Improved Standardization of Flow Cytometry Diagnostic Screening of Primary Immunodeficiency by Software-Based Automated Gating

Linskens, Eleni A.; Diks, Annieck M; Neirinck, Jana; Pérez-Andrés, Martín; Maertelaere, Emilie De; Berkowska, Magdalena A.; Kerre, Tessa; Hofmans, Mattias; Órfão, Alberto; Dongen, Jacques J.M. van; Haerynck, Filomeen; Philippé, Jan; Bonroy, Carolien

doi:10.3389/fimmu.2020.584646

Cited by 16 publications

(19 citation statements)

References 22 publications

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“…It is dif cult to de ne an appropriate gating method due to the limited reference standard and analyzed sample numbers [16]. In the present study, certain inconsistent results were observed from post-HSCT patients, presumably because post-HSCT patients are more likely to develop aberrant cell expression or unexpected phenotyping cells during the engraftment that are not observed in healthy individuals.…”

Section: Discussionmentioning

confidence: 65%

Comparison of Automated and Manual Gating of Lymphocyte Subsets in Hematopoietic Stem Cell Transplantation Recipients

et al. 2022

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Background: Lymphocyte subset analysis is essential to evaluate the engraftment status in hematopoietic stem cell transplantation (HSCT). Automated gating tools are widely used for flow cytometry analysis. Unlike healthy individuals, different cell populations and aberrant expressions may occur in HSCT samples. In the present study, we evaluated the applicability of automated gating in HSCT recipients by comparing it to expertbased manual gating. Methods: Lymphocyte subset was performed using Beckman Coulter Navios (Beckman Coulter, USA) flow cytometry. Data files from 22 patients with hematologic malignancies were analyzed in parallel by manual gating and automated gating using Navios Tetra software. Quality control results and reproducibility were evaluated using IMMUNO-TROL controls. Results: Spearman rank correlation coefficients between the two gating methods were > 0.970 in all cell populations except CD8+ T cells. CD8+ T cell counts via automated gating were higher than those of manual gating in all cases due to the T cell populations with reduced CD8 expression. Automated gating program failed to identify CD4+CD8+ double-positive T cell population. Moreover, it excluded certain lymphocytes with low forward scatter (FSC) and high side scatter (SSC). Furthermore, two HSCT recipients revealed a high percentage of CD56−CD16+ NK cells, we found the need to add CD16 reagent to the Navios system. All coefficients of variation were < 10% except for CD56+ NK cells via automated gating. Conclusions: Manual gating confirmation via flow cytometry histogram is necessary to identify the aberrant phenotypes and unexpected cell populations in HSCT recipients.

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Section: Discussionmentioning

confidence: 65%

Comparison of Automated and Manual Gating of Lymphocyte Subsets in Hematopoietic Stem Cell Transplantation Recipients

et al. 2022

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“…To ensure objective data analysis, we used the automated gating and identification (AGI) tool in the Infinicyt software. This tool was shown to reduce intra-and interoperator variability and increase reproducibility of the analysis [40,[48][49][50]. This is especially important for studies with big data from multiple samples, which cannot be analyzed by a single operator within a reasonable timeframe.…”

Section: Discussionmentioning

confidence: 99%

Age and Primary Vaccination Background Influence the Plasma Cell Response to Pertussis Booster Vaccination

et al. 2022

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Pertussis is a vaccine-preventable disease caused by the bacterium Bordetella pertussis. Over the past years, the incidence and mortality of pertussis increased significantly. A possible cause is the switch from whole-cell to acellular pertussis vaccines, although other factors may also contribute. Here, we applied high-dimensional flow cytometry to investigate changes in B cells in individuals of different ages and distinct priming backgrounds upon administration of an acellular pertussis booster vaccine. Participants were divided over four age cohorts. We compared longitudinal kinetics within each cohort and between the different cohorts. Changes in the B-cell compartment were correlated to numbers of vaccine-specific B- and plasma cells and serum Ig levels. Expansion and maturation of plasma cells 7 days postvaccination was the most prominent cellular change in all age groups and was most pronounced for more mature IgG1+ plasma cells. Plasma cell responses were stronger in individuals primed with whole-cell vaccine than in individuals primed with acellular vaccine. Moreover, IgG1+ and IgA1+ plasma cell expansion correlated with FHA-, Prn-, or PT- specific serum IgG or IgA levels. Our study indicates plasma cells as a potential early cellular marker of an immune response and contributes to understanding differences in immune responses between age groups and primary vaccination backgrounds.

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“…To ensure objective data analysis, we used the automated gating and identification (AGI) tool in the Infinicyt software, which has been demonstrated to reduce intra-and inter-operator variability and increase reproducibility of the analysis [40,[48][49][50]. This is especially important for studies with big data from multiple samples, which cannot be analyzed by a single operator within a reasonable timeframe.…”

Section: Discussionmentioning

confidence: 99%

Evaluation of B-Cell Kinetics After Acellular Pertussis Vaccination in Four Cohorts of Different Age and Priming Background

Diks¹,

Versteegen²,

Teodósio³

et al. 2021

Preprint

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Pertussis is a vaccine-preventable disease caused by the bacterium Bordetella pertussis. Over the past years, the incidence and mortality of pertussis increased significantly. A possible cause is the switch from whole cell to acellular pertussis vaccines, although other factors may also contribute. To develop future vaccines and improve current vaccination strategies, it is critical to understand factors influencing the generation of immunological memory. We applied high-dimensional flow cytometry to investigate changes in B cells in individuals of different ages and distinct priming backgrounds upon administration of an acellular pertussis booster vaccine. These findings were correlated to vaccine-specific plasma cells and serum Ig levels. Expansion and maturation of plasma cells 7 days post-vaccination was the most prominent cellular change in all age groups, and was most pronounced for more mature IgG1+ plasma cells. Cellular responses were stronger in individuals primed with whole cell vaccine than in individuals primed with acellular vaccine. Moreover, IgG1+ plasma cell expansion weakly correlated with Prn- and PT- specific serum IgG levels. Our study points at plasma cells as a potential early cellular marker of an immune response and contributes to understanding differences in immune responses between age groups and priming backgrounds.

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Improved Standardization of Flow Cytometry Diagnostic Screening of Primary Immunodeficiency by Software-Based Automated Gating

Cited by 16 publications

References 22 publications

Comparison of Automated and Manual Gating of Lymphocyte Subsets in Hematopoietic Stem Cell Transplantation Recipients

Comparison of Automated and Manual Gating of Lymphocyte Subsets in Hematopoietic Stem Cell Transplantation Recipients

Age and Primary Vaccination Background Influence the Plasma Cell Response to Pertussis Booster Vaccination

Evaluation of B-Cell Kinetics After Acellular Pertussis Vaccination in Four Cohorts of Different Age and Priming Background

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