Analysis of factors contributing to the formation of mononuclear cell aggregates (?escapees?) in flow cytometric immunophenotyping

Gratama, Jan W.; Linden, Reinier van der; Bolhuis, R. L. H.; Winkel, Jan G. J. van de

doi:10.1002/(sici)1097-0320(19971101)29:3<250::aid-cyto8>3.0.co;2-f

Cited by 16 publications

(9 citation statements)

References 14 publications

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“…It is known that lysing reagents may affect leucocyte populations differently; some reagents may even cause significant loss of leucocytes, which may contribute to error especially in samples from HIV‐infected patients whose cells may be more susceptible to lysis (11, 25). Artifacts such as cell aggregates, cell debris, background fluorescence and, more importantly, the interference by some monocytes contaminating in the lymphocyte boundary may further lead to lymphocyte impurities (11, 26) and influence CD4+ or CD8+ T‐lymphocyte count accuracy.…”

Section: Discussionmentioning

confidence: 99%

Evaluation of a single‐platform microcapillary flow cytometer for enumeration of absolute CD4+ T‐lymphocyte counts in HIV‐1 infected Thai patients

Pattanapanyasat

Phuang-Ngern

Lerdwana

et al. 2007

Cytometry Part B Clinical

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Background: Various assays are used to enumerate peripheral blood absolute CD4+ T-lymphocytes. Flow cytometry is considered the gold standard for this purpose. However, the high cost of available flow cytometers and monoclonal antibody reagents make it difficult to implement such methods in the resource-poor settings. In this study, we evaluated a cheaper, recently developed single-platform microcapillary cytometer for CD4+ T-lymphocyte enumeration, the personal cell analyzer (PCA), from Guava V V R Technologies. Methods: CD4+ and CD8+ T-lymphocyte counts in whole blood samples from 250 HIV-1 infected Thais were determined, using a two-color reagent kit and the Guava PCA, and compared with the results obtained with two reference microbead-based methods from Becton Dickinson Biosciences: the threecolor TruCOUNT TM tube method and the two-color FACSCount TM method. Statistical correlations and agreements were determined using linear correlation and Bland-Altman analysis.Results: Absolute CD4+ T-lymphocyte counts obtained using the Guava PCA method highly correlated with those obtained using TruCOUNT method (R 2 = 0.95, mean bias +13.1 cells/ml, limit of agreement [LOA] 2117.9 to +144.1 cells/ml) and the FACSCount method (R 2 = 0.94, mean bias = +33.2 cells/ml, LOA 2101.8 to +168.3 cells/ml). Absolute CD8+ T-lymphocyte counts obtained using the Guava PCA method also highly correlated with those obtained with the two reference methods (R 2 = 0.92 and 0.88, respectively).Conclusion: This study shows that the enumeration of CD4+ T-lymphocytes using the Guava microcapillary cytometer PCA method performed well when compared with the two reference bead-based methods. However, like the two reference methods, this new method needs substantial technical expertise. q 2007 Clinical Cytometry Society

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Section: Discussionmentioning

confidence: 99%

Evaluation of a single‐platform microcapillary flow cytometer for enumeration of absolute CD4+ T‐lymphocyte counts in HIV‐1 infected Thai patients

Pattanapanyasat

Phuang-Ngern

Lerdwana

et al. 2007

Cytometry Part B Clinical

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“…Fc-mediated binding to cells that do not express the antigen of interest is in this context considered undesirable antibody binding. The level of this type of undesirable antibody binding can be reduced by blocking Fc-receptors with anti-Fc-receptor antibodies (28), Fab or F(ab) 2 fragments thereof, or with pooled immunoglobulins, or even with whole serum.…”

Section: Background Caused By Undesirablementioning

confidence: 99%

Considerations for the control of background fluorescence in clinical flow cytometry

Hulspas¹,

O’Gorman

Wood

et al. 2009

Cytometry Part B Clinical

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“…A further artefact scenario, described previously in cytometry [10], involves monocytes binding to T cells through Fcg receptors. Antibodies against T lymphocytes are found in the majority of SLE patients and, whereas these are generally detected as cold-reactive IgM, there is also evidence for the presence of IgG antibodies [11].…”

Section: Discussionmentioning

confidence: 94%

Serendipitous evidence of T lymphocyte activation in close female relatives of patients with systemic lupus erythematosus

Green¹,

Kennell²,

Larché

et al. 2009

Clinical and Experimental Immunology

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SummaryA well-recognized characteristic of the autoimmune disease, systemic lupus erythematosus (SLE), is the high level of activated T cells present in the blood. Because of the increased size and granularity of activated T cells, in flow cytometry one might expect to find increased numbers of cells falling outside a standard light-scatter lymphocyte gate, and indeed we now report that the percentage of T lymphocytes in the gate (% TiG) was below the normal range in 23 of 58 (40%) female patients because of increased scatter values. However, the surprising additional observation was made that 18 of 30 (60%) female first-degree relatives of the patients also fell below the normal % TiG range, suggesting the presence of T cell activation in these relatives. This view is strengthened by the strong inverse correlation between plasma total immunoglobulin G(IgG), which was raised in some relatives, and % TiG, as T cell activation is a requirement for IgG production. Conversely, there was no correlation with IgM, which has no comparable link with T cell activation. While a definitive interpretation must await the demonstration of activation antigen expression in relatives, these findings suggest the existence of a T cell activation trait, not harmful in itself, which, however, contributes to the development of disease in patients with SLE.

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Analysis of factors contributing to the formation of mononuclear cell aggregates (?escapees?) in flow cytometric immunophenotyping

Cited by 16 publications

References 14 publications

Evaluation of a single‐platform microcapillary flow cytometer for enumeration of absolute CD4+ T‐lymphocyte counts in HIV‐1 infected Thai patients

Evaluation of a single‐platform microcapillary flow cytometer for enumeration of absolute CD4+ T‐lymphocyte counts in HIV‐1 infected Thai patients

Considerations for the control of background fluorescence in clinical flow cytometry

Serendipitous evidence of T lymphocyte activation in close female relatives of patients with systemic lupus erythematosus

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