flow cytometry staining analysis  (Bioss)

Journal: Journal of Inflammation Research

Article Title: Multi-Omics Analysis Identifies Immune Regulatory Networks in Sepsis-Associated Liver Injury: Experimental Validation and Clinical Relevance

doi: 10.2147/JIR.S515615

Figure Lengend Snippet: Immune markers were used to detect the type of immune cells in mice with sepsis and liver immune cell typing by single cell RNA sequencing. ( A ) Flow cytometry results of immune T lymphocytes (CD3+). ( B ) Flow cytometry results of immune B lymphocytes (CD19+). ( C ) Flow cytometry results of immune Macrophages (F4/80+). ( D ) Flow cytometry results of immune NK cells (NK1.1+). ( E ) Single-cell sequencing screened out the T-cell-related immune gene FCER1G in liver injury caused by sepsis. ( F ) Single-cell sequencing screened out the T-cell-related immune gene IL2RB in liver injury caused by sepsis. ( G ) Single-cell sequencing screened out the T-cell-related immune gene PTGDR in liver injury caused by sepsis. ( H ) Single-cell sequencing screened out the T-cell-related immune gene XCL1 in liver injury caused by sepsis.

Article Snippet: Flow Cytometry Staining & Analysis:T cells (Bioss Cat# bsm-30149A, PE, 1:150);B cells (Bioss Cat# bsm-30156A, FITC, 1:50);NK cells (Bioss Cat# bsm-30177M, PE, 1:100);Macrophages (Bioss Cat# bsm-30277A, PE, 1:50).

Techniques: RNA Sequencing, Flow Cytometry, Sequencing

Journal: Cells

Article Title: Insulin Modulates NK Cell Activity in Liver Fibrosis MASH Patients via the STING Pathway

doi: 10.3390/cells14130941

Figure Lengend Snippet: NK cell dysfunction is linked to suppressed STING expression in MASH patients with advanced liver fibrosis. The percentages of cells expressing ( a ) CD107a, ( b ) NKp46, ( c ) LAIR-1, and ( d ) Siglec7 in NK from healthy controls and metabolic dysfunction-associated steatohepatitis (MASH) patients with early (F1/F2) and advanced (F3/F4) fibrosis scores as determined by METAVIR were measured by flow cytometry. ( e ) Quantitation of STING mRNA in peripheral NK cells obtained from healthy individuals (considered as control) and patients with MASH with varying degrees of fibrosis scoring were assessed via RT-PCR. The data are presented as a fold change compared with healthy controls. The housekeeping gene GAPDH was used as the internal control for normalization. Flow cytometry analysis showed the percentages of cells expressing ( f ) STING and ( g ) IFN-γ in NK from healthy controls and MASH patients with early (F1/F2) and advanced (F3/F4) fibrosis. ( h ) A coculture cytotoxicity assay using a 1:1 ratio of NK cells (from healthy donors and MASH patients with early and advanced fibrosis scores) and pHSCs obtained from F3/F4 MASH patients was performed, and annexin V (%) expression was assessed in the pHSCs as a marker of apoptosis using flow cytometry. The horizontal dotted line indicates the baseline results from monoculture conditions. In the flow cytometry experiments, an isotype-matched IgG antibody conjugated to the appropriate fluorochrome was used as a control. Data are expressed as the mean ± standard deviation (SD). Statistical significance was assessed using Newman–Keuls two-way analysis of variance (ANOVA), with significance levels denoted as follows: * p < 0.01, ** p < 0.001, *** p < 0.0001, and **** p < 0.00001. ns = not significant. Group sizes were as follows: healthy controls ( n = 10), early liver fibrosis ( n = 10), and advanced liver fibrosis ( n = 10).

Article Snippet: Prior to flow cytometry analysis, the viability of pHSCs and NK cells (100 μL at 1 × 10 6 cells/mL) was assessed using propidium iodide (PI) staining, following the manufacturer’s protocol (A35110, R&D Systems, Minneapolis, MN, USA).

