Human intestinal barrier function is crucial for health. Beneficial microbes, such as commensal gut bacteria and probiotics, are known to contribute to the regulation of this barrier function. Interactions between bacteria and human intestinal cells can be analyzed by co-culturing bacteria with mamm...
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Bio-protocol LLC
2025-06-01
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| Series: | Bio-Protocol |
| Online Access: | https://bio-protocol.org/en/bpdetail?id=5361&type=0 |
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| author | Ajitpal Purba Rachel Anderson Dulantha Ulluwishewa |
| author_facet | Ajitpal Purba Rachel Anderson Dulantha Ulluwishewa |
| author_sort | Ajitpal Purba |
| collection | DOAJ |
| description | Human intestinal barrier function is crucial for health. Beneficial microbes, such as commensal gut bacteria and probiotics, are known to contribute to the regulation of this barrier function. Interactions between bacteria and human intestinal cells can be analyzed by co-culturing bacteria with mammalian cells in vitro. Here, we describe a method to assess the effect of individual bacterial strains on intestinal barrier function using automated transepithelial electrical resistance (TEER) measurements. Caco-2 cells are used as a model of the intestinal epithelium, as these cells spontaneously differentiate into small intestinal epithelial-like cells characterized by tight junctions between adjacent cells. These cells are seeded on polyester filter inserts and cultured for 17 days to form a differentiated monolayer prior to the co-culture experiment. Bacteria are grown on agar, and a single colony is used to prepare a liquid culture in bacterial broth appropriate for the bacteria of interest. On the day of the co-culture experiment, the bacterial culture is resuspended in cell culture medium at the desired concentration. Inserts are transferred to cellZscope cell modules to enable automated TEER measurements, and the medium in the insert is replaced with cell culture medium containing the bacteria of interest. This method allows for intestinal tight junction barrier function to be assessed non-invasively and in real-time in response to probiotics. The use of the automated cellZscope system eliminates the need for labor-intensive manual TEER measurements, which reduces the variability in data that results from human handling and temperature changes that occur when cells are removed from the incubator. |
| format | Article |
| id | doaj-art-739210e5598a47cfaf087719d4e08ce7 |
| institution | Kabale University |
| issn | 2331-8325 |
| language | English |
| publishDate | 2025-06-01 |
| publisher | Bio-protocol LLC |
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| series | Bio-Protocol |
| spelling | doaj-art-739210e5598a47cfaf087719d4e08ce72025-08-20T03:32:23ZengBio-protocol LLCBio-Protocol2331-83252025-06-01151210.21769/BioProtoc.5361In Vitro Co-culture of Bacterial and Mammalian Cells to Investigate Effects of Potential Probiotics on Intestinal Barrier FunctionAjitpal Purba0Rachel Anderson1Dulantha Ulluwishewa2AgResearch, Joint Food Science Facility, Palmerston North, New ZealandAgResearch, Joint Food Science Facility, Palmerston North, New ZealandAgResearch, Joint Food Science Facility, Palmerston North, New ZealandHuman intestinal barrier function is crucial for health. Beneficial microbes, such as commensal gut bacteria and probiotics, are known to contribute to the regulation of this barrier function. Interactions between bacteria and human intestinal cells can be analyzed by co-culturing bacteria with mammalian cells in vitro. Here, we describe a method to assess the effect of individual bacterial strains on intestinal barrier function using automated transepithelial electrical resistance (TEER) measurements. Caco-2 cells are used as a model of the intestinal epithelium, as these cells spontaneously differentiate into small intestinal epithelial-like cells characterized by tight junctions between adjacent cells. These cells are seeded on polyester filter inserts and cultured for 17 days to form a differentiated monolayer prior to the co-culture experiment. Bacteria are grown on agar, and a single colony is used to prepare a liquid culture in bacterial broth appropriate for the bacteria of interest. On the day of the co-culture experiment, the bacterial culture is resuspended in cell culture medium at the desired concentration. Inserts are transferred to cellZscope cell modules to enable automated TEER measurements, and the medium in the insert is replaced with cell culture medium containing the bacteria of interest. This method allows for intestinal tight junction barrier function to be assessed non-invasively and in real-time in response to probiotics. The use of the automated cellZscope system eliminates the need for labor-intensive manual TEER measurements, which reduces the variability in data that results from human handling and temperature changes that occur when cells are removed from the incubator.https://bio-protocol.org/en/bpdetail?id=5361&type=0 |
| spellingShingle | Ajitpal Purba Rachel Anderson Dulantha Ulluwishewa In Vitro Co-culture of Bacterial and Mammalian Cells to Investigate Effects of Potential Probiotics on Intestinal Barrier Function Bio-Protocol |
| title | In Vitro Co-culture of Bacterial and Mammalian Cells to Investigate Effects of Potential Probiotics on Intestinal Barrier Function |
| title_full | In Vitro Co-culture of Bacterial and Mammalian Cells to Investigate Effects of Potential Probiotics on Intestinal Barrier Function |
| title_fullStr | In Vitro Co-culture of Bacterial and Mammalian Cells to Investigate Effects of Potential Probiotics on Intestinal Barrier Function |
| title_full_unstemmed | In Vitro Co-culture of Bacterial and Mammalian Cells to Investigate Effects of Potential Probiotics on Intestinal Barrier Function |
| title_short | In Vitro Co-culture of Bacterial and Mammalian Cells to Investigate Effects of Potential Probiotics on Intestinal Barrier Function |
| title_sort | in vitro co culture of bacterial and mammalian cells to investigate effects of potential probiotics on intestinal barrier function |
| url | https://bio-protocol.org/en/bpdetail?id=5361&type=0 |
| work_keys_str_mv | AT ajitpalpurba invitrococultureofbacterialandmammaliancellstoinvestigateeffectsofpotentialprobioticsonintestinalbarrierfunction AT rachelanderson invitrococultureofbacterialandmammaliancellstoinvestigateeffectsofpotentialprobioticsonintestinalbarrierfunction AT dulanthaulluwishewa invitrococultureofbacterialandmammaliancellstoinvestigateeffectsofpotentialprobioticsonintestinalbarrierfunction |