Using the Autofluorescence Finder on the Sony ID7000TM Spectral Cell Analyzer to Identify and Unmix Multiple Highly Autofluorescent Murine Lung Populations
Autofluorescence (AF) is a feature of all cell types, though some have more than others. In tissues with complex heterogeneous cellularity, AF is frequently a source of high background, masking faint fluorescent signals and reducing the available dynamic range of detectors for detecting fluorescence...
| Main Authors: | , , , , |
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| Format: | Article |
| Language: | English |
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Frontiers Media S.A.
2022-03-01
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| Series: | Frontiers in Bioengineering and Biotechnology |
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| Online Access: | https://www.frontiersin.org/articles/10.3389/fbioe.2022.827987/full |
| _version_ | 1818695078261030912 |
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| author | Nicholas Wanner Jerry Barnhart Nicholas Apostolakis Violetta Zlojutro Kewal Asosingh Kewal Asosingh |
| author_facet | Nicholas Wanner Jerry Barnhart Nicholas Apostolakis Violetta Zlojutro Kewal Asosingh Kewal Asosingh |
| author_sort | Nicholas Wanner |
| collection | DOAJ |
| description | Autofluorescence (AF) is a feature of all cell types, though some have more than others. In tissues with complex heterogeneous cellularity, AF is frequently a source of high background, masking faint fluorescent signals and reducing the available dynamic range of detectors for detecting fluorescence signals from markers of interest in a flow cytometry panel. Pulmonary flow cytometry presents unique challenges because lung cells are heterogeneous and contain varying amounts of high AF. The goal of this study was to demonstrate how a novel AF Finder tool on the Sony ID7000™ Spectral Cell Analyzer can be used to identify and screen multiple AF subsets in complex highly AF tissues like murine lungs. In lung single cell suspensions, the AF Finder tool identified four distinct AF spectra from six highly AF subsets. The subtraction of these distinct AF spectra resulted in a resolution increase by several log decades in several fluorescent channels. The major immune and lung tissue resident cells in a murine model of asthma were easily identified in a multi-color panel using AF subtraction. The findings demonstrate the practicality of the AF Finder tool, particularly when analyzing samples with multiple AF populations of varying intensities, in order to reduce fluorescence background and increase signal resolution in spectral flow cytometry. |
| first_indexed | 2024-12-17T13:39:45Z |
| format | Article |
| id | doaj.art-c297625d841247c88383c153cf4b71e0 |
| institution | Directory Open Access Journal |
| issn | 2296-4185 |
| language | English |
| last_indexed | 2024-12-17T13:39:45Z |
| publishDate | 2022-03-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Bioengineering and Biotechnology |
| spelling | doaj.art-c297625d841247c88383c153cf4b71e02022-12-21T21:46:19ZengFrontiers Media S.A.Frontiers in Bioengineering and Biotechnology2296-41852022-03-011010.3389/fbioe.2022.827987827987Using the Autofluorescence Finder on the Sony ID7000TM Spectral Cell Analyzer to Identify and Unmix Multiple Highly Autofluorescent Murine Lung PopulationsNicholas Wanner0Jerry Barnhart1Nicholas Apostolakis2Violetta Zlojutro3Kewal Asosingh4Kewal Asosingh5Asosingh Lab, Department of Inflammation and Immunity, Cleveland Clinic, Lerner Research Institute, Cleveland, OH, United StatesSony Biotechnology Inc., San Jose, CA, United StatesAsosingh Lab, Department of Inflammation and Immunity, Cleveland Clinic, Lerner Research Institute, Cleveland, OH, United StatesAsosingh Lab, Department of Inflammation and Immunity, Cleveland Clinic, Lerner Research Institute, Cleveland, OH, United StatesAsosingh Lab, Department of Inflammation and Immunity, Cleveland Clinic, Lerner Research Institute, Cleveland, OH, United StatesFlow Cytometry Core, Cleveland Clinic, Lerner Research Institute, Cleveland, OH, United StatesAutofluorescence (AF) is a feature of all cell types, though some have more than others. In tissues with complex heterogeneous cellularity, AF is frequently a source of high background, masking faint fluorescent signals and reducing the available dynamic range of detectors for detecting fluorescence signals from markers of interest in a flow cytometry panel. Pulmonary flow cytometry presents unique challenges because lung cells are heterogeneous and contain varying amounts of high AF. The goal of this study was to demonstrate how a novel AF Finder tool on the Sony ID7000™ Spectral Cell Analyzer can be used to identify and screen multiple AF subsets in complex highly AF tissues like murine lungs. In lung single cell suspensions, the AF Finder tool identified four distinct AF spectra from six highly AF subsets. The subtraction of these distinct AF spectra resulted in a resolution increase by several log decades in several fluorescent channels. The major immune and lung tissue resident cells in a murine model of asthma were easily identified in a multi-color panel using AF subtraction. The findings demonstrate the practicality of the AF Finder tool, particularly when analyzing samples with multiple AF populations of varying intensities, in order to reduce fluorescence background and increase signal resolution in spectral flow cytometry.https://www.frontiersin.org/articles/10.3389/fbioe.2022.827987/fullautofluorescencespectral flow cytometrymurine lungSony ID7000autofluorescence finder |
| spellingShingle | Nicholas Wanner Jerry Barnhart Nicholas Apostolakis Violetta Zlojutro Kewal Asosingh Kewal Asosingh Using the Autofluorescence Finder on the Sony ID7000TM Spectral Cell Analyzer to Identify and Unmix Multiple Highly Autofluorescent Murine Lung Populations Frontiers in Bioengineering and Biotechnology autofluorescence spectral flow cytometry murine lung Sony ID7000 autofluorescence finder |
| title | Using the Autofluorescence Finder on the Sony ID7000TM Spectral Cell Analyzer to Identify and Unmix Multiple Highly Autofluorescent Murine Lung Populations |
| title_full | Using the Autofluorescence Finder on the Sony ID7000TM Spectral Cell Analyzer to Identify and Unmix Multiple Highly Autofluorescent Murine Lung Populations |
| title_fullStr | Using the Autofluorescence Finder on the Sony ID7000TM Spectral Cell Analyzer to Identify and Unmix Multiple Highly Autofluorescent Murine Lung Populations |
| title_full_unstemmed | Using the Autofluorescence Finder on the Sony ID7000TM Spectral Cell Analyzer to Identify and Unmix Multiple Highly Autofluorescent Murine Lung Populations |
| title_short | Using the Autofluorescence Finder on the Sony ID7000TM Spectral Cell Analyzer to Identify and Unmix Multiple Highly Autofluorescent Murine Lung Populations |
| title_sort | using the autofluorescence finder on the sony id7000tm spectral cell analyzer to identify and unmix multiple highly autofluorescent murine lung populations |
| topic | autofluorescence spectral flow cytometry murine lung Sony ID7000 autofluorescence finder |
| url | https://www.frontiersin.org/articles/10.3389/fbioe.2022.827987/full |
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