Techniques: Expressing, Flow Cytometry, Quantitation Assay, Control, Reverse Transcription Polymerase Chain Reaction, Cytotoxicity Assay, Marker, Standard Deviation

Journal: Cells

Article Title: Insulin Modulates NK Cell Activity in Liver Fibrosis MASH Patients via the STING Pathway

doi: 10.3390/cells14130941

Figure Lengend Snippet: STING signaling orchestrates NK Cell impairment along liver fibrosis progression in MASH. Following isolation, NK cells from healthy individuals and MASH patients with early (F1/F2) and advanced (F3/F4) stages of liver fibrosis were seeded at a concentration of 1 × 10 6 cells/mL. The cells were then treated for 24 h with STING agonists 5,6-dimethylxanthenone-4-acetic acid (DMXAA; D5817, MERCK) or 2′3′-cyclic GMP-AMP (2′3′-cGAMP; tlrl-nacga23-02, InvivoGen) each at a final concentration of 10 µM. DMSO was used as the vehicle control. The expression levels of ( a ) CD107a, ( b ) NKp46, and ( c ) LAIR-1 on NK cells were assessed by flow cytometry. Isotype-matched IgG antibodies conjugated with appropriate fluorochromes were used as controls in the flow cytometry assays. Data are presented as the mean ± standard deviation (SD). Statistical significance was determined using Newman–Keuls two-way ANOVA with the following notations: * p < 0.01, ** p < 0.001, *** p < 0.0001, and **** p < 0.00001. ns = not significant. Group sizes included healthy controls ( n = 5), patients with early fibrosis ( n = 5), and patients with advanced fibrosis ( n = 5).

Article Snippet: Prior to flow cytometry analysis, the viability of pHSCs and NK cells (100 μL at 1 × 10 6 cells/mL) was assessed using propidium iodide (PI) staining, following the manufacturer’s protocol (A35110, R&D Systems, Minneapolis, MN, USA).

Techniques: Isolation, Concentration Assay, Control, Expressing, Flow Cytometry, Standard Deviation

Journal: Cells

Article Title: Insulin Modulates NK Cell Activity in Liver Fibrosis MASH Patients via the STING Pathway

doi: 10.3390/cells14130941

Figure Lengend Snippet: Insulin signaling enhances STING expression and restores NK Cell function in MASH patients. Following isolation, NK cells from healthy controls and MASH patients with early (F1/F2) and advanced (F3/F4) fibrosis scores were seeded at 1 × 10 6 cells/mL and treated with the STING agonists 2′3′-cyclic GMP-AMP (2′3′-cGAMP; tlrl-nacga23-02, InvivoGen, California, USA) at a final concentration of 10 µM or recombinant human insulin (91077C, MERCK, Jerusalem, Israel), or both for 24 h. DMSO is used as a vehicle. The percentages of cells expressing ( a ) insulin receptor (IR), ( b ) STING, ( c ) CD107a, and ( d ) PD-1in NK cells were measured by flow cytometry. In the flow cytometry experiments, an isotype IgG antibody labeled with the relevant fluorochrome was used as a control. The data are presented as the average ± SD. Significance was determined using Newman–Keuls two-way analysis of variance (ANOVA), * p < 0.01, ** p < 0.001, *** p < 0.0001, and **** p < 0.00001. ns = not significant. Healthy ( n = 5), early liver fibrosis ( n = 5), and advanced liver fibrosis ( n = 5).

Article Snippet: Prior to flow cytometry analysis, the viability of pHSCs and NK cells (100 μL at 1 × 10 6 cells/mL) was assessed using propidium iodide (PI) staining, following the manufacturer’s protocol (A35110, R&D Systems, Minneapolis, MN, USA).

