Comparison of NET quantification methods based on immunofluorescence microscopy: Hand-counting, semi-automated and automated evaluations
Formation of neutrophil extracellular traps was first described in 2004, showing that NETs are composed of decondensed chromatin fibers and nuclear and granule components. Free DNA is often used to quantify NETs, but to differentiate NETosis from necrotic DNA-release, immunofluorescence microscopy w...
| Main Authors: | , , , , , , , , , , |
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| Format: | Article |
| Language: | English |
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Elsevier
2023-06-01
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| Series: | Heliyon |
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| Online Access: | http://www.sciencedirect.com/science/article/pii/S2405844023041890 |
| _version_ | 1797808428338380800 |
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| author | Timo Henneck Christina Krüger Andreas Nerlich Melissa Langer Leonie Fingerhut Marta C. Bonilla Marita Meurer Sönke von den Berg Nicole de Buhr Katja Branitzki-Heinemann Maren von Köckritz-Blickwede |
| author_facet | Timo Henneck Christina Krüger Andreas Nerlich Melissa Langer Leonie Fingerhut Marta C. Bonilla Marita Meurer Sönke von den Berg Nicole de Buhr Katja Branitzki-Heinemann Maren von Köckritz-Blickwede |
| author_sort | Timo Henneck |
| collection | DOAJ |
| description | Formation of neutrophil extracellular traps was first described in 2004, showing that NETs are composed of decondensed chromatin fibers and nuclear and granule components. Free DNA is often used to quantify NETs, but to differentiate NETosis from necrotic DNA-release, immunofluorescence microscopy with NET-specific markers is required. Although evaluation by hand is time-consuming and difficult to standardize, it is still widespread. Unfortunately, no standardized method and only limited software tools are available for NET evaluation. This study provides an overview of recent techniques in use and aims to compare two published computer-based methods with hand counting. We found that the selected semi-automated quantification method and fully automated quantification via NETQUANT differed significantly from results obtained by hand and exhibited problems in detection of complex NET structures with partially illogical results. In contrast to that, trained persons were able to adapt to varying settings. Future approaches aimed at developing deep-learning algorithms for fast and reproducible quantification of NETs are needed. |
| first_indexed | 2024-03-13T06:37:21Z |
| format | Article |
| id | doaj.art-b3581cf2ab59418cb5cb794d761cb183 |
| institution | Directory Open Access Journal |
| issn | 2405-8440 |
| language | English |
| last_indexed | 2024-03-13T06:37:21Z |
| publishDate | 2023-06-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Heliyon |
| spelling | doaj.art-b3581cf2ab59418cb5cb794d761cb1832023-06-09T04:28:40ZengElsevierHeliyon2405-84402023-06-0196e16982Comparison of NET quantification methods based on immunofluorescence microscopy: Hand-counting, semi-automated and automated evaluationsTimo Henneck0Christina Krüger1Andreas Nerlich2Melissa Langer3Leonie Fingerhut4Marta C. Bonilla5Marita Meurer6Sönke von den Berg7Nicole de Buhr8Katja Branitzki-Heinemann9Maren von Köckritz-Blickwede10Institute of Biochemistry, University of Veterinary Medicine Hannover, Foundation, 30559, Hannover, GermanyInstitute of Biochemistry, University of Veterinary Medicine Hannover, Foundation, 30559, Hannover, GermanyInstitute for Microbiology, University of Veterinary Medicine Hannover, Foundation, 30173, Hannover, GermanyInstitute of Biochemistry, University of Veterinary Medicine Hannover, Foundation, 30559, Hannover, GermanyInstitute of Biochemistry, University of Veterinary Medicine Hannover, Foundation, 30559, Hannover, Germany; Research Center for Emerging Infections and Zoonoses (RIZ), University of Veterinary Medicine Hannover, Foundation, 30559, Hannover, GermanyInstitute of Biochemistry, University of Veterinary Medicine Hannover, Foundation, 30559, Hannover, Germany; Research Center