interleukin 2 il 2 dependent natural killer cell cell line nk 92mi  (ATCC)


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    ATCC interleukin 2 il 2 dependent natural killer cell cell line nk 92mi
    Effects on <t>NK</t> <t>cell</t> proliferation of B-cell supernatants containing 0.5 mg/mL extracts.
    Interleukin 2 Il 2 Dependent Natural Killer Cell Cell Line Nk 92mi, supplied by ATCC, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    1) Product Images from "Enhanced Immunomodulatory Activity of Gelatin-Encapsulated Rubus coreanus Miquel Nanoparticles"

    Article Title: Enhanced Immunomodulatory Activity of Gelatin-Encapsulated Rubus coreanus Miquel Nanoparticles

    Journal: International Journal of Molecular Sciences

    doi: 10.3390/ijms12129031

    Effects on NK cell proliferation of B-cell supernatants containing 0.5 mg/mL extracts.
    Figure Legend Snippet: Effects on NK cell proliferation of B-cell supernatants containing 0.5 mg/mL extracts.

    Techniques Used:

    natural killer cell line nk 92  (ATCC)


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    ATCC natural killer cell line nk 92
    Natural Killer Cell Line Nk 92, supplied by ATCC, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    natural killer 92 nk 92 cell line  (ATCC)


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    ATCC natural killer 92 nk 92 cell line
    Generation of <t>NK-92-CAR.19-IL-15/IL15Rα</t> cells. (A) Scheme of CAR constructs targeting CD19 (CAR.19) under the control of spleen focus-forming virus (SFFV) promoter. CAR consists of scFv, CD8 hinge and transmembrane region, 41BB costimulatory molecule and CD3ζ signaling molecule. Where indicated, the CAR sequences are followed by a self-cleaving peptide (T2A) and IL-15 or IL-15/IL-15Rα (IL-15 and IL-15Rα fused through a flexible linker). (B, C) NK-92 cells were transduced with the described CAR constructs followed by immunomagnetic enrichment in two steps. (B) Representative dot plots show flow cytometric analysis of CAR expression on freshly transduced and enriched NK-92-CAR.19 cells. (C) The graph shows the frequency of CAR-positive cells before selection, after first and second selection, and 30 days after second selection.
    Natural Killer 92 Nk 92 Cell Line, supplied by ATCC, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    1) Product Images from "Engineering NK-CAR.19 cells with the IL-15/IL-15Rα complex improved proliferation and anti-tumor effect in vivo"

    Article Title: Engineering NK-CAR.19 cells with the IL-15/IL-15Rα complex improved proliferation and anti-tumor effect in vivo

    Journal: Frontiers in Immunology

    doi: 10.3389/fimmu.2023.1226518

    Generation of NK-92-CAR.19-IL-15/IL15Rα cells. (A) Scheme of CAR constructs targeting CD19 (CAR.19) under the control of spleen focus-forming virus (SFFV) promoter. CAR consists of scFv, CD8 hinge and transmembrane region, 41BB costimulatory molecule and CD3ζ signaling molecule. Where indicated, the CAR sequences are followed by a self-cleaving peptide (T2A) and IL-15 or IL-15/IL-15Rα (IL-15 and IL-15Rα fused through a flexible linker). (B, C) NK-92 cells were transduced with the described CAR constructs followed by immunomagnetic enrichment in two steps. (B) Representative dot plots show flow cytometric analysis of CAR expression on freshly transduced and enriched NK-92-CAR.19 cells. (C) The graph shows the frequency of CAR-positive cells before selection, after first and second selection, and 30 days after second selection.
    Figure Legend Snippet: Generation of NK-92-CAR.19-IL-15/IL15Rα cells. (A) Scheme of CAR constructs targeting CD19 (CAR.19) under the control of spleen focus-forming virus (SFFV) promoter. CAR consists of scFv, CD8 hinge and transmembrane region, 41BB costimulatory molecule and CD3ζ signaling molecule. Where indicated, the CAR sequences are followed by a self-cleaving peptide (T2A) and IL-15 or IL-15/IL-15Rα (IL-15 and IL-15Rα fused through a flexible linker). (B, C) NK-92 cells were transduced with the described CAR constructs followed by immunomagnetic enrichment in two steps. (B) Representative dot plots show flow cytometric analysis of CAR expression on freshly transduced and enriched NK-92-CAR.19 cells. (C) The graph shows the frequency of CAR-positive cells before selection, after first and second selection, and 30 days after second selection.

    Techniques Used: Construct, Virus, Transduction, Expressing, Selection

    NK-92-CAR.19-IL-15/IL15Rα cells proliferate independently of exogenous IL-2. Proliferation of parental and genetically modified NK-92 cells: NK-92-CAR.19, NK-92-CAR.19-IL-15 and NK-92-CAR.19-IL-15/IL15Rα in X-Vivo 10 supplemented with 5% of human plasma. (A) Cells were cultured either in presence of IL-2 (500 UI/mL) or (B) in absence of exogenous cytokines. (C) Viability of cell culture with cytokines and (D) without cytokines. Cell culture was followed over 21 days. Pooled data of two independent experiments (each one performed in duplicates) are shown. (E) Quantification of soluble IL-15 in supernatant of NK cell culture by Luminex (n=2). Two-way ANOVA statistical test, Tukey's post-multiple comparison posttest. ****P<0.0001.
    Figure Legend Snippet: NK-92-CAR.19-IL-15/IL15Rα cells proliferate independently of exogenous IL-2. Proliferation of parental and genetically modified NK-92 cells: NK-92-CAR.19, NK-92-CAR.19-IL-15 and NK-92-CAR.19-IL-15/IL15Rα in X-Vivo 10 supplemented with 5% of human plasma. (A) Cells were cultured either in presence of IL-2 (500 UI/mL) or (B) in absence of exogenous cytokines. (C) Viability of cell culture with cytokines and (D) without cytokines. Cell culture was followed over 21 days. Pooled data of two independent experiments (each one performed in duplicates) are shown. (E) Quantification of soluble IL-15 in supernatant of NK cell culture by Luminex (n=2). Two-way ANOVA statistical test, Tukey's post-multiple comparison posttest. ****P<0.0001.

    Techniques Used: Genetically Modified, Cell Culture, Luminex, Comparison

    NK-92-CAR.19 cells coexpressing IL-15/Rα enhance CAR-mediated anti-tumor Ncell function. NK-92, NK-92-CAR.19, NK-92-CAR.19-IL-15 and NK-92-CAR.19-IL-15/IL-15Rα cells were co-cultivated with (A) Raji (B) NALM-6 and (C) K562 in 2:1 and 10:1 effector:target cells ratio, for 5 hours. Cytotoxicity was measured by flow cytometry cytotoxicity assay. One-way ANOVA statistical test, Tukey's multiple comparison posttest. ****P < 0.0001; ***P < 0.001; **P < 0.01; *P < 0.05. Mean values + SD are shown. Data pooled from three independent experiments.
    Figure Legend Snippet: NK-92-CAR.19 cells coexpressing IL-15/Rα enhance CAR-mediated anti-tumor Ncell function. NK-92, NK-92-CAR.19, NK-92-CAR.19-IL-15 and NK-92-CAR.19-IL-15/IL-15Rα cells were co-cultivated with (A) Raji (B) NALM-6 and (C) K562 in 2:1 and 10:1 effector:target cells ratio, for 5 hours. Cytotoxicity was measured by flow cytometry cytotoxicity assay. One-way ANOVA statistical test, Tukey's multiple comparison posttest. ****P < 0.0001; ***P < 0.001; **P < 0.01; *P < 0.05. Mean values + SD are shown. Data pooled from three independent experiments.

    Techniques Used: Flow Cytometry, Cytotoxicity Assay, Comparison

    NK-92-CAR.19-IL-15/IL15Rα cells produce high amount of anti-tumor pro-inflammatory cytokines. NK-92, CAR.19, CAR.19-IL-15 and CAR.19-IL-15Rα NK-92 cells (2.5 × 10 5 ) were incubated for 5 h with Raji lymphoma cells (A, C) or with Nalm-6 (B, D) at an E/T ratio of 10:1. Supernatants were collected, and the levels of IFN-γ (A, B) and TNF-α (C, D) , granzyme A (E, F) ), granzyme B (G, H) and perforin (I, J) were measured using the Luminex MAGPIX system. NK cells without target cells were included as controls. One-way ANOVA statistical test, Tukey's multiple comparison posttest. Mean values ± SD are shown; n=2. ***P < 0.001; **P < 0.01; *P < 0.05. Data pooled from three independent experiments.
    Figure Legend Snippet: NK-92-CAR.19-IL-15/IL15Rα cells produce high amount of anti-tumor pro-inflammatory cytokines. NK-92, CAR.19, CAR.19-IL-15 and CAR.19-IL-15Rα NK-92 cells (2.5 × 10 5 ) were incubated for 5 h with Raji lymphoma cells (A, C) or with Nalm-6 (B, D) at an E/T ratio of 10:1. Supernatants were collected, and the levels of IFN-γ (A, B) and TNF-α (C, D) , granzyme A (E, F) ), granzyme B (G, H) and perforin (I, J) were measured using the Luminex MAGPIX system. NK cells without target cells were included as controls. One-way ANOVA statistical test, Tukey's multiple comparison posttest. Mean values ± SD are shown; n=2. ***P < 0.001; **P < 0.01; *P < 0.05. Data pooled from three independent experiments.