Techniques: Expressing, Cell Function Assay, Isolation, Concentration Assay, Recombinant, Flow Cytometry, Labeling, Control

Journal: Cancer Immunology, Immunotherapy : CII

Article Title: Prostaglandin F2 receptor negative regulator as a potential target for chimeric antigen receptor-T cell therapy for glioblastoma

doi: 10.1007/s00262-025-03979-4

Figure Lengend Snippet: Identification of 5E17 as a GBM-specific mAb. A Strategy for the identification of the GBM-specific mAb 5E17. B Representative flow cytometry findings of 5E17 mAb bound to GBM and nonmalignant brain cells from patients. Analyses of live (PI-negative) cells are shown. The results of staining with the isotype or negative control instead of anti-5E17 mAb, CD31, CD45, or CD90 are shown to draw the gate for 5E17-positive cells. Analyses of the other patients are shown in Supplementary Fig. 2. Blue histogram indicates isotype control. SSC, side scatter. C Mean fluorescence intensity (means ± standard error of the mean) of flow cytometry findings bound to GBM ( n = 7) and nonmalignant brain cells ( n = 6) from patients. Each plot shows the mean fluorescence intensity. GBM cells were assessed as CD31-CD45-CD90 + cells, and nonmalignant tumor cells were assessed as CD31-CD45- cells. * p < 0.05, calculated using a Mann–Whitney U -test

Article Snippet: C Flow cytometry analysis of 5E17 (top row) and commercially available anti-PTGFRN antibody (R&D systems, inc.) (bottom row) reactivity to wild/5E17 overexpressed Ba/F3 cells and wild/PTGFRN KO U87MG cells.

Techniques: Flow Cytometry, Staining, Negative Control, Control, Fluorescence, MANN-WHITNEY

Journal: Cancer Immunology, Immunotherapy : CII

Article Title: Prostaglandin F2 receptor negative regulator as a potential target for chimeric antigen receptor-T cell therapy for glioblastoma

doi: 10.1007/s00262-025-03979-4

Figure Lengend Snippet: 5E17 recognized PTGFRN. A Procedure for expression cloning to identify antigens recognized by 5E17 monoclonal antibody. B Flow cytometry plots showing the process of 5E17-positive cell enrichment in the expression cloning of the 5E17 antigen. SSC, side scatter. C Flow cytometry analysis of 5E17 (top row) and commercially available anti-PTGFRN antibody (R&D systems, inc.) (bottom row) reactivity to wild/5E17 overexpressed Ba/F3 cells and wild/PTGFRN KO U87MG cells. WT, wild type; KO, knock out

Article Snippet: C Flow cytometry analysis of 5E17 (top row) and commercially available anti-PTGFRN antibody (R&D systems, inc.) (bottom row) reactivity to wild/5E17 overexpressed Ba/F3 cells and wild/PTGFRN KO U87MG cells.

Techniques: Expressing, Cloning, Flow Cytometry, Knock-Out

Journal: Cancer Immunology, Immunotherapy : CII

Article Title: Prostaglandin F2 receptor negative regulator as a potential target for chimeric antigen receptor-T cell therapy for glioblastoma

doi: 10.1007/s00262-025-03979-4

Figure Lengend Snippet: Development of CAR-T cells for the 5E17 antigen. A Establishment of CAR-T cells using the 5E17 monoclonal antibody in the indicated construct. VH, variable heavy; VL, variable light. B Flow cytometry plots of 5E17 CAR or Mock transduction efficiency in human T cells 10 days after CAR transduction. SSC, side scatter. C Growth of 5E17 CAR-T or Mock T cells during in vitro culture

Article Snippet: C Flow cytometry analysis of 5E17 (top row) and commercially available anti-PTGFRN antibody (R&D systems, inc.) (bottom row) reactivity to wild/5E17 overexpressed Ba/F3 cells and wild/PTGFRN KO U87MG cells.

Techniques: Construct, Flow Cytometry, Transduction, In Vitro