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Lymphocyte enrichment using CD81‐targeted immunoaffinity matrix
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| Zeitschriftentitel: | Cytometry Part A |
|---|---|
| Personen und Körperschaften: | , , , , , , , , , , |
| In: | Cytometry Part A, 91, 2017, 1, S. 62-72 |
| Format: | E-Article |
| Sprache: | Englisch |
| veröffentlicht: |
Wiley
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| Schlagwörter: |
| author_facet |
Pelák, Ondřej Kužílková, Daniela Thürner, Daniel Kiene, Marie‐Luise Stanar, Kristian Stuchlý, Jan Vášková, Martina Starý, Jan Hrušák, Ondřej Stadler, Herbert Kalina, Tomáš Pelák, Ondřej Kužílková, Daniela Thürner, Daniel Kiene, Marie‐Luise Stanar, Kristian Stuchlý, Jan Vášková, Martina Starý, Jan Hrušák, Ondřej Stadler, Herbert Kalina, Tomáš |
|---|---|
| author |
Pelák, Ondřej Kužílková, Daniela Thürner, Daniel Kiene, Marie‐Luise Stanar, Kristian Stuchlý, Jan Vášková, Martina Starý, Jan Hrušák, Ondřej Stadler, Herbert Kalina, Tomáš |
| spellingShingle |
Pelák, Ondřej Kužílková, Daniela Thürner, Daniel Kiene, Marie‐Luise Stanar, Kristian Stuchlý, Jan Vášková, Martina Starý, Jan Hrušák, Ondřej Stadler, Herbert Kalina, Tomáš Cytometry Part A Lymphocyte enrichment using CD81‐targeted immunoaffinity matrix Cell Biology Histology Pathology and Forensic Medicine |
| author_sort |
pelák, ondřej |
| spelling |
Pelák, Ondřej Kužílková, Daniela Thürner, Daniel Kiene, Marie‐Luise Stanar, Kristian Stuchlý, Jan Vášková, Martina Starý, Jan Hrušák, Ondřej Stadler, Herbert Kalina, Tomáš 1552-4922 1552-4930 Wiley Cell Biology Histology Pathology and Forensic Medicine http://dx.doi.org/10.1002/cyto.a.22918 <jats:title>Abstract</jats:title><jats:p>In mass cytometry, the isolation of pure lymphocytes is very important to obtain reproducible results and to shorten the time spent on data acquisition. To prepare highly purified cell suspensions of peripheral blood lymphocytes for further analysis on mass cytometer, we used the new CD81+ immune affinity chromatography cell isolation approach. Using 21 metal conjugated antibodies in a single tube we were able to identify all basic cell subsets and compare their relative abundance in final products obtained by density gradient (Ficoll‐Paque) and immune affinity chromatography (CD81+ T‐catch™) isolation approach. We show that T‐catch isolation approach results in purer final product than Ficoll‐Paque (<jats:italic>P</jats:italic> values 0.0156), with fewer platelets bound to target cells. As a result acquisition time of 10<jats:sup>5</jats:sup> nucleated cells was 3.5 shorter. We then applied unsupervised high dimensional analysis viSNE algorithm to compare the two isolation protocols, which allowed us to evaluate the contribution of unsupervised analysis over supervised manual gating. ViSNE algorithm effectively characterized almost all supervised cell subsets. Moreover, viSNE uncovered previously overseen cell subsets and showed inaccuracies in Maxpar™ Human peripheral blood phenotyping panel kit recommended gating strategy. These findings emphasize the use of unsupervised analysis tools in parallel with conventional gating strategy to mine the complete information from a set of samples. They also stress the importance of the impurity removal to sensitively detect rare cell populations in unsupervised analysis. © 2016 International Society for Advancement of Cytometry</jats:p> Lymphocyte enrichment using CD81‐targeted immunoaffinity matrix Cytometry Part A |
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Cytometry Part A |
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| title |
Lymphocyte enrichment using CD81‐targeted immunoaffinity matrix |
| title_unstemmed |
Lymphocyte enrichment using CD81‐targeted immunoaffinity matrix |
| title_full |
Lymphocyte enrichment using CD81‐targeted immunoaffinity matrix |
| title_fullStr |
Lymphocyte enrichment using CD81‐targeted immunoaffinity matrix |
| title_full_unstemmed |
Lymphocyte enrichment using CD81‐targeted immunoaffinity matrix |
| title_short |
Lymphocyte enrichment using CD81‐targeted immunoaffinity matrix |
| title_sort |
lymphocyte enrichment using cd81‐targeted immunoaffinity matrix |
| topic |
Cell Biology Histology Pathology and Forensic Medicine |
| url |
http://dx.doi.org/10.1002/cyto.a.22918 |
| publishDate |
2017 |
| physical |
62-72 |
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<jats:title>Abstract</jats:title><jats:p>In mass cytometry, the isolation of pure lymphocytes is very important to obtain reproducible results and to shorten the time spent on data acquisition. To prepare highly purified cell suspensions of peripheral blood lymphocytes for further analysis on mass cytometer, we used the new CD81+ immune affinity chromatography cell isolation approach. Using 21 metal conjugated antibodies in a single tube we were able to identify all basic cell subsets and compare their relative abundance in final products obtained by density gradient (Ficoll‐Paque) and immune affinity chromatography (CD81+ T‐catch™) isolation approach. We show that T‐catch isolation approach results in purer final product than Ficoll‐Paque (<jats:italic>P</jats:italic> values 0.0156), with fewer platelets bound to target cells. As a result acquisition time of 10<jats:sup>5</jats:sup> nucleated cells was 3.5 shorter. We then applied unsupervised high dimensional analysis viSNE algorithm to compare the two isolation protocols, which allowed us to evaluate the contribution of unsupervised analysis over supervised manual gating. ViSNE algorithm effectively characterized almost all supervised cell subsets. Moreover, viSNE uncovered previously overseen cell subsets and showed inaccuracies in Maxpar™ Human peripheral blood phenotyping panel kit recommended gating strategy. These findings emphasize the use of unsupervised analysis tools in parallel with conventional gating strategy to mine the complete information from a set of samples. They also stress the importance of the impurity removal to sensitively detect rare cell populations in unsupervised analysis. © 2016 International Society for Advancement of Cytometry</jats:p> |