for Emerging Infections and Zoonoses (RIZ), University of Veterinary Medicine Hannover, Foundation, 30559, Hannover, GermanyInstitute of Biochemistry, University of Veterinary Medicine Hannover, Foundation, 30559, Hannover, Germany; Research Center for Emerging Infections and Zoonoses (RIZ), University of Veterinary Medicine Hannover, Foundation, 30559, Hannover, GermanyInstitute for Zoology, University of Veterinary Medicine Hannover, Foundation, 30559, Hannover, GermanyInstitute of Biochemistry, University of Veterinary Medicine Hannover, Foundation, 30559, Hannover, Germany; Research Center for Emerging Infections and Zoonoses (RIZ), University of Veterinary Medicine Hannover, Foundation, 30559, Hannover, GermanyInstitute of Biochemistry, University of Veterinary Medicine Hannover, Foundation, 30559, Hannover, Germany; Research Center for Emerging Infections and Zoonoses (RIZ), University of Veterinary Medicine Hannover, Foundation, 30559, Hannover, GermanyInstitute of Biochemistry, University of Veterinary Medicine Hannover, Foundation, 30559, Hannover, Germany; Research Center for Emerging Infections and Zoonoses (RIZ), University of Veterinary Medicine Hannover, Foundation, 30559, Hannover, Germany; Corresponding author. Department of Biochemistry, University of Veterinary Medicine Hannover, Foundation, Bünteweg 17, 30559, Hannover, Germany.Formation of neutrophil extracellular traps was first described in 2004, showing that NETs are composed of decondensed chromatin fibers and nuclear and granule components. Free DNA is often used to quantify NETs, but to differentiate NETosis from necrotic DNA-release, immunofluorescence microscopy with NET-specific markers is required. Although evaluation by hand is time-consuming and difficult to standardize, it is still widespread. Unfortunately, no standardized method and only limited software tools are available for NET evaluation. This study provides an overview of recent techniques in use and aims to compare two published computer-based methods with hand counting. We found that the selected semi-automated quantification method and fully automated quantification via NETQUANT differed significantly from results obtained by hand and exhibited problems in detection of complex NET structures with partially illogical results. In contrast to that, trained persons were able to adapt to varying settings. Future approaches aimed at developing deep-learning algorithms for fast and reproducible quantification of NETs are needed.http://www.sciencedirect.com/science/article/pii/S2405844023041890NETsImageJQuantificationCell countingNETQUANTIn vitro |
| spellingShingle | Timo Henneck Christina Krüger Andreas Nerlich Melissa Langer Leonie Fingerhut Marta C. Bonilla Marita Meurer Sönke von den Berg Nicole de Buhr Katja Branitzki-Heinemann Maren von Köckritz-Blickwede Comparison of NET quantification methods based on immunofluorescence microscopy: Hand-counting, semi-automated and automated evaluations Heliyon NETs ImageJ Quantification Cell counting NETQUANT In vitro |
| title | Comparison of NET quantification methods based on immunofluorescence microscopy: Hand-counting, semi-automated and automated evaluations |
| title_full | Comparison of NET quantification methods based on immunofluorescence microscopy: Hand-counting, semi-automated and automated evaluations |
| title_fullStr | Comparison of NET quantification methods based on immunofluorescence microscopy: Hand-counting, semi-automated and automated evaluations |
| title_full_unstemmed | Comparison of NET quantification methods based on immunofluorescence microscopy: Hand-counting, semi-automated and automated evaluations |
| title_short | Comparison of NET quantification methods based on immunofluorescence microscopy: Hand-counting, semi-automated and automated evaluations |
| title_sort | comparison of net quantification methods based on immunofluorescence microscopy hand counting semi automated and automated evaluations |
| topic | NETs ImageJ Quantification Cell counting NETQUANT In vitro |
| url | http://www.sciencedirect.com/science/article/pii/S2405844023041890 |
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