    Techniques Used: Incubation, Luminex, Comparison

    Metascape functional analysis of transcriptome profiles of NK-92-CAR.19 cells after coculture with a target B cell line. (A) The Circos plot shows how genes from the different input gene lists after coculture setups overlap. On the outside, the arc represents the identity of each gene list. On the inside, the orange color represents the genes that appear in multiple lists, and the light orange color represents genes that are unique to that gene list. Purple lines link the same genes shared by multiple gene lists. Blue lines link the genes that fall into the same ontology term (the term has to be statistically significantly enriched). The greater the number of purple links and the longer the dark orange arcs, the greater is the overlap among the input gene lists. Blue links indicate the amount of functional overlap among the input gene lists. (B) Networks of Protein-Protein interaction (PPI) identified in the gene lists for comparing across NK92-CAR groups and NK92-WT, and illustrated using Cytoscape 3.9.1. (C) Summary of enrichment analysis done using the DEG_up from (A) and the Trancription_Factor_Targets as an ontology category (Metascape).
    Figure Legend Snippet: Metascape functional analysis of transcriptome profiles of NK-92-CAR.19 cells after coculture with a target B cell line. (A) The Circos plot shows how genes from the different input gene lists after coculture setups overlap. On the outside, the arc represents the identity of each gene list. On the inside, the orange color represents the genes that appear in multiple lists, and the light orange color represents genes that are unique to that gene list. Purple lines link the same genes shared by multiple gene lists. Blue lines link the genes that fall into the same ontology term (the term has to be statistically significantly enriched). The greater the number of purple links and the longer the dark orange arcs, the greater is the overlap among the input gene lists. Blue links indicate the amount of functional overlap among the input gene lists. (B) Networks of Protein-Protein interaction (PPI) identified in the gene lists for comparing across NK92-CAR groups and NK92-WT, and illustrated using Cytoscape 3.9.1. (C) Summary of enrichment analysis done using the DEG_up from (A) and the Trancription_Factor_Targets as an ontology category (Metascape).

    Techniques Used: Functional Assay

    Transgenic IL-15 or IL-15/Rα maintains low expression of immune checkpoint molecules in NK-92-CAR.19 cell variants. NK-92-CAR.19 cells were cultured unstimulated (-) or stimulated (+) with Raji cells at an E:T ratio of 1:2, followed by restimulation with fresh target cells after 24 and 48 hours. Three days after the beginning of the experiment, the surface expression of LAG-3 (A) , PD-1 (B) and TIM-3 (C) were analyzed by flow cytometry. Three independent experiments were performed. One-way ANOVA statistical test, Tukey's post-hoc multiple comparison. Mean values + SD are shown. Gray: NK-92, Black: coculture of NK-92 and Raji cells, Orange: CAR.19, Brown: coculture of CAR.19 and Raji cells, Pink: CAR.19-IL-15, Red: cocultured of CAR.19-IL-15 and Raji cells, Light blue: CAR.19-IL-15RA, and Blue: coculture of CAR.19-IL-15 and Raji cells. (D) Representative flow cytometry histograms plots showing the mean fluorescence intensity of LAG-3, PD-1 and TIM-3 in NK-92 cell and, in the NK-92-CAR.19 cell variants. ***P < 0.001; **P < 0.01; *P < 0.05.
    Figure Legend Snippet: Transgenic IL-15 or IL-15/Rα maintains low expression of immune checkpoint molecules in NK-92-CAR.19 cell variants. NK-92-CAR.19 cells were cultured unstimulated (-) or stimulated (+) with Raji cells at an E:T ratio of 1:2, followed by restimulation with fresh target cells after 24 and 48 hours. Three days after the beginning of the experiment, the surface expression of LAG-3 (A) , PD-1 (B) and TIM-3 (C) were analyzed by flow cytometry. Three independent experiments were performed. One-way ANOVA statistical test, Tukey's post-hoc multiple comparison. Mean values + SD are shown. Gray: NK-92, Black: coculture of NK-92 and Raji cells, Orange: CAR.19, Brown: coculture of CAR.19 and Raji cells, Pink: CAR.19-IL-15, Red: cocultured of CAR.19-IL-15 and Raji cells, Light blue: CAR.19-IL-15RA, and Blue: coculture of CAR.19-IL-15 and Raji cells. (D) Representative flow cytometry histograms plots showing the mean fluorescence intensity of LAG-3, PD-1 and TIM-3 in NK-92 cell and, in the NK-92-CAR.19 cell variants. ***P < 0.001; **P < 0.01; *P < 0.05.

    Techniques Used: Transgenic Assay, Expressing, Cell Culture, Flow Cytometry, Comparison, Fluorescence

    NK-92-CAR.19-IL15/IL-15Rα cells eliminate lymphoma B-cells and reduce tumor burden in vivo . (A) Scheme of an in vivo study demonstrating the anti-tumor activity of transduced NK-92-CAR.19 cells in a xenogeneic NSG mouse model of disseminated human B-cell hematologic malignancy. NSG mice were intravenously injected with 2 × 10 4 Raji-Luc lymphoma B-cells. On days 4, 7, 10, 12, 15, and 19, animals were treated by intravenous injection of 7 × 10 6 NK-92 cells (n=5). (B) Lymphoma development was monitored by in vivo bioluminescence imaging. Images were taken on days 8, 14, 21, 26, and 29. (C) Bioluminescence intensity was quantified and means + SEM are shown. (D) Bioluminescence of individual mice shown on day 29. Means + SEM are shown. Statistics were determined by the Mann-Whitney test. Each experimental group consisted of five mice. **P < 0.01; *P < 0.05.
    Figure Legend Snippet: NK-92-CAR.19-IL15/IL-15Rα cells eliminate lymphoma B-cells and reduce tumor burden in vivo . (A) Scheme of an in vivo study demonstrating the anti-tumor activity of transduced NK-92-CAR.19 cells in a xenogeneic NSG mouse model of disseminated human B-cell hematologic malignancy. NSG mice were intravenously injected with 2 × 10 4 Raji-Luc lymphoma B-cells. On days 4, 7, 10, 12, 15, and 19, animals were treated by intravenous injection of 7 × 10 6 NK-92 cells (n=5). (B) Lymphoma development was monitored by in vivo bioluminescence imaging. Images were taken on days 8, 14, 21, 26, and 29. (C) Bioluminescence intensity was quantified and means + SEM are shown. (D) Bioluminescence of individual mice shown on day 29. Means + SEM are shown. Statistics were determined by the Mann-Whitney test. Each experimental group consisted of five mice. **P < 0.01; *P < 0.05.

    Techniques Used: In Vivo, Activity Assay, Injection, Imaging, MANN-WHITNEY

    Proposed model of action of engineered NK-92-CAR-19-IL-15/IL-15Rα cells. In the absence of exogenous IL-2: (A) NK-92-CAR.19 cells remain non-proliferative. (B) NK-92-CAR.19-IL-15 cells exhibit modest proliferation up to day 12, as they release soluble IL-15 that subsequently engages with IL-2/IL-15Rβ–γc receptors on their own membrane or neighboring NK cells. (C) NK-92-CAR.19-IL-15/IL-15Rα cells display robust proliferation, driven by the presence of IL-15/IL-15Rα complexes on their cell membrane. This activates the cells through cis-presentation or may stimulate adjacent NK cells expressing membrane-bound IL-2/IL-15Rβ–γc receptors via trans-presentation. The signaling from IL-15 prompts all three NK-92-CAR-19 cell variants to upregulate the PI3K/AKT and JAK/STAT pathways. However, this upregulation is notably more pronounced in NK-92-CAR.19-IL-15/IL-15Rα cells compared to the other variants. Consequently, this substantial enhancement in signaling leads to heightened proliferative capacity, increased secretion of proinflammatory cytokines, and exceptionally potent cytotoxic activity in these pioneering engineered NK-92 cells.
    Figure Legend Snippet: Proposed model of action of engineered NK-92-CAR-19-IL-15/IL-15Rα cells. In the absence of exogenous IL-2: (A) NK-92-CAR.19 cells remain non-proliferative. (B) NK-92-CAR.19-IL-15 cells exhibit modest proliferation up to day 12, as they release soluble IL-15 that subsequently engages with IL-2/IL-15Rβ–γc receptors on their own membrane or neighboring NK cells. (C) NK-92-CAR.19-IL-15/IL-15Rα cells display robust proliferation, driven by the presence of IL-15/IL-15Rα complexes on their cell membrane. This activates the cells through cis-presentation or may stimulate adjacent NK cells expressing membrane-bound IL-2/IL-15Rβ–γc receptors via trans-presentation. The signaling from IL-15 prompts all three NK-92-CAR-19 cell variants to upregulate the PI3K/AKT and JAK/STAT pathways. However, this upregulation is notably more pronounced in NK-92-CAR.19-IL-15/IL-15Rα cells compared to the other variants. Consequently, this substantial enhancement in signaling leads to heightened proliferative capacity, increased secretion of proinflammatory cytokines, and exceptionally potent cytotoxic activity in these pioneering engineered NK-92 cells.

