Influence of 4‐ and 6‐color flow cytometers and acquisition/analysis softwares on the determination of lymphocyte subsets in HIV infection

Ashman, Margarita; Sachdeva, Naresh; D'Avila, Leonardo Hekman; Scott, Gwendolyn B.; Mitchell, Charles D.; Cintron, L.; Rathore, Mahendra Singh; Asthana, Deshratn

doi:10.1002/cyto.b.20178

Cited by 13 publications

(13 citation statements)

References 13 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…These findings need further investigation as the reduced T-cell function in smokers will have long lasting effects on overall immune response to vaccinations or other infections. Reduced IL-2 production by CD4+ T-cells has been associated with decreased antibody responses to influenza vaccination in older people [40], [41]. The quality of CD4+ T-cells in smokers has been significantly affected and this defect in their immune function could explain why smokers develop serious complications during an infectious disease.…”

Section: Discussionmentioning

confidence: 99%

Tobacco Smoking Increases Immune Activation and Impairs T-Cell Function in HIV Infected Patients on Antiretrovirals: A Cross-Sectional Pilot Study

et al. 2014

View full text Add to dashboard Cite

BackgroundThe influence of tobacco smoking on the immune system of HIV infected individuals is largely unknown. We investigated the impact of tobacco smoking on immune activation, microbial translocation, immune exhaustion and T-cell function in HIV infected individuals.MethodHIV infected smokers and non-smokers (n = 25 each) with documented viral suppression on combination antiretroviral therapy and HIV uninfected smokers and non-smokers (n = 15 each) were enrolled. Markers of immune activation (CD38 and HLA-DR) and immune exhaustion (PD1, Tim3 and CTLA4) were analyzed in peripheral blood mononuclear cells (PBMCs) by flow cytometry. Plasma markers of microbial translocation (soluble-CD14 - sCD14 and lipopolysaccharide - LPS) were measured. Antigen specific functions of CD4+ and CD8+ T-cells were measured, by flow cytometry, in PBMCs after 6 hours stimulation with Cytomegalovirus, Epstein-Barr virus and Influenza Virus (CEF) peptide pool.ResultsCompared to non-smokers, smokers of HIV infected and uninfected groups showed significantly higher CD4+ and CD8+ T-cell activation with increased frequencies of CD38+HLA-DR+ cells with a higher magnitude in HIV infected smokers. Expressions of immune exhaustion markers (PD1, Tim3 and CTLA4) either alone or in combinations were significantly higher in smokers, especially on CD4+ T-cells. Compared to HIV uninfected non-smokers, microbial translocation (sCD14 and LPS) was higher in smokers of both groups and directly correlated with CD4+ and CD8+ T-cell activation. Antigen specific T-cell function showed significantly lower cytokine response of CD4+ and CD8+ T-cells to CEF peptide-pool stimulation in smokers of both HIV infected and uninfected groups.ConclusionsOur results suggest that smoking and HIV infection independently influence T-cell immune activation and function and together they present the worst immune profile. Since smoking is widespread among HIV infected individuals, studies are warranted to further evaluate the cumulative effect of smoking on impairment of the immune system and accelerated disease progression.

show abstract

Section: Discussionmentioning

confidence: 99%

Tobacco Smoking Increases Immune Activation and Impairs T-Cell Function in HIV Infected Patients on Antiretrovirals: A Cross-Sectional Pilot Study

et al. 2014

View full text Add to dashboard Cite

show abstract

“…The recent availability of MFC has allowed the absolute enumeration of the lymphocyte subpopulation in a single tube, including B lymphocytes or NK cells, at reduced cost and with shorter assay times [18-20]. Determination of lymphocyte subpopulations is very important in many kinds of immunological [20] and hematological disorders [21].…”

Section: Discussionmentioning

confidence: 99%

“…Although some clinical laboratories routinely use a single-tube assay with lyse-no-wash methodology, which reduces inter-laboratory variability, a single-tube assay requires complex analysis with a multiple gating strategy [17-20]. The use of complex instruments with multicolor analysis, in which every fluorochrome has to be accurately compensated for, especially in a lyse-no-wash technique, can be problematic for an inexperienced operator [18].…”

Section: Introductionmentioning

confidence: 99%

Single-color Multitarget Flow Cytometry Using Monoclonal Antibodies Labeled with Different Intensities of the Same Fluorochrome

