Human umbilical cord blood cells suffer major modification by fixatives and anticoagulants
Introduction: Developing techniques for the tagless isolation of homogeneous cell populations in physiological-like conditions is of great interest in medical research. A particular case is Gravitational Field-Flow Fractionation (GrFFF), which can be run avoiding cell fixation, and that was already...
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| Format: | Article |
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Frontiers Media S.A.
2023-03-01
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| Series: | Frontiers in Physiology |
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| Online Access: | https://www.frontiersin.org/articles/10.3389/fphys.2023.1070474/full |
| _version_ | 1827989972159823872 |
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| author | Roberta Danusso Roberta Danusso Riccardo Rosati Luca Possenti Elena Lombardini Francesca Gigli Maria Laura Costantino Enrico Ferrazzi Giustina Casagrande Giustina Casagrande Debora Lattuada |
| author_facet | Roberta Danusso Roberta Danusso Riccardo Rosati Luca Possenti Elena Lombardini Francesca Gigli Maria Laura Costantino Enrico Ferrazzi Giustina Casagrande Giustina Casagrande Debora Lattuada |
| author_sort | Roberta Danusso |
| collection | DOAJ |
| description | Introduction: Developing techniques for the tagless isolation of homogeneous cell populations in physiological-like conditions is of great interest in medical research. A particular case is Gravitational Field-Flow Fractionation (GrFFF), which can be run avoiding cell fixation, and that was already used to separate viable cells. Cell dimensions have a key role in this process. However, their dimensions under physiological-like conditions are not easily known since the most diffused measurement techniques are performed on fixed cells, and the fixation used to preserve tissues can alter the cell size. This work aims to obtain and compare cell size data under physiological-like conditions and in the presence of a fixative.Methods: We developed a new protocol that allows the analysis of blood cells in different conditions. Then, we applied it to obtain a dataset of human cord blood cell dimensions from 32 subjects, comparing two tubes with anticoagulants (EDTA and Citrate) and two tubes with different preservatives (CellRescue and CellSave). We analyzed a total of 2071 cells by using confocal microscopy via bio-imaging to assess dimensions (cellular and nuclear) and morphology.Results: Cell diameter measured does not differ when using the different anticoagulants, except for the increase reported for monocyte in the presence of citrate. Instead, cell dimensions differ when comparing anticoagulants and cell preservative tubes, with a few exceptions. Cells characterized by high cytoplasm content show a reduction in their size, while morphology appears always preserved. In a subgroup of cells, 3D reconstruction was performed. Cell and nucleus volumes were estimated using different methods (specific 3D tool or reconstruction from 2D projection).Discussion: We found that some cell types benefit from a complete 3D analysis because they contain non-spherical structures (mainly for cells characterized by poly-lobated nucleus). Overall, we showed the effect of the preservatives mixture on cell dimensions. Such an effect must be considered when dealing with problems highly dependent on cell size, such as GrFFF. Additionally, such information is crucial in computational models increasingly being employed to simulate biological events. |
| first_indexed | 2024-04-10T00:28:42Z |
| format | Article |
| id | doaj.art-5ecb923043484690a1b22f9bc5e492d2 |
| institution | Directory Open Access Journal |
| issn | 1664-042X |
| language | English |
| last_indexed | 2024-04-10T00:28:42Z |
| publishDate | 2023-03-01 |
| publisher | Frontiers Media S.A. |
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| series | Frontiers in Physiology |