    Techniques Used: Membrane, Expressing, Activity Assay

    natural killer nk like cell line yt  (ATCC)


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    ATCC natural killer nk like cell line yt
    Natural Killer Nk Like Cell Line Yt, supplied by ATCC, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    natural killer nk cell line nk  (ATCC)


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    ATCC natural killer nk cell line nk
    Natural Killer Nk Cell Line Nk, supplied by ATCC, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    natural killer nk cell line nk  (ATCC)


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    ATCC natural killer nk cell line nk
    The quantitation of TGF-β1 ( A ), IL-1β ( B ), IL-18 ( C ), and IL-21 ( D ) released from the co-culture of HCC SK-Hep1 and <t>NK-92</t> with or without CoCl 2 (250 μM) and IL-6 antibody (2 ng/ml) by ELISA assay. * p < 0.05 is versus non-treated; ** p < 0.05 is versus CoCl 2 -treated. All data are presented as the means ± SD in three independent experiments.
    Natural Killer Nk Cell Line Nk, supplied by ATCC, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/natural killer nk cell line nk/product/ATCC
    Average 86 stars, based on 1 article reviews
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    1) Product Images from "Role of Interleukin(IL)-6 in NK Activity to Hypoxic-Induced Highly Invasive Hepatocellular Carcinoma(HCC) Cells"

    Article Title: Role of Interleukin(IL)-6 in NK Activity to Hypoxic-Induced Highly Invasive Hepatocellular Carcinoma(HCC) Cells

    Journal: Journal of Microbiology and Biotechnology

    doi: 10.4014/jmb.2304.04023

    The quantitation of TGF-β1 ( A ), IL-1β ( B ), IL-18 ( C ), and IL-21 ( D ) released from the co-culture of HCC SK-Hep1 and NK-92 with or without CoCl 2 (250 μM) and IL-6 antibody (2 ng/ml) by ELISA assay. * p < 0.05 is versus non-treated; ** p < 0.05 is versus CoCl 2 -treated. All data are presented as the means ± SD in three independent experiments.
    Figure Legend Snippet: The quantitation of TGF-β1 ( A ), IL-1β ( B ), IL-18 ( C ), and IL-21 ( D ) released from the co-culture of HCC SK-Hep1 and NK-92 with or without CoCl 2 (250 μM) and IL-6 antibody (2 ng/ml) by ELISA assay. * p < 0.05 is versus non-treated; ** p < 0.05 is versus CoCl 2 -treated. All data are presented as the means ± SD in three independent experiments.

    Techniques Used: Quantitation Assay, Co-Culture Assay, Enzyme-linked Immunosorbent Assay

    ( A ) Cytotoxic ability of NK-92 co-cultured with HCC SK-Hep1 with or without the treatment of CoCl 2 (250 μM) or IL-6 antibody by LDH-release assay. ( B, D ) Granules (Grz-B and perforin) in NK-92 cells cocultured with HCC SK-Hep1 treated with or without CoCl 2 (250 μM) or IL-6 antibody. The expression of Grz-B and perforin was measured by Western blot analysis and intracellular staining with fluorescent perforin antibody, respectively. The expression of mRNA of ( C ) grz-B and ( E ) perforin in NK-92 co-cultured with HCC SK-Hep1 is analyzed using RT-qPCR. All experiments were conducted in three independent as means ± SD. * p < 0.05 vs. non-treated; ** p < 0.05 vs. CoCl 2 .
    Figure Legend Snippet: ( A ) Cytotoxic ability of NK-92 co-cultured with HCC SK-Hep1 with or without the treatment of CoCl 2 (250 μM) or IL-6 antibody by LDH-release assay. ( B, D ) Granules (Grz-B and perforin) in NK-92 cells cocultured with HCC SK-Hep1 treated with or without CoCl 2 (250 μM) or IL-6 antibody. The expression of Grz-B and perforin was measured by Western blot analysis and intracellular staining with fluorescent perforin antibody, respectively. The expression of mRNA of ( C ) grz-B and ( E ) perforin in NK-92 co-cultured with HCC SK-Hep1 is analyzed using RT-qPCR. All experiments were conducted in three independent as means ± SD. * p < 0.05 vs. non-treated; ** p < 0.05 vs. CoCl 2 .

    Techniques Used: Cell Culture, Lactate Dehydrogenase Assay, Expressing, Western Blot, Staining, Quantitative RT-PCR

    Target cells were incubated in a 6-well plate overnight and then co-cultured with effector cells (NK-92) at a ratio of 2:1 (E:T) for 24 h with or without CoCl 2 (250 μM) or IL-6 antibody (2 ng/ml). After removing the effector cells, target cells were harvested and lysed using protein extraction buffer. The extracted proteins were analyzed using Western blot analysis. The expression of ( A ) (cleaved or) caspase-3, ( B ) (cleaved or) caspase-7, ( C ) (cleaved or) caspase-9, and ( D ) (cleaved or) caspase-8 in HCC SK-Hep1 co-cultured with NK-92. *,** p < 0.05. The quantitation of bands was analyzed in three independent experiments.
    Figure Legend Snippet: Target cells were incubated in a 6-well plate overnight and then co-cultured with effector cells (NK-92) at a ratio of 2:1 (E:T) for 24 h with or without CoCl 2 (250 μM) or IL-6 antibody (2 ng/ml). After removing the effector cells, target cells were harvested and lysed using protein extraction buffer. The extracted proteins were analyzed using Western blot analysis. The expression of ( A ) (cleaved or) caspase-3, ( B ) (cleaved or) caspase-7, ( C ) (cleaved or) caspase-9, and ( D ) (cleaved or) caspase-8 in HCC SK-Hep1 co-cultured with NK-92. *,** p < 0.05. The quantitation of bands was analyzed in three independent experiments.

    Techniques Used: Incubation, Cell Culture, Protein Extraction, Western Blot, Expressing, Quantitation Assay

    The expression of HIF-1α and (p)Stat3 was analyzed using Western blot analysis. The expression of ( A, B ) HIF-1α and ( C, D ) (p)Stat3 in HCC SK-Hep1 cells co-cultured with NK-92 cells with or without the treatment of CoCl 2 (250 μM) or IL-6 antibody (2 ng/ml), and hrIL-21. *,** p < 0.05. All data were presented as the means ± SD in three independent experiments.
    Figure Legend Snippet: The expression of HIF-1α and (p)Stat3 was analyzed using Western blot analysis. The expression of ( A, B ) HIF-1α and ( C, D ) (p)Stat3 in HCC SK-Hep1 cells co-cultured with NK-92 cells with or without the treatment of CoCl 2 (250 μM) or IL-6 antibody (2 ng/ml), and hrIL-21. *,** p < 0.05. All data were presented as the means ± SD in three independent experiments.

    Techniques Used: Expressing, Western Blot, Cell Culture

    natural killer nk like cell line yt  (ATCC)


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    ATCC natural killer nk like cell line yt
    Natural Killer Nk Like Cell Line Yt, supplied by ATCC, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    human natural killer cell line nk  (ATCC)


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    ATCC human natural killer cell line nk
    Human Natural Killer Cell Line Nk, supplied by ATCC, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    human natural killer cell line nk  (ATCC)


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    ATCC human natural killer cell line nk
    Influence of GOS on <t>NK-92</t> cell viability. Cells were exposed to culture medium alone (control) or GOS at concentrations of 10, 50, 100, and 500 µg/mL for 48 h. Cell viability was determined by examination of metabolic activity using the MTT assay. Results are presented as mean ± SEM of at least six measurements. Statistically significant differences compared to the control at p < 0.05 (*), p < 0.001 (***). One-way ANOVA test; Dunnett’s post-hoc test.
    Human Natural Killer Cell Line Nk, supplied by ATCC, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Average 86 stars, based on 1 article reviews
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    1) Product Images from "(1→3)-α- d -Glucooligosaccharides Increase the Killing Capacity of NK Cells against Selected Human Colon Cancer Cells"

    Article Title: (1→3)-α- d -Glucooligosaccharides Increase the Killing Capacity of NK Cells against Selected Human Colon Cancer Cells

    Journal: Molecules

    doi: 10.3390/molecules28104212

    Influence of GOS on NK-92 cell viability. Cells were exposed to culture medium alone (control) or GOS at concentrations of 10, 50, 100, and 500 µg/mL for 48 h. Cell viability was determined by examination of metabolic activity using the MTT assay. Results are presented as mean ± SEM of at least six measurements. Statistically significant differences compared to the control at p < 0.05 (*), p < 0.001 (***). One-way ANOVA test; Dunnett’s post-hoc test.
    Figure Legend Snippet: Influence of GOS on NK-92 cell viability. Cells were exposed to culture medium alone (control) or GOS at concentrations of 10, 50, 100, and 500 µg/mL for 48 h. Cell viability was determined by examination of metabolic activity using the MTT assay. Results are presented as mean ± SEM of at least six measurements. Statistically significant differences compared to the control at p < 0.05 (*), p < 0.001 (***). One-way ANOVA test; Dunnett’s post-hoc test.