Park

Han

2012

Ann Lab Med

View full text Add to dashboard Cite

BackgroundWe developed a single-color multitarget flow cytometry (SM-FC) assay, a single-tube assay with graded mean fluorescence intensities (MFIs). We evaluated the repeatability of SM-FC, and its correlation with multicolor flow cytometry (MFC), to assess its application as a routine FC assay.MethodsWe selected CD19, CD3, CD4, and CD8 as antigen targets to analyze a lymphocyte subset. MFIs were graded by adjusting monoclonal antibody (mAb) volumes to detect several cell populations. Dimly labeled mAb was prepared by decreasing mAb volume and the optimum diluted volume was determined by serial dilution. SM-FC repeatability was analyzed 10 times in 2 normal controls. The correlation between SM-FC and MFC was evaluated in 20 normal and 23 patient samples.ResultsCV values (0.8-5.0% and 1.3-4.1% in samples 1 and 2, respectively) acquired by SM-FC with CD3-fluorescein α-isothyocyanate (FITC)dim+CD4-FITCbright and with CD19-FITCdim+CD3-FITCbright showed good repeatability, comparable to that acquired by MFC (1.6-3.7% and 1.0-4.8% in samples 1 and 2, respectively). Excellent correlation was observed between the 2 methods in the 20 normal samples (B cells, T cells, non-Thelper cells, and Thelper cells; r2=0.87, 0.97, 0.97, and 0.98, respectively; P<0.05). There were also linear relationships between SM-FC with CD19-FITCdim+CD3-FITCbright and CD8-PEdim+CD4-PEbright, and MFC, in the 23 patient samples (B cells, T cells, Tcytotoxic cells, and Thelper cells; r2≥0.98, 0.99, 0.99, and 0.99, respectively; P<0.05).ConclusionsThe multicolor, single-tube SM-FC technique is a potential alternative tool for identifying a lymphocyte subset.

show abstract

“…Immunophenotyping is an important diagnostic tool for decision making in various diseases [1][2][3][4][5][6]. Typical examples are: -diagnosis of immunodeficiencies [7], -monitoring of systemic and chronic diseases [8][9][10][11][12], -cellular diagnostics of allergy [13,14], -functional characterization of cells [15,16], -genetic investigation [17,18], -differential diagnosis of lymphocytosis [19], -diagnosis of leukemia [5,[20][21][22][23][24], -investigation of T-cell receptor expression in Kawasaki syndrome [25][26], -investigation of pregnancy disturbances [27], -quality control in hemotherapy [28][29][30].…”

Section: Introductionmentioning

confidence: 99%

Age-Related Lymphocyte Subset Changes in the Peripheral Blood of Healthy Children – a Meta-Study

Sack¹,

Gerling²,

Tárnok³

2007

Transfus Med Hemother

View full text Add to dashboard Cite

Immunophenotyping is an important diagnostic tool for decision making in various diseases. Although clinical interpretation relies on measurable aberrations of the patient’s values from normal ranges, agedependent are rarely available. Materials and Methods: The present study is aimed to combine published data about normal values of peripheral blood lymphocyte subpopulations to describe age-dependent changes from the neonate to the adult. These values could serve in a better way as normal values in comparison to present ones. Furthermore, this investigation allows us to define valid data even for short periods in children’s life and to investigate the influence of technical approaches, sample preparation, antibody selection, measurement equipment, and data analysis. Results: Development-related alterations of lymphocyte subset counts in children could be extracted from the pre-existing papers for diagnostic use. These results were mostly independent from gender, ethnic factors, procedure of sample collection, anticoagulation, pre-analytical procedures, time to workbench, applied method for immunophenotyping, staining procedure, selected monoclonal antibodies, technical devices, and software products. Conclusion: Our data indicate that previous normal values are not sufficiently precise for the interpretation of lymphocyte subsets in children. Mainly during the 1st year of life, count and subset distribution of lymphocytes is different from that of adults. Therefore, a close meshed data set of normal values is required to guarantee adequate diagnostic interpretation.

show abstract

Influence of 4‐ and 6‐color flow cytometers and acquisition/analysis softwares on the determination of lymphocyte subsets in HIV infection

Cited by 13 publications

References 13 publications

Tobacco Smoking Increases Immune Activation and Impairs T-Cell Function in HIV Infected Patients on Antiretrovirals: A Cross-Sectional Pilot Study

Tobacco Smoking Increases Immune Activation and Impairs T-Cell Function in HIV Infected Patients on Antiretrovirals: A Cross-Sectional Pilot Study

Single-color Multitarget Flow Cytometry Using Monoclonal Antibodies Labeled with Different Intensities of the Same Fluorochrome

Age-Related Lymphocyte Subset Changes in the Peripheral Blood of Healthy Children – a Meta-Study

Contact Info

Product

Resources

About