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| author | Pelák, Ondřej, Kužílková, Daniela, Thürner, Daniel, Kiene, Marie‐Luise, Stanar, Kristian, Stuchlý, Jan, Vášková, Martina, Starý, Jan, Hrušák, Ondřej, Stadler, Herbert, Kalina, Tomáš |
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| description | <jats:title>Abstract</jats:title><jats:p>In mass cytometry, the isolation of pure lymphocytes is very important to obtain reproducible results and to shorten the time spent on data acquisition. To prepare highly purified cell suspensions of peripheral blood lymphocytes for further analysis on mass cytometer, we used the new CD81+ immune affinity chromatography cell isolation approach. Using 21 metal conjugated antibodies in a single tube we were able to identify all basic cell subsets and compare their relative abundance in final products obtained by density gradient (Ficoll‐Paque) and immune affinity chromatography (CD81+ T‐catch™) isolation approach. We show that T‐catch isolation approach results in purer final product than Ficoll‐Paque (<jats:italic>P</jats:italic> values 0.0156), with fewer platelets bound to target cells. As a result acquisition time of 10<jats:sup>5</jats:sup> nucleated cells was 3.5 shorter. We then applied unsupervised high dimensional analysis viSNE algorithm to compare the two isolation protocols, which allowed us to evaluate the contribution of unsupervised analysis over supervised manual gating. ViSNE algorithm effectively characterized almost all supervised cell subsets. Moreover, viSNE uncovered previously overseen cell subsets and showed inaccuracies in Maxpar™ Human peripheral blood phenotyping panel kit recommended gating strategy. These findings emphasize the use of unsupervised analysis tools in parallel with conventional gating strategy to mine the complete information from a set of samples. They also stress the importance of the impurity removal to sensitively detect rare cell populations in unsupervised analysis. © 2016 International Society for Advancement of Cytometry</jats:p> |
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| spelling | Pelák, Ondřej Kužílková, Daniela Thürner, Daniel Kiene, Marie‐Luise Stanar, Kristian Stuchlý, Jan Vášková, Martina Starý, Jan Hrušák, Ondřej Stadler, Herbert Kalina, Tomáš 1552-4922 1552-4930 Wiley Cell Biology Histology Pathology and Forensic Medicine http://dx.doi.org/10.1002/cyto.a.22918 <jats:title>Abstract</jats:title><jats:p>In mass cytometry, the isolation of pure lymphocytes is very important to obtain reproducible results and to shorten the time spent on data acquisition. To prepare highly purified cell suspensions of peripheral blood lymphocytes for further analysis on mass cytometer, we used the new CD81+ immune affinity chromatography cell isolation approach. Using 21 metal conjugated antibodies in a single tube we were able to identify all basic cell subsets and compare their relative abundance in final products obtained by density gradient (Ficoll‐Paque) and immune affinity chromatography (CD81+ T‐catch™) isolation approach. We show that T‐catch isolation approach results in purer final product than Ficoll‐Paque (<jats:italic>P</jats:italic> values 0.0156), with fewer platelets bound to target cells. As a result acquisition time of 10<jats:sup>5</jats:sup> nucleated cells was 3.5 shorter. We then applied unsupervised high dimensional analysis viSNE algorithm to compare the two isolation protocols, which allowed us to evaluate the contribution of unsupervised analysis over supervised manual gating. ViSNE algorithm effectively characterized almost all supervised cell subsets. Moreover, viSNE uncovered previously overseen cell subsets and showed inaccuracies in Maxpar™ Human peripheral blood phenotyping panel kit recommended gating strategy. These findings emphasize the use of unsupervised analysis tools in parallel with conventional gating strategy to mine the complete information from a set of samples. They also stress the importance of the impurity removal to sensitively detect rare cell populations in unsupervised analysis. © 2016 International Society for Advancement of Cytometry</jats:p> Lymphocyte enrichment using CD81‐targeted immunoaffinity matrix Cytometry Part A |
| spellingShingle | Pelák, Ondřej, Kužílková, Daniela, Thürner, Daniel, Kiene, Marie‐Luise, Stanar, Kristian, Stuchlý, Jan, Vášková, Martina, Starý, Jan, Hrušák, Ondřej, Stadler, Herbert, Kalina, Tomáš, Cytometry Part A, Lymphocyte enrichment using CD81‐targeted immunoaffinity matrix, Cell Biology, Histology, Pathology and Forensic Medicine |
| title | Lymphocyte enrichment using CD81‐targeted immunoaffinity matrix |
| title_full | Lymphocyte enrichment using CD81‐targeted immunoaffinity matrix |
| title_fullStr | Lymphocyte enrichment using CD81‐targeted immunoaffinity matrix |
| title_full_unstemmed | Lymphocyte enrichment using CD81‐targeted immunoaffinity matrix |
| title_short | Lymphocyte enrichment using CD81‐targeted immunoaffinity matrix |
| title_sort | lymphocyte enrichment using cd81‐targeted immunoaffinity matrix |
| title_unstemmed | Lymphocyte enrichment using CD81‐targeted immunoaffinity matrix |
| topic | Cell Biology, Histology, Pathology and Forensic Medicine |
| url | http://dx.doi.org/10.1002/cyto.a.22918 |