    Techniques Used: Activity Assay, MTT Assay

    Influence of GOS on NK-92 cell cytotoxicity against human adenocarcinoma cell lines HT-29 and LS180. Cancer cells were exposed to culture medium alone (control) or GOS (10, 50, 100, and 500 µg/mL) used alone or NK-92 cells in the absence or presence of GOS for 48 h. NK-92 cells were used in the same or twofold higher concentration as the treated cancer cells. The viability of the investigated colon cancer cells in response to GOS used alone or together with the NK cells was determined by examination of cell metabolic activity using the MTT assay. Results are presented as mean ± SEM of at least five measurements. Statistically significant differences compared to the control at p < 0.001 (***). Statistically significant differences between cancer cells treated with GOS-activated NK-92 cells vs. cancer cells treated with only NK cells at p < 0.01 (^^), p < 0.001 (^^^). Statistically significant differences between cancer cells treated with GOS-activated NK-92 cells vs. cancer cells exposed to GOS at a corresponding concentration as that used for NK cell activation at p < 0.001 (’’’). One-way ANOVA test; Tukey’s post-hoc test. Cancer cell death in response to GOS as well as NK-92 cells activated by the tested compound was visualized by cell double staining with a mixture of propidium iodide and Hoechst 33342. The representative pictures from fluorescence microscopy are presented; the magnification is 200×.
    Figure Legend Snippet: Influence of GOS on NK-92 cell cytotoxicity against human adenocarcinoma cell lines HT-29 and LS180. Cancer cells were exposed to culture medium alone (control) or GOS (10, 50, 100, and 500 µg/mL) used alone or NK-92 cells in the absence or presence of GOS for 48 h. NK-92 cells were used in the same or twofold higher concentration as the treated cancer cells. The viability of the investigated colon cancer cells in response to GOS used alone or together with the NK cells was determined by examination of cell metabolic activity using the MTT assay. Results are presented as mean ± SEM of at least five measurements. Statistically significant differences compared to the control at p < 0.001 (***). Statistically significant differences between cancer cells treated with GOS-activated NK-92 cells vs. cancer cells treated with only NK cells at p < 0.01 (^^), p < 0.001 (^^^). Statistically significant differences between cancer cells treated with GOS-activated NK-92 cells vs. cancer cells exposed to GOS at a corresponding concentration as that used for NK cell activation at p < 0.001 (’’’). One-way ANOVA test; Tukey’s post-hoc test. Cancer cell death in response to GOS as well as NK-92 cells activated by the tested compound was visualized by cell double staining with a mixture of propidium iodide and Hoechst 33342. The representative pictures from fluorescence microscopy are presented; the magnification is 200×.

    Techniques Used: Concentration Assay, Activity Assay, MTT Assay, Activation Assay, Double Staining, Fluorescence, Microscopy

    interleukin 2 il 2 dependent natural killer cell cell line nk 92mi  (ATCC)


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    ATCC interleukin 2 il 2 dependent natural killer cell cell line nk 92mi
    Effects on <t>NK</t> <t>cell</t> proliferation of B-cell supernatants containing 0.5 mg/mL extracts.
    Interleukin 2 Il 2 Dependent Natural Killer Cell Cell Line Nk 92mi, supplied by ATCC, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    1) Product Images from "Enhanced Immunomodulatory Activity of Gelatin-Encapsulated Rubus coreanus Miquel Nanoparticles"

    Article Title: Enhanced Immunomodulatory Activity of Gelatin-Encapsulated Rubus coreanus Miquel Nanoparticles

    Journal: International Journal of Molecular Sciences

    doi: 10.3390/ijms12129031

    Effects on NK cell proliferation of B-cell supernatants containing 0.5 mg/mL extracts.
    Figure Legend Snippet: Effects on NK cell proliferation of B-cell supernatants containing 0.5 mg/mL extracts.

    Techniques Used:

    nk92 egfp cd16 pta 8836 cell lines  (ATCC)


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    ATCC nk92 egfp cd16 pta 8836 cell lines
    Neuroblastoma-derived sEVs modulate NK cell maturation in vivo and NK cell-mediated ADCC in vitro. (A) Schematic of NK cell subpopulations. Created with BioRender.com. (B) Representative flow cytometry plots of splenic NK cell subpopulations isolated from 9464D-GD2 tumor-bearing mice receiving the indicated treatments as described in . (C) Quantification of the percentage of immature splenic NK cell (CD27 +CD11b-; left panel) and mature NK cell (CD27- CD11b+; right panel) subpopulations in 9464D-GD2 tumor-bearing mice treated as described in . Mean±SEM, n=7 per group. Student’s t-test. *p<0.05; **p<0.01; ***p<0.001. (D–G) Human (IMR32) or murine (9464D-GD2) neuroblastoma cells treated as indicated in the presence or absence of <t>NK92-CD16-EGFP</t> cells (NK) and monitored for viability utilizing cell impermeant nucleic acid stain YOYO-3 with the IncuCyte S3 Live-Cell Analysis System. The cell-by-cell analysis module was used to quantify viable tumor cells (YOYO3- EGFP-). (D) Kinetic analysis of the IMR32 in vitro NK-cell-mediated ADCC assay. Mean±SEM, n=6. Viable cells calculated as percentage of viable cells in each treatment condition divided by viable cells in untreated control group. (E) Percentage of viable IMR32 cells at 24 hours, n=6. Mean±SD. One-way ANOVA with Tukey’s/Sidak’s post hoc tests. **p<0.01; ***p<0.001. (F) Kinetic analysis of the 9464D-GD2 in vitro NK-cell-mediated ADCC assay. Mean±SEM, n=4. Viable cells calculated as percentage of viable cells in each treatment condition divided by viable cells in untreated control group. (G) Percentage of viable 9464D-GD2 cells at 24 hours, n=4. Mean±SD. One-way ANOVA with Tukey’s/Sidak’s post hoc tests. *p<0.05; **p<0.01; ***p<0.001. ADCC, antibody-dependent cell-mediated cytotoxicity; ANOVA, analysis of variance; DN, double negative; DP, double positive; sEV, small extracellular vesicle.
    Nk92 Egfp Cd16 Pta 8836 Cell Lines, supplied by ATCC, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    1) Product Images from "Small extracellular vesicles induce resistance to anti-GD2 immunotherapy unveiling tipifarnib as an adjunct to neuroblastoma immunotherapy"

    Article Title: Small extracellular vesicles induce resistance to anti-GD2 immunotherapy unveiling tipifarnib as an adjunct to neuroblastoma immunotherapy

