NOMO-1 cells expressing an NF-κB luciferase reporter gene facilitate a simple, rapid monocyte activation test that can detect a wide range of pyrogens.
Pyrogens, which include endotoxin and non-endotoxin pyrogens (NEPs), act on immune cells in the bloodstream, causing various effects such as fever and endotoxic shock. The limulus amebocyte lysate test, a commonly used endotoxin test in the manufacturing of pharmaceuticals and medical devices, can d...
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| Format: | Article |
| Language: | English |
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Public Library of Science (PLoS)
2025-01-01
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| Series: | PLoS ONE |
| Online Access: | https://doi.org/10.1371/journal.pone.0326408 |
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| author | Tomohisa Nanao Yuki Marutani Katsuko Sato Tomohiro Mori Takeshi Kitagawa Teruaki Oku Takahiro Nishibu |
| author_facet | Tomohisa Nanao Yuki Marutani Katsuko Sato Tomohiro Mori Takeshi Kitagawa Teruaki Oku Takahiro Nishibu |
| author_sort | Tomohisa Nanao |
| collection | DOAJ |
| description | Pyrogens, which include endotoxin and non-endotoxin pyrogens (NEPs), act on immune cells in the bloodstream, causing various effects such as fever and endotoxic shock. The limulus amebocyte lysate test, a commonly used endotoxin test in the manufacturing of pharmaceuticals and medical devices, can detect endotoxin but not NEPs. The monocyte activation test (MAT), which uses monocytes, is a testing method included in the European Pharmacopoeia (EP 11.5; 07/2024:20630) that can detect NEPs. The MAT detects the cellular response following activation of Toll-like receptors (TLRs) by pyrogens; released cytokines, such as IL-6, are often the targets of detection. This cytokine release is regulated by the transcription factor NF-κB. In this study, we investigated whether it is possible to detect pyrogens with an NF-κB reporter gene-expressing cell line, using the NOMO-1 cell line as a model monocyte-like line. This study demonstrates that the reporter gene-expressing cells can detect 0.0125 EU/mL lipopolysaccharide (LPS) after 3 hours of incubation, and a stable calibration curve for LPS quantification can be created. Moreover, these cells can detect agonists for TLR1-9 in a concentration-dependent manner. Pharmaceuticals, including blood products and antibody drugs, were used in LPS recovery tests to confirm that they do not interfere with LPS detection. This study demonstrates that NF-κB reporter cells facilitate a simpler, more concise MAT, eliminating the complexity associated with enzyme-linked immunosorbent assays. Moreover, using the NOMO-1 cell line allows for the detection of a wider range of NEPs compared with using existing reporter gene-expressing cell lines. |
| format | Article |
| id | doaj-art-4ce0f2f12f4c4af0b49df469f0c5e203 |
| institution | Kabale University |
| issn | 1932-6203 |
| language | English |
| publishDate | 2025-01-01 |
| publisher | Public Library of Science (PLoS) |
| record_format | Article |
| series | PLoS ONE |
| spelling | doaj-art-4ce0f2f12f4c4af0b49df469f0c5e2032025-08-20T03:26:35ZengPublic Library of Science (PLoS)PLoS ONE1932-62032025-01-01206e032640810.1371/journal.pone.0326408NOMO-1 cells expressing an NF-κB luciferase reporter gene facilitate a simple, rapid monocyte activation test that can detect a wide range of pyrogens.Tomohisa NanaoYuki MarutaniKatsuko SatoTomohiro MoriTakeshi KitagawaTeruaki OkuTakahiro NishibuPyrogens, which include endotoxin and non-endotoxin pyrogens (NEPs), act on immune cells in the bloodstream, causing various effects such as fever and endotoxic shock. The limulus amebocyte lysate test, a commonly used endotoxin test in the manufacturing of pharmaceuticals and medical devices, can detect endotoxin but not NEPs. The monocyte activation test (MAT), which uses monocytes, is a testing method included in the European Pharmacopoeia (EP 11.5; 07/2024:20630) that can detect NEPs. The MAT detects the cellular response following activation of Toll-like receptors (TLRs) by pyrogens; released cytokines, such as IL-6, are often the targets of detection. This cytokine release is regulated by the transcription factor NF-κB. In this study, we investigated whether it is possible to detect pyrogens with an NF-κB reporter gene-expressing cell line, using the NOMO-1 cell line as a model monocyte-like line. This study demonstrates that the reporter gene-expressing cells can detect 0.0125 EU/mL lipopolysaccharide (LPS) after 3 hours of incubation, and a stable calibration curve for LPS quantification can be created. Moreover, these cells can detect agonists for TLR1-9 in a concentration-dependent manner. Pharmaceuticals, including blood products and antibody drugs, were used in LPS recovery tests to confirm that they do not interfere with LPS detection. This study demonstrates that NF-κB reporter cells facilitate a simpler, more concise MAT, eliminating the complexity associated with enzyme-linked immunosorbent assays. Moreover, using the NOMO-1 cell line allows for the detection of a wider range of NEPs compared with using existing reporter gene-expressing cell lines.https://doi.org/10.1371/journal.pone.0326408 |
| spellingShingle | Tomohisa Nanao Yuki Marutani Katsuko Sato Tomohiro Mori Takeshi Kitagawa Teruaki Oku Takahiro Nishibu NOMO-1 cells expressing an NF-κB luciferase reporter gene facilitate a simple, rapid monocyte activation test that can detect a wide range of pyrogens. PLoS ONE |
| title | NOMO-1 cells expressing an NF-κB luciferase reporter gene facilitate a simple, rapid monocyte activation test that can detect a wide range of pyrogens. |
| title_full | NOMO-1 cells expressing an NF-κB luciferase reporter gene facilitate a simple, rapid monocyte activation test that can detect a wide range of pyrogens. |
| title_fullStr | NOMO-1 cells expressing an NF-κB luciferase reporter gene facilitate a simple, rapid monocyte activation test that can detect a wide range of pyrogens. |
| title_full_unstemmed | NOMO-1 cells expressing an NF-κB luciferase reporter gene facilitate a simple, rapid monocyte activation test that can detect a wide range of pyrogens. |
| title_short | NOMO-1 cells expressing an NF-κB luciferase reporter gene facilitate a simple, rapid monocyte activation test that can detect a wide range of pyrogens. |
| title_sort | nomo 1 cells expressing an nf κb luciferase reporter gene facilitate a simple rapid monocyte activation test that can detect a wide range of pyrogens |
| url | https://doi.org/10.1371/journal.pone.0326408 |
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