Direct labeling of milk cells without centrifugation for counting total and differential somatic cells using flow cytometry
ABSTRACT: Somatic cell count (SCC) in milk is an essential indicator for defining and managing udder health. However, analyzing differential SCC (dSCC) can be helpful in determining the type or evolution stage of mastitis. A high abundance of polymorphonuclear cells (PMN) is associated with acute ma...
| Main Authors: | , , , , , |
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| Format: | Article |
| Language: | English |
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Elsevier
2022-11-01
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| Series: | Journal of Dairy Science |
| Subjects: | |
| Online Access: | http://www.sciencedirect.com/science/article/pii/S0022030222005537 |
| _version_ | 1797990646051504128 |
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| author | Jérôme Widmer Laurence Descloux Cédric Brügger Marielle-Louise Jäger Thomas Berger Lotti Egger |
| author_facet | Jérôme Widmer Laurence Descloux Cédric Brügger Marielle-Louise Jäger Thomas Berger Lotti Egger |
| author_sort | Jérôme Widmer |
| collection | DOAJ |
| description | ABSTRACT: Somatic cell count (SCC) in milk is an essential indicator for defining and managing udder health. However, analyzing differential SCC (dSCC) can be helpful in determining the type or evolution stage of mastitis. A high abundance of polymorphonuclear cells (PMN) is associated with acute mastitis; however, the status of a chronic disease is less well characterized. A method capable of analyzing SCC and dSCC can prove to be a helpful tool for monitoring the status of evolution of mastitis disease in a better way. Therefore, a new direct-flow cytometry method was developed to count and differentiate somatic cells in milk without the steps of centrifugation or washing, avoiding variabilities that occur due to enrichment or loss of specific cell types. In this new method, SCC is analyzed using the method of DNA staining with Hoechst stain, whereas dSCC are analyzed using specific antibodies targeting 2 main cell types associated with mastitis: PMN cells and antigen-presenting cells, which are associated with innate and adaptive immunity. Equivalent SCC values were obtained between the new method and the routine ISO 13366-2 method in a comparison of 240 raw milk samples. Furthermore, dSCC results were confirmed by microscopy after May-Gründwald-Giemsa staining in 165 quarter milk samples from healthy and diseased cows. The method was verified with fluorescence microscopy on the 2 targeted cell types and in raw milk samples. The newly developed method is independent of any instrument and can be further designed to differentiate other cell types and animal species by selecting appropriate antibodies. |
| first_indexed | 2024-04-11T08:38:41Z |
| format | Article |
| id | doaj.art-5eb56d458fc844ef96acaaa77e7c9fe1 |
| institution | Directory Open Access Journal |
| issn | 0022-0302 |
| language | English |
| last_indexed | 2024-04-11T08:38:41Z |
| publishDate | 2022-11-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Journal of Dairy Science |
| spelling | doaj.art-5eb56d458fc844ef96acaaa77e7c9fe12022-12-22T04:34:15ZengElsevierJournal of Dairy Science0022-03022022-11-011051187058717Direct labeling of milk cells without centrifugation for counting total and differential somatic cells using flow cytometryJérôme Widmer0Laurence Descloux1Cédric Brügger2Marielle-Louise Jäger3Thomas Berger4Lotti Egger5Method Development and Analytics, Agroscope Liebefeld, 3003 Bern, SwitzerlandMethod Development and Analytics, Agroscope Liebefeld, 3003 Bern, SwitzerlandMethod Development and Analytics, Agroscope Liebefeld, 3003 Bern, SwitzerlandMethod Development and Analytics, Agroscope Liebefeld, 3003 Bern, SwitzerlandMethod Development and Analytics, Agroscope Liebefeld, 3003 Bern, SwitzerlandCorresponding author; Method Development and Analytics, Agroscope Liebefeld, 3003 Bern, SwitzerlandABSTRACT: Somatic cell count (SCC) in milk is an essential indicator for defining and managing udder health. However, analyzing differential SCC (dSCC) can be helpful in determining the type or evolution stage of mastitis. A high abundance of polymorphonuclear cells (PMN) is associated with acute mastitis; however, the status of a chronic disease is less well characterized. A method capable of analyzing SCC and dSCC can prove to be a helpful tool for monitoring the status of evolution of mastitis disease in a better way. Therefore, a new direct-flow cytometry method was developed to count and differentiate somatic cells in milk without the steps of centrifugation or washing, avoiding variabilities that occur due to enrichment or loss of specific cell types. In this new method, SCC is analyzed using the method of DNA staining with Hoechst stain, whereas dSCC are analyzed using specific antibodies targeting 2 main cell types associated with mastitis: PMN cells and antigen-presenting cells, which are associated with innate and adaptive immunity. Equivalent SCC values were obtained between the new method and the routine ISO 13366-2 method in a comparison of 240 raw milk samples. Furthermore, dSCC results were confirmed by microscopy after May-Gründwald-Giemsa staining in 165 quarter milk samples from healthy and diseased cows. The method was verified with fluorescence microscopy on the 2 targeted cell types and in raw milk samples. The newly developed method is independent of any instrument and can be further designed to differentiate other cell types and animal species by selecting appropriate antibodies.http://www.sciencedirect.com/science/article/pii/S0022030222005537total somatic cells in milkmastitisdifferential somatic cells in milkflow cytometry |
| spellingShingle | Jérôme Widmer Laurence Descloux Cédric Brügger Marielle-Louise Jäger Thomas Berger Lotti Egger Direct labeling of milk cells without centrifugation for counting total and differential somatic cells using flow cytometry Journal of Dairy Science total somatic cells in milk mastitis differential somatic cells in milk flow cytometry |
| title | Direct labeling of milk cells without centrifugation for counting total and differential somatic cells using flow cytometry |
| title_full | Direct labeling of milk cells without centrifugation for counting total and differential somatic cells using flow cytometry |
| title_fullStr | Direct labeling of milk cells without centrifugation for counting total and differential somatic cells using flow cytometry |
| title_full_unstemmed | Direct labeling of milk cells without centrifugation for counting total and differential somatic cells using flow cytometry |
| title_short | Direct labeling of milk cells without centrifugation for counting total and differential somatic cells using flow cytometry |
| title_sort | direct labeling of milk cells without centrifugation for counting total and differential somatic cells using flow cytometry |
| topic | total somatic cells in milk mastitis differential somatic cells in milk flow cytometry |
| url | http://www.sciencedirect.com/science/article/pii/S0022030222005537 |
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