    Journal: Journal for Immunotherapy of Cancer

    doi: 10.1136/jitc-2021-004399

    Neuroblastoma-derived sEVs modulate NK cell maturation in vivo and NK cell-mediated ADCC in vitro. (A) Schematic of NK cell subpopulations. Created with BioRender.com. (B) Representative flow cytometry plots of splenic NK cell subpopulations isolated from 9464D-GD2 tumor-bearing mice receiving the indicated treatments as described in . (C) Quantification of the percentage of immature splenic NK cell (CD27 +CD11b-; left panel) and mature NK cell (CD27- CD11b+; right panel) subpopulations in 9464D-GD2 tumor-bearing mice treated as described in . Mean±SEM, n=7 per group. Student’s t-test. *p<0.05; **p<0.01; ***p<0.001. (D–G) Human (IMR32) or murine (9464D-GD2) neuroblastoma cells treated as indicated in the presence or absence of NK92-CD16-EGFP cells (NK) and monitored for viability utilizing cell impermeant nucleic acid stain YOYO-3 with the IncuCyte S3 Live-Cell Analysis System. The cell-by-cell analysis module was used to quantify viable tumor cells (YOYO3- EGFP-). (D) Kinetic analysis of the IMR32 in vitro NK-cell-mediated ADCC assay. Mean±SEM, n=6. Viable cells calculated as percentage of viable cells in each treatment condition divided by viable cells in untreated control group. (E) Percentage of viable IMR32 cells at 24 hours, n=6. Mean±SD. One-way ANOVA with Tukey’s/Sidak’s post hoc tests. **p<0.01; ***p<0.001. (F) Kinetic analysis of the 9464D-GD2 in vitro NK-cell-mediated ADCC assay. Mean±SEM, n=4. Viable cells calculated as percentage of viable cells in each treatment condition divided by viable cells in untreated control group. (G) Percentage of viable 9464D-GD2 cells at 24 hours, n=4. Mean±SD. One-way ANOVA with Tukey’s/Sidak’s post hoc tests. *p<0.05; **p<0.01; ***p<0.001. ADCC, antibody-dependent cell-mediated cytotoxicity; ANOVA, analysis of variance; DN, double negative; DP, double positive; sEV, small extracellular vesicle.
    Figure Legend Snippet: Neuroblastoma-derived sEVs modulate NK cell maturation in vivo and NK cell-mediated ADCC in vitro. (A) Schematic of NK cell subpopulations. Created with BioRender.com. (B) Representative flow cytometry plots of splenic NK cell subpopulations isolated from 9464D-GD2 tumor-bearing mice receiving the indicated treatments as described in . (C) Quantification of the percentage of immature splenic NK cell (CD27 +CD11b-; left panel) and mature NK cell (CD27- CD11b+; right panel) subpopulations in 9464D-GD2 tumor-bearing mice treated as described in . Mean±SEM, n=7 per group. Student’s t-test. *p<0.05; **p<0.01; ***p<0.001. (D–G) Human (IMR32) or murine (9464D-GD2) neuroblastoma cells treated as indicated in the presence or absence of NK92-CD16-EGFP cells (NK) and monitored for viability utilizing cell impermeant nucleic acid stain YOYO-3 with the IncuCyte S3 Live-Cell Analysis System. The cell-by-cell analysis module was used to quantify viable tumor cells (YOYO3- EGFP-). (D) Kinetic analysis of the IMR32 in vitro NK-cell-mediated ADCC assay. Mean±SEM, n=6. Viable cells calculated as percentage of viable cells in each treatment condition divided by viable cells in untreated control group. (E) Percentage of viable IMR32 cells at 24 hours, n=6. Mean±SD. One-way ANOVA with Tukey’s/Sidak’s post hoc tests. **p<0.01; ***p<0.001. (F) Kinetic analysis of the 9464D-GD2 in vitro NK-cell-mediated ADCC assay. Mean±SEM, n=4. Viable cells calculated as percentage of viable cells in each treatment condition divided by viable cells in untreated control group. (G) Percentage of viable 9464D-GD2 cells at 24 hours, n=4. Mean±SD. One-way ANOVA with Tukey’s/Sidak’s post hoc tests. *p<0.05; **p<0.01; ***p<0.001. ADCC, antibody-dependent cell-mediated cytotoxicity; ANOVA, analysis of variance; DN, double negative; DP, double positive; sEV, small extracellular vesicle.

    Techniques Used: Derivative Assay, In Vivo, In Vitro, Flow Cytometry, Isolation, Staining, ADCC Assay

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    ATCC interleukin 2 il 2 dependent natural killer cell cell line nk 92mi
    Effects on <t>NK</t> <t>cell</t> proliferation of B-cell supernatants containing 0.5 mg/mL extracts.
    Interleukin 2 Il 2 Dependent Natural Killer Cell Cell Line Nk 92mi, supplied by ATCC, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    86
    ATCC natural killer cell line nk 92
    Effects on <t>NK</t> <t>cell</t> proliferation of B-cell supernatants containing 0.5 mg/mL extracts.
    Natural Killer Cell Line Nk 92, supplied by ATCC, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    86
    ATCC natural killer 92 nk 92 cell line
    Generation of <t>NK-92-CAR.19-IL-15/IL15Rα</t> cells. (A) Scheme of CAR constructs targeting CD19 (CAR.19) under the control of spleen focus-forming virus (SFFV) promoter. CAR consists of scFv, CD8 hinge and transmembrane region, 41BB costimulatory molecule and CD3ζ signaling molecule. Where indicated, the CAR sequences are followed by a self-cleaving peptide (T2A) and IL-15 or IL-15/IL-15Rα (IL-15 and IL-15Rα fused through a flexible linker). (B, C) NK-92 cells were transduced with the described CAR constructs followed by immunomagnetic enrichment in two steps. (B) Representative dot plots show flow cytometric analysis of CAR expression on freshly transduced and enriched NK-92-CAR.19 cells. (C) The graph shows the frequency of CAR-positive cells before selection, after first and second selection, and 30 days after second selection.
    Natural Killer 92 Nk 92 Cell Line, supplied by ATCC, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    ATCC natural killer nk like cell line yt
    Generation of <t>NK-92-CAR.19-IL-15/IL15Rα</t> cells. (A) Scheme of CAR constructs targeting CD19 (CAR.19) under the control of spleen focus-forming virus (SFFV) promoter. CAR consists of scFv, CD8 hinge and transmembrane region, 41BB costimulatory molecule and CD3ζ signaling molecule. Where indicated, the CAR sequences are followed by a self-cleaving peptide (T2A) and IL-15 or IL-15/IL-15Rα (IL-15 and IL-15Rα fused through a flexible linker). (B, C) NK-92 cells were transduced with the described CAR constructs followed by immunomagnetic enrichment in two steps. (B) Representative dot plots show flow cytometric analysis of CAR expression on freshly transduced and enriched NK-92-CAR.19 cells. (C) The graph shows the frequency of CAR-positive cells before selection, after first and second selection, and 30 days after second selection.
    Natural Killer Nk Like Cell Line Yt, supplied by ATCC, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    ATCC natural killer nk cell line nk
    Generation of <t>NK-92-CAR.19-IL-15/IL15Rα</t> cells. (A) Scheme of CAR constructs targeting CD19 (CAR.19) under the control of spleen focus-forming virus (SFFV) promoter. CAR consists of scFv, CD8 hinge and transmembrane region, 41BB costimulatory molecule and CD3ζ signaling molecule. Where indicated, the CAR sequences are followed by a self-cleaving peptide (T2A) and IL-15 or IL-15/IL-15Rα (IL-15 and IL-15Rα fused through a flexible linker). (B, C) NK-92 cells were transduced with the described CAR constructs followed by immunomagnetic enrichment in two steps. (B) Representative dot plots show flow cytometric analysis of CAR expression on freshly transduced and enriched NK-92-CAR.19 cells. (C) The graph shows the frequency of CAR-positive cells before selection, after first and second selection, and 30 days after second selection.
    Natural Killer Nk Cell Line Nk, supplied by ATCC, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    ATCC human natural killer cell line nk
    Generation of <t>NK-92-CAR.19-IL-15/IL15Rα</t> cells. (A) Scheme of CAR constructs targeting CD19 (CAR.19) under the control of spleen focus-forming virus (SFFV) promoter. CAR consists of scFv, CD8 hinge and transmembrane region, 41BB costimulatory molecule and CD3ζ signaling molecule. Where indicated, the CAR sequences are followed by a self-cleaving peptide (T2A) and IL-15 or IL-15/IL-15Rα (IL-15 and IL-15Rα fused through a flexible linker). (B, C) NK-92 cells were transduced with the described CAR constructs followed by immunomagnetic enrichment in two steps. (B) Representative dot plots show flow cytometric analysis of CAR expression on freshly transduced and enriched NK-92-CAR.19 cells. (C) The graph shows the frequency of CAR-positive cells before selection, after first and second selection, and 30 days after second selection.
    Human Natural Killer Cell Line Nk, supplied by ATCC, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    ATCC nk92 egfp cd16 pta 8836 cell lines
    Neuroblastoma-derived sEVs modulate NK cell maturation in vivo and NK cell-mediated ADCC in vitro. (A) Schematic of NK cell subpopulations. Created with BioRender.com. (B) Representative flow cytometry plots of splenic NK cell subpopulations isolated from 9464D-GD2 tumor-bearing mice receiving the indicated treatments as described in . (C) Quantification of the percentage of immature splenic NK cell (CD27 +CD11b-; left panel) and mature NK cell (CD27- CD11b+; right panel) subpopulations in 9464D-GD2 tumor-bearing mice treated as described in . Mean±SEM, n=7 per group. Student’s t-test. *p<0.05; **p<0.01; ***p<0.001. (D–G) Human (IMR32) or murine (9464D-GD2) neuroblastoma cells treated as indicated in the presence or absence of <t>NK92-CD16-EGFP</t> cells (NK) and monitored for viability utilizing cell impermeant nucleic acid stain YOYO-3 with the IncuCyte S3 Live-Cell Analysis System. The cell-by-cell analysis module was used to quantify viable tumor cells (YOYO3- EGFP-). (D) Kinetic analysis of the IMR32 in vitro NK-cell-mediated ADCC assay. Mean±SEM, n=6. Viable cells calculated as percentage of viable cells in each treatment condition divided by viable cells in untreated control group. (E) Percentage of viable IMR32 cells at 24 hours, n=6. Mean±SD. One-way ANOVA with Tukey’s/Sidak’s post hoc tests. **p<0.01; ***p<0.001. (F) Kinetic analysis of the 9464D-GD2 in vitro NK-cell-mediated ADCC assay. Mean±SEM, n=4. Viable cells calculated as percentage of viable cells in each treatment condition divided by viable cells in untreated control group. (G) Percentage of viable 9464D-GD2 cells at 24 hours, n=4. Mean±SD. One-way ANOVA with Tukey’s/Sidak’s post hoc tests. *p<0.05; **p<0.01; ***p<0.001. ADCC, antibody-dependent cell-mediated cytotoxicity; ANOVA, analysis of variance; DN, double negative; DP, double positive; sEV, small extracellular vesicle.
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    Effects on NK cell proliferation of B-cell supernatants containing 0.5 mg/mL extracts.