| spelling | doaj.art-5ecb923043484690a1b22f9bc5e492d22023-03-15T05:09:59ZengFrontiers Media S.A.Frontiers in Physiology1664-042X2023-03-011410.3389/fphys.2023.10704741070474Human umbilical cord blood cells suffer major modification by fixatives and anticoagulantsRoberta Danusso0Roberta Danusso1Riccardo Rosati2Luca Possenti3Elena Lombardini4Francesca Gigli5Maria Laura Costantino6Enrico Ferrazzi7Giustina Casagrande8Giustina Casagrande9Debora Lattuada10Department of Women-Child-Newborn, Foundation IRCCS Ca’ Granda Ospedale Maggiore Policlinico, Milano, ItalyDepartment of Medicine and Surgery, University of Milano-Bicocca, Monza, ItalyDepartment of Women-Child-Newborn, Foundation IRCCS Ca’ Granda Ospedale Maggiore Policlinico, Milano, ItalyLaBS, Department of Chemistry, Materials and Chemical Engineering “Giulio Natta”, Politecnico di Milano, Milan, ItalyDepartment of Women-Child-Newborn, Foundation IRCCS Ca’ Granda Ospedale Maggiore Policlinico, Milano, ItalyDepartment of Women-Child-Newborn, Foundation IRCCS Ca’ Granda Ospedale Maggiore Policlinico, Milano, ItalyLaBS, Department of Chemistry, Materials and Chemical Engineering “Giulio Natta”, Politecnico di Milano, Milan, ItalyDepartment of Women-Child-Newborn, Foundation IRCCS Ca’ Granda Ospedale Maggiore Policlinico, Milano, ItalyDepartment of Women-Child-Newborn, Foundation IRCCS Ca’ Granda Ospedale Maggiore Policlinico, Milano, ItalyLaBS, Department of Chemistry, Materials and Chemical Engineering “Giulio Natta”, Politecnico di Milano, Milan, ItalyDepartment of Women-Child-Newborn, Foundation IRCCS Ca’ Granda Ospedale Maggiore Policlinico, Milano, ItalyIntroduction: Developing techniques for the tagless isolation of homogeneous cell populations in physiological-like conditions is of great interest in medical research. A particular case is Gravitational Field-Flow Fractionation (GrFFF), which can be run avoiding cell fixation, and that was already used to separate viable cells. Cell dimensions have a key role in this process. However, their dimensions under physiological-like conditions are not easily known since the most diffused measurement techniques are performed on fixed cells, and the fixation used to preserve tissues can alter the cell size. This work aims to obtain and compare cell size data under physiological-like conditions and in the presence of a fixative.Methods: We developed a new protocol that allows the analysis of blood cells in different conditions. Then, we applied it to obtain a dataset of human cord blood cell dimensions from 32 subjects, comparing two tubes with anticoagulants (EDTA and Citrate) and two tubes with different preservatives (CellRescue and CellSave). We analyzed a total of 2071 cells by using confocal microscopy via bio-imaging to assess dimensions (cellular and nuclear) and morphology.Results: Cell diameter measured does not differ when using the different anticoagulants, except for the increase reported for monocyte in the presence of citrate. Instead, cell dimensions differ when comparing anticoagulants and cell preservative tubes, with a few exceptions. Cells characterized by high cytoplasm content show a reduction in their size, while morphology appears always preserved. In a subgroup of cells, 3D reconstruction was performed. Cell and nucleus volumes were estimated using different methods (specific 3D tool or reconstruction from 2D projection).Discussion: We found that some cell types benefit from a complete 3D analysis because they contain non-spherical structures (mainly for cells characterized by poly-lobated nucleus). Overall, we showed the effect of the preservatives mixture on cell dimensions. Such an effect must be considered when dealing with problems highly dependent on cell size, such as GrFFF. Additionally, such information is crucial in computational models increasingly being employed to simulate biological events.https://www.frontiersin.org/articles/10.3389/fphys.2023.1070474/fullcell dimensionumbilical cord blood cellsfixativescell preservativesanticoagulantsbio-image analysis |
| spellingShingle | Roberta Danusso Roberta Danusso Riccardo Rosati Luca Possenti Elena Lombardini Francesca Gigli Maria Laura Costantino Enrico Ferrazzi Giustina Casagrande Giustina Casagrande Debora Lattuada Human umbilical cord blood cells suffer major modification by fixatives and anticoagulants Frontiers in Physiology cell dimension umbilical cord blood cells fixatives cell preservatives anticoagulants bio-image analysis |
| title | Human umbilical cord blood cells suffer major modification by fixatives and anticoagulants |
| title_full | Human umbilical cord blood cells suffer major modification by fixatives and anticoagulants |
| title_fullStr | Human umbilical cord blood cells suffer major modification by fixatives and anticoagulants |
| title_full_unstemmed | Human umbilical cord blood cells suffer major modification by fixatives and anticoagulants |
| title_short | Human umbilical cord blood cells suffer major modification by fixatives and anticoagulants |
| title_sort | human umbilical cord blood cells suffer major modification by fixatives and anticoagulants |
| topic | cell dimension umbilical cord blood cells fixatives cell preservatives anticoagulants bio-image analysis |
| url | https://www.frontiersin.org/articles/10.3389/fphys.2023.1070474/full |
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