    Journal: International Journal of Molecular Sciences

    Article Title: Enhanced Immunomodulatory Activity of Gelatin-Encapsulated Rubus coreanus Miquel Nanoparticles

    doi: 10.3390/ijms12129031

    Figure Lengend Snippet: Effects on NK cell proliferation of B-cell supernatants containing 0.5 mg/mL extracts.

    Article Snippet: The interleukin-2 (IL-2) dependent Natural Killer Cell cell line NK-92MI, (ATCC, CRL-2408) was diluted to 2 × 10 7 cells/mL using 2 mM L-glutamine, 0.2 mM myoinositol, 20 mM folic acid, 0.1 mM 2-mercaptoethanol, and 12.5% FBS in MEM and cultured in T-25 flasks.

    Techniques:

    Generation of NK-92-CAR.19-IL-15/IL15Rα cells. (A) Scheme of CAR constructs targeting CD19 (CAR.19) under the control of spleen focus-forming virus (SFFV) promoter. CAR consists of scFv, CD8 hinge and transmembrane region, 41BB costimulatory molecule and CD3ζ signaling molecule. Where indicated, the CAR sequences are followed by a self-cleaving peptide (T2A) and IL-15 or IL-15/IL-15Rα (IL-15 and IL-15Rα fused through a flexible linker). (B, C) NK-92 cells were transduced with the described CAR constructs followed by immunomagnetic enrichment in two steps. (B) Representative dot plots show flow cytometric analysis of CAR expression on freshly transduced and enriched NK-92-CAR.19 cells. (C) The graph shows the frequency of CAR-positive cells before selection, after first and second selection, and 30 days after second selection.

    Journal: Frontiers in Immunology

    Article Title: Engineering NK-CAR.19 cells with the IL-15/IL-15Rα complex improved proliferation and anti-tumor effect in vivo

    doi: 10.3389/fimmu.2023.1226518

    Figure Lengend Snippet: Generation of NK-92-CAR.19-IL-15/IL15Rα cells. (A) Scheme of CAR constructs targeting CD19 (CAR.19) under the control of spleen focus-forming virus (SFFV) promoter. CAR consists of scFv, CD8 hinge and transmembrane region, 41BB costimulatory molecule and CD3ζ signaling molecule. Where indicated, the CAR sequences are followed by a self-cleaving peptide (T2A) and IL-15 or IL-15/IL-15Rα (IL-15 and IL-15Rα fused through a flexible linker). (B, C) NK-92 cells were transduced with the described CAR constructs followed by immunomagnetic enrichment in two steps. (B) Representative dot plots show flow cytometric analysis of CAR expression on freshly transduced and enriched NK-92-CAR.19 cells. (C) The graph shows the frequency of CAR-positive cells before selection, after first and second selection, and 30 days after second selection.

    Article Snippet: The Natural Killer 92 (NK-92) cell line, as described by Gong et al. (1994) ( ) and obtained from ATCC (CRL-2407™), was cultured using serum-free X-VIVO 10 media (Lonza, Cologne, Germany). containing 5% human heat inactivated AB plasma (Brazilian Blood Donation Service of the Regional Blood Center of Ribeirão Preto), supplemented with 500 IU/ml IL-2 (Clinigen), as previously described ( , ).

    Techniques: Construct, Virus, Transduction, Expressing, Selection

    NK-92-CAR.19-IL-15/IL15Rα cells proliferate independently of exogenous IL-2. Proliferation of parental and genetically modified NK-92 cells: NK-92-CAR.19, NK-92-CAR.19-IL-15 and NK-92-CAR.19-IL-15/IL15Rα in X-Vivo 10 supplemented with 5% of human plasma. (A) Cells were cultured either in presence of IL-2 (500 UI/mL) or (B) in absence of exogenous cytokines. (C) Viability of cell culture with cytokines and (D) without cytokines. Cell culture was followed over 21 days. Pooled data of two independent experiments (each one performed in duplicates) are shown. (E) Quantification of soluble IL-15 in supernatant of NK cell culture by Luminex (n=2). Two-way ANOVA statistical test, Tukey's post-multiple comparison posttest. ****P<0.0001.

    Journal: Frontiers in Immunology

    Article Title: Engineering NK-CAR.19 cells with the IL-15/IL-15Rα complex improved proliferation and anti-tumor effect in vivo

    doi: 10.3389/fimmu.2023.1226518

    Figure Lengend Snippet: NK-92-CAR.19-IL-15/IL15Rα cells proliferate independently of exogenous IL-2. Proliferation of parental and genetically modified NK-92 cells: NK-92-CAR.19, NK-92-CAR.19-IL-15 and NK-92-CAR.19-IL-15/IL15Rα in X-Vivo 10 supplemented with 5% of human plasma. (A) Cells were cultured either in presence of IL-2 (500 UI/mL) or (B) in absence of exogenous cytokines. (C) Viability of cell culture with cytokines and (D) without cytokines. Cell culture was followed over 21 days. Pooled data of two independent experiments (each one performed in duplicates) are shown. (E) Quantification of soluble IL-15 in supernatant of NK cell culture by Luminex (n=2). Two-way ANOVA statistical test, Tukey's post-multiple comparison posttest. ****P<0.0001.

    Article Snippet: The Natural Killer 92 (NK-92) cell line, as described by Gong et al. (1994) ( ) and obtained from ATCC (CRL-2407™), was cultured using serum-free X-VIVO 10 media (Lonza, Cologne, Germany). containing 5% human heat inactivated AB plasma (Brazilian Blood Donation Service of the Regional Blood Center of Ribeirão Preto), supplemented with 500 IU/ml IL-2 (Clinigen), as previously described ( , ).

    Techniques: Genetically Modified, Cell Culture, Luminex, Comparison

    NK-92-CAR.19 cells coexpressing IL-15/Rα enhance CAR-mediated anti-tumor Ncell function. NK-92, NK-92-CAR.19, NK-92-CAR.19-IL-15 and NK-92-CAR.19-IL-15/IL-15Rα cells were co-cultivated with (A) Raji (B) NALM-6 and (C) K562 in 2:1 and 10:1 effector:target cells ratio, for 5 hours. Cytotoxicity was measured by flow cytometry cytotoxicity assay. One-way ANOVA statistical test, Tukey's multiple comparison posttest. ****P < 0.0001; ***P < 0.001; **P < 0.01; *P < 0.05. Mean values + SD are shown. Data pooled from three independent experiments.

    Journal: Frontiers in Immunology

    Article Title: Engineering NK-CAR.19 cells with the IL-15/IL-15Rα complex improved proliferation and anti-tumor effect in vivo

    doi: 10.3389/fimmu.2023.1226518

    Figure Lengend Snippet: NK-92-CAR.19 cells coexpressing IL-15/Rα enhance CAR-mediated anti-tumor Ncell function. NK-92, NK-92-CAR.19, NK-92-CAR.19-IL-15 and NK-92-CAR.19-IL-15/IL-15Rα cells were co-cultivated with (A) Raji (B) NALM-6 and (C) K562 in 2:1 and 10:1 effector:target cells ratio, for 5 hours. Cytotoxicity was measured by flow cytometry cytotoxicity assay. One-way ANOVA statistical test, Tukey's multiple comparison posttest. ****P < 0.0001; ***P < 0.001; **P < 0.01; *P < 0.05. Mean values + SD are shown. Data pooled from three independent experiments.

    Article Snippet: The Natural Killer 92 (NK-92) cell line, as described by Gong et al. (1994) ( ) and obtained from ATCC (CRL-2407™), was cultured using serum-free X-VIVO 10 media (Lonza, Cologne, Germany). containing 5% human heat inactivated AB plasma (Brazilian Blood Donation Service of the Regional Blood Center of Ribeirão Preto), supplemented with 500 IU/ml IL-2 (Clinigen), as previously described ( , ).

    Techniques: Flow Cytometry, Cytotoxicity Assay, Comparison

    NK-92-CAR.19-IL-15/IL15Rα cells produce high amount of anti-tumor pro-inflammatory cytokines. NK-92, CAR.19, CAR.19-IL-15 and CAR.19-IL-15Rα NK-92 cells (2.5 × 10 5 ) were incubated for 5 h with Raji lymphoma cells (A, C) or with Nalm-6 (B, D) at an E/T ratio of 10:1. Supernatants were collected, and the levels of IFN-γ (A, B) and TNF-α (C, D) , granzyme A (E, F) ), granzyme B (G, H) and perforin (I, J) were measured using the Luminex MAGPIX system. NK cells without target cells were included as controls. One-way ANOVA statistical test, Tukey's multiple comparison posttest. Mean values ± SD are shown; n=2. ***P < 0.001; **P < 0.01; *P < 0.05. Data pooled from three independent experiments.

    Journal: Frontiers in Immunology

    Article Title: Engineering NK-CAR.19 cells with the IL-15/IL-15Rα complex improved proliferation and anti-tumor effect in vivo

    doi: 10.3389/fimmu.2023.1226518

    Figure Lengend Snippet: NK-92-CAR.19-IL-15/IL15Rα cells produce high amount of anti-tumor pro-inflammatory cytokines. NK-92, CAR.19, CAR.19-IL-15 and CAR.19-IL-15Rα NK-92 cells (2.5 × 10 5 ) were incubated for 5 h with Raji lymphoma cells (A, C) or with Nalm-6 (B, D) at an E/T ratio of 10:1. Supernatants were collected, and the levels of IFN-γ (A, B) and TNF-α (C, D) , granzyme A (E, F) ), granzyme B (G, H) and perforin (I, J) were measured using the Luminex MAGPIX system. NK cells without target cells were included as controls. One-way ANOVA statistical test, Tukey's multiple comparison posttest. Mean values ± SD are shown; n=2. ***P < 0.001; **P < 0.01; *P < 0.05. Data pooled from three independent experiments.

    Article Snippet: The Natural Killer 92 (NK-92) cell line, as described by Gong et al. (1994) ( ) and obtained from ATCC (CRL-2407™), was cultured using serum-free X-VIVO 10 media (Lonza, Cologne, Germany). containing 5% human heat inactivated AB plasma (Brazilian Blood Donation Service of the Regional Blood Center of Ribeirão Preto), supplemented with 500 IU/ml IL-2 (Clinigen), as previously described ( , ).

    Techniques: Incubation, Luminex, Comparison

    Metascape functional analysis of transcriptome profiles of NK-92-CAR.19 cells after coculture with a target B cell line. (A) The Circos plot shows how genes from the different input gene lists after coculture setups overlap. On the outside, the arc represents the identity of each gene list. On the inside, the orange color represents the genes that appear in multiple lists, and the light orange color represents genes that are unique to that gene list. Purple lines link the same genes shared by multiple gene lists. Blue lines link the genes that fall into the same ontology term (the term has to be statistically significantly enriched). The greater the number of purple links and the longer the dark orange arcs, the greater is the overlap among the input gene lists. Blue links indicate the amount of functional overlap among the input gene lists. (B) Networks of Protein-Protein interaction (PPI) identified in the gene lists for comparing across NK92-CAR groups and NK92-WT, and illustrated using Cytoscape 3.9.1. (C) Summary of enrichment analysis done using the DEG_up from (A) and the Trancription_Factor_Targets as an ontology category (Metascape).

    Journal: Frontiers in Immunology

    Article Title: Engineering NK-CAR.19 cells with the IL-15/IL-15Rα complex improved proliferation and anti-tumor effect in vivo

    doi: 10.3389/fimmu.2023.1226518

    Figure Lengend Snippet: Metascape functional analysis of transcriptome profiles of NK-92-CAR.19 cells after coculture with a target B cell line. (A) The Circos plot shows how genes from the different input gene lists after coculture setups overlap. On the outside, the arc represents the identity of each gene list. On the inside, the orange color represents the genes that appear in multiple lists, and the light orange color represents genes that are unique to that gene list. Purple lines link the same genes shared by multiple gene lists. Blue lines link the genes that fall into the same ontology term (the term has to be statistically significantly enriched). The greater the number of purple links and the longer the dark orange arcs, the greater is the overlap among the input gene lists. Blue links indicate the amount of functional overlap among the input gene lists. (B) Networks of Protein-Protein interaction (PPI) identified in the gene lists for comparing across NK92-CAR groups and NK92-WT, and illustrated using Cytoscape 3.9.1. (C) Summary of enrichment analysis done using the DEG_up from (A) and the Trancription_Factor_Targets as an ontology category (Metascape).

    Article Snippet: The Natural Killer 92 (NK-92) cell line, as described by Gong et al. (1994) ( ) and obtained from ATCC (CRL-2407™), was cultured using serum-free X-VIVO 10 media (Lonza, Cologne, Germany). containing 5% human heat inactivated AB plasma (Brazilian Blood Donation Service of the Regional Blood Center of Ribeirão Preto), supplemented with 500 IU/ml IL-2 (Clinigen), as previously described ( , ).

    Techniques: Functional Assay

    Transgenic IL-15 or IL-15/Rα maintains low expression of immune checkpoint molecules in NK-92-CAR.19 cell variants. NK-92-CAR.19 cells were cultured unstimulated (-) or stimulated (+) with Raji cells at an E:T ratio of 1:2, followed by restimulation with fresh target cells after 24 and 48 hours. Three days after the beginning of the experiment, the surface expression of LAG-3 (A) , PD-1 (B) and TIM-3 (C) were analyzed by flow cytometry. Three independent experiments were performed. One-way ANOVA statistical test, Tukey's post-hoc multiple comparison. Mean values + SD are shown. Gray: NK-92, Black: coculture of NK-92 and Raji cells, Orange: CAR.19, Brown: coculture of CAR.19 and Raji cells, Pink: CAR.19-IL-15, Red: cocultured of CAR.19-IL-15 and Raji cells, Light blue: CAR.19-IL-15RA, and Blue: coculture of CAR.19-IL-15 and Raji cells. (D) Representative flow cytometry histograms plots showing the mean fluorescence intensity of LAG-3, PD-1 and TIM-3 in NK-92 cell and, in the NK-92-CAR.19 cell variants. ***P < 0.001; **P < 0.01; *P < 0.05.

    Journal: Frontiers in Immunology

    Article Title: Engineering NK-CAR.19 cells with the IL-15/IL-15Rα complex improved proliferation and anti-tumor effect in vivo

    doi: 10.3389/fimmu.2023.1226518

    Figure Lengend Snippet: Transgenic IL-15 or IL-15/Rα maintains low expression of immune checkpoint molecules in NK-92-CAR.19 cell variants. NK-92-CAR.19 cells were cultured unstimulated (-) or stimulated (+) with Raji cells at an E:T ratio of 1:2, followed by restimulation with fresh target cells after 24 and 48 hours. Three days after the beginning of the experiment, the surface expression of LAG-3 (A) , PD-1 (B) and TIM-3 (C) were analyzed by flow cytometry. Three independent experiments were performed. One-way ANOVA statistical test, Tukey's post-hoc multiple comparison. Mean values + SD are shown. Gray: NK-92, Black: coculture of NK-92 and Raji cells, Orange: CAR.19, Brown: coculture of CAR.19 and Raji cells, Pink: CAR.19-IL-15, Red: cocultured of CAR.19-IL-15 and Raji cells, Light blue: CAR.19-IL-15RA, and Blue: coculture of CAR.19-IL-15 and Raji cells. (D) Representative flow cytometry histograms plots showing the mean fluorescence intensity of LAG-3, PD-1 and TIM-3 in NK-92 cell and, in the NK-92-CAR.19 cell variants. ***P < 0.001; **P < 0.01; *P < 0.05.

    Article Snippet: The Natural Killer 92 (NK-92) cell line, as described by Gong et al. (1994) ( ) and obtained from ATCC (CRL-2407™), was cultured using serum-free X-VIVO 10 media (Lonza, Cologne, Germany). containing 5% human heat inactivated AB plasma (Brazilian Blood Donation Service of the Regional Blood Center of Ribeirão Preto), supplemented with 500 IU/ml IL-2 (Clinigen), as previously described ( , ).

    Techniques: Transgenic Assay, Expressing, Cell Culture, Flow Cytometry, Comparison, Fluorescence

    NK-92-CAR.19-IL15/IL-15Rα cells eliminate lymphoma B-cells and reduce tumor burden in vivo . (A) Scheme of an in vivo study demonstrating the anti-tumor activity of transduced NK-92-CAR.19 cells in a xenogeneic NSG mouse model of disseminated human B-cell hematologic malignancy. NSG mice were intravenously injected with 2 × 10 4 Raji-Luc lymphoma B-cells. On days 4, 7, 10, 12, 15, and 19, animals were treated by intravenous injection of 7 × 10 6 NK-92 cells (n=5). (B) Lymphoma development was monitored by in vivo bioluminescence imaging. Images were taken on days 8, 14, 21, 26, and 29. (C) Bioluminescence intensity was quantified and means + SEM are shown. (D) Bioluminescence of individual mice shown on day 29. Means + SEM are shown. Statistics were determined by the Mann-Whitney test. Each experimental group consisted of five mice. **P < 0.01; *P < 0.05.

    Journal: Frontiers in Immunology

    Article Title: Engineering NK-CAR.19 cells with the IL-15/IL-15Rα complex improved proliferation and anti-tumor effect in vivo

    doi: 10.3389/fimmu.2023.1226518

    Figure Lengend Snippet: NK-92-CAR.19-IL15/IL-15Rα cells eliminate lymphoma B-cells and reduce tumor burden in vivo . (A) Scheme of an in vivo study demonstrating the anti-tumor activity of transduced NK-92-CAR.19 cells in a xenogeneic NSG mouse model of disseminated human B-cell hematologic malignancy. NSG mice were intravenously injected with 2 × 10 4 Raji-Luc lymphoma B-cells. On days 4, 7, 10, 12, 15, and 19, animals were treated by intravenous injection of 7 × 10 6 NK-92 cells (n=5). (B) Lymphoma development was monitored by in vivo bioluminescence imaging. Images were taken on days 8, 14, 21, 26, and 29. (C) Bioluminescence intensity was quantified and means + SEM are shown. (D) Bioluminescence of individual mice shown on day 29. Means + SEM are shown. Statistics were determined by the Mann-Whitney test. Each experimental group consisted of five mice. **P < 0.01; *P < 0.05.

    Article Snippet: The Natural Killer 92 (NK-92) cell line, as described by Gong et al. (1994) ( ) and obtained from ATCC (CRL-2407™), was cultured using serum-free X-VIVO 10 media (Lonza, Cologne, Germany). containing 5% human heat inactivated AB plasma (Brazilian Blood Donation Service of the Regional Blood Center of Ribeirão Preto), supplemented with 500 IU/ml IL-2 (Clinigen), as previously described ( , ).

    Techniques: In Vivo, Activity Assay, Injection, Imaging, MANN-WHITNEY

    Proposed model of action of engineered NK-92-CAR-19-IL-15/IL-15Rα cells. In the absence of exogenous IL-2: (A) NK-92-CAR.19 cells remain non-proliferative. (B) NK-92-CAR.19-IL-15 cells exhibit modest proliferation up to day 12, as they release soluble IL-15 that subsequently engages with IL-2/IL-15Rβ–γc receptors on their own membrane or neighboring NK cells. (C) NK-92-CAR.19-IL-15/IL-15Rα cells display robust proliferation, driven by the presence of IL-15/IL-15Rα complexes on their cell membrane. This activates the cells through cis-presentation or may stimulate adjacent NK cells expressing membrane-bound IL-2/IL-15Rβ–γc receptors via trans-presentation. The signaling from IL-15 prompts all three NK-92-CAR-19 cell variants to upregulate the PI3K/AKT and JAK/STAT pathways. However, this upregulation is notably more pronounced in NK-92-CAR.19-IL-15/IL-15Rα cells compared to the other variants. Consequently, this substantial enhancement in signaling leads to heightened proliferative capacity, increased secretion of proinflammatory cytokines, and exceptionally potent cytotoxic activity in these pioneering engineered NK-92 cells.

    Journal: Frontiers in Immunology

    Article Title: Engineering NK-CAR.19 cells with the IL-15/IL-15Rα complex improved proliferation and anti-tumor effect in vivo

    doi: 10.3389/fimmu.2023.1226518

    Figure Lengend Snippet: Proposed model of action of engineered NK-92-CAR-19-IL-15/IL-15Rα cells. In the absence of exogenous IL-2: (A) NK-92-CAR.19 cells remain non-proliferative. (B) NK-92-CAR.19-IL-15 cells exhibit modest proliferation up to day 12, as they release soluble IL-15 that subsequently engages with IL-2/IL-15Rβ–γc receptors on their own membrane or neighboring NK cells. (C) NK-92-CAR.19-IL-15/IL-15Rα cells display robust proliferation, driven by the presence of IL-15/IL-15Rα complexes on their cell membrane. This activates the cells through cis-presentation or may stimulate adjacent NK cells expressing membrane-bound IL-2/IL-15Rβ–γc receptors via trans-presentation. The signaling from IL-15 prompts all three NK-92-CAR-19 cell variants to upregulate the PI3K/AKT and JAK/STAT pathways. However, this upregulation is notably more pronounced in NK-92-CAR.19-IL-15/IL-15Rα cells compared to the other variants. Consequently, this substantial enhancement in signaling leads to heightened proliferative capacity, increased secretion of proinflammatory cytokines, and exceptionally potent cytotoxic activity in these pioneering engineered NK-92 cells.

    Article Snippet: The Natural Killer 92 (NK-92) cell line, as described by Gong et al. (1994) ( ) and obtained from ATCC (CRL-2407™), was cultured using serum-free X-VIVO 10 media (Lonza, Cologne, Germany). containing 5% human heat inactivated AB plasma (Brazilian Blood Donation Service of the Regional Blood Center of Ribeirão Preto), supplemented with 500 IU/ml IL-2 (Clinigen), as previously described ( , ).

    Techniques: Membrane, Expressing, Activity Assay

    Neuroblastoma-derived sEVs modulate NK cell maturation in vivo and NK cell-mediated ADCC in vitro. (A) Schematic of NK cell subpopulations. Created with BioRender.com. (B) Representative flow cytometry plots of splenic NK cell subpopulations isolated from 9464D-GD2 tumor-bearing mice receiving the indicated treatments as described in . (C) Quantification of the percentage of immature splenic NK cell (CD27 +CD11b-; left panel) and mature NK cell (CD27- CD11b+; right panel) subpopulations in 9464D-GD2 tumor-bearing mice treated as described in . Mean±SEM, n=7 per group. Student’s t-test. *p<0.05; **p<0.01; ***p<0.001. (D–G) Human (IMR32) or murine (9464D-GD2) neuroblastoma cells treated as indicated in the presence or absence of NK92-CD16-EGFP cells (NK) and monitored for viability utilizing cell impermeant nucleic acid stain YOYO-3 with the IncuCyte S3 Live-Cell Analysis System. The cell-by-cell analysis module was used to quantify viable tumor cells (YOYO3- EGFP-). (D) Kinetic analysis of the IMR32 in vitro NK-cell-mediated ADCC assay. Mean±SEM, n=6. Viable cells calculated as percentage of viable cells in each treatment condition divided by viable cells in untreated control group. (E) Percentage of viable IMR32 cells at 24 hours, n=6. Mean±SD. One-way ANOVA with Tukey’s/Sidak’s post hoc tests. **p<0.01; ***p<0.001. (F) Kinetic analysis of the 9464D-GD2 in vitro NK-cell-mediated ADCC assay. Mean±SEM, n=4. Viable cells calculated as percentage of viable cells in each treatment condition divided by viable cells in untreated control group. (G) Percentage of viable 9464D-GD2 cells at 24 hours, n=4. Mean±SD. One-way ANOVA with Tukey’s/Sidak’s post hoc tests. *p<0.05; **p<0.01; ***p<0.001. ADCC, antibody-dependent cell-mediated cytotoxicity; ANOVA, analysis of variance; DN, double negative; DP, double positive; sEV, small extracellular vesicle.

    Journal: Journal for Immunotherapy of Cancer

    Article Title: Small extracellular vesicles induce resistance to anti-GD2 immunotherapy unveiling tipifarnib as an adjunct to neuroblastoma immunotherapy

    doi: 10.1136/jitc-2021-004399

    Figure Lengend Snippet: Neuroblastoma-derived sEVs modulate NK cell maturation in vivo and NK cell-mediated ADCC in vitro. (A) Schematic of NK cell subpopulations. Created with BioRender.com. (B) Representative flow cytometry plots of splenic NK cell subpopulations isolated from 9464D-GD2 tumor-bearing mice receiving the indicated treatments as described in . (C) Quantification of the percentage of immature splenic NK cell (CD27 +CD11b-; left panel) and mature NK cell (CD27- CD11b+; right panel) subpopulations in 9464D-GD2 tumor-bearing mice treated as described in . Mean±SEM, n=7 per group. Student’s t-test. *p<0.05; **p<0.01; ***p<0.001. (D–G) Human (IMR32) or murine (9464D-GD2) neuroblastoma cells treated as indicated in the presence or absence of NK92-CD16-EGFP cells (NK) and monitored for viability utilizing cell impermeant nucleic acid stain YOYO-3 with the IncuCyte S3 Live-Cell Analysis System. The cell-by-cell analysis module was used to quantify viable tumor cells (YOYO3- EGFP-). (D) Kinetic analysis of the IMR32 in vitro NK-cell-mediated ADCC assay. Mean±SEM, n=6. Viable cells calculated as percentage of viable cells in each treatment condition divided by viable cells in untreated control group. (E) Percentage of viable IMR32 cells at 24 hours, n=6. Mean±SD. One-way ANOVA with Tukey’s/Sidak’s post hoc tests. **p<0.01; ***p<0.001. (F) Kinetic analysis of the 9464D-GD2 in vitro NK-cell-mediated ADCC assay. Mean±SEM, n=4. Viable cells calculated as percentage of viable cells in each treatment condition divided by viable cells in untreated control group. (G) Percentage of viable 9464D-GD2 cells at 24 hours, n=4. Mean±SD. One-way ANOVA with Tukey’s/Sidak’s post hoc tests. *p<0.05; **p<0.01; ***p<0.001. ADCC, antibody-dependent cell-mediated cytotoxicity; ANOVA, analysis of variance; DN, double negative; DP, double positive; sEV, small extracellular vesicle.

    Article Snippet: Human IMR32 neuroblastoma (CCL-127), HEK 293T/17 (CRL-11268), and NK92-EGFP-CD16 (PTA-8836) cell lines were purchased from ATCC.

    Techniques: Derivative Assay, In Vivo, In Vitro, Flow Cytometry, Isolation, Staining, ADCC Assay