Journal: Biology

Article Title: Engineering a Dual Specificity γδ T-Cell Receptor for Cancer Immunotherapy

doi: 10.3390/biology13030196

Figure Lengend Snippet: Evaluation of αvβ6-dependent activation of chimeric G115 γδ TCRs. ( A ) Analysis of integrin expression on A375 puro and A375-β6 cells. Red–integrin; blue–isotype control. Data are representative of 3 independent replicates. Co-cultures were performed between A375 puro ( B ) and A375-β6 ( C ) tumour cells and untransduced (untrans) or transduced T-cell populations at an effector-to-target ratio of 1:1 for 72 h. Residual tumour cell viability was determined using an MTT assay (mean ± SEM of indicated replicates). Statistical analysis was performed using one-way ANOVA; **** p < 0.0001. ( D ) Supernatants collected from co-cultures described in ( B , C ) were analysed for IFN-γ by ELISA (mean ± SEM; n = 21–45 from 4–6 independent donors). Statistical analysis was performed using one-way ANOVA; * p < 0.05; ** p < 0.01.

Article Snippet: Integrin expression was detected by flow cytometry using the following antibodies: αvβ3 integrin (FAB3050A, R&D Systems, Minneapolis, MN, USA), αvβ5 integrin (FAB2528A, R&D systems), αvβ6 integrin (FAB4155A, R&D Systems) and αvβ8 integrin (FAB4775A, R&D systems).

Techniques: Activation Assay, Expressing, Control, MTT Assay, Enzyme-linked Immunosorbent Assay

Journal: Biology

Article Title: Engineering a Dual Specificity γδ T-Cell Receptor for Cancer Immunotherapy

doi: 10.3390/biology13030196

Figure Lengend Snippet: Evaluation of PAg-dependent activation of chimeric G115 γδ TCRs by ffLuc-expressing K562 cells. ( A ) Analysis of αvβ6 integrin expression on ffLuc + K562 cells. Red–integrin; blue–isotype control. Data are representative of 3 independent replicates. ( B ) Firefly luciferase-expressing K562 cells were pre-incubated with the indicated Zol concentration for 24 h prior to the establishment of co-cultures with untrans(duced) or transduced T-cell populations at an effector to target ratio 1:1 for 72 h. Data show mean ± SEM of residual K562 viability (n = 6–11 from 4 independent donors), as determined by luciferase assay. Statistical analysis was performed using two-way ANOVA; *** p < 0.001; **** p < 0.0001. ( C ) Supernatants collected from co-cultures described in B were analysed for IFN-γ by ELISA (mean ± SEM; n = 19 from 3 independent donors). Statistical analysis was performed using two-way ANOVA; *** p < 0.001; **** p < 0.0001.

Article Snippet: Integrin expression was detected by flow cytometry using the following antibodies: αvβ3 integrin (FAB3050A, R&D Systems, Minneapolis, MN, USA), αvβ5 integrin (FAB2528A, R&D systems), αvβ6 integrin (FAB4155A, R&D Systems) and αvβ8 integrin (FAB4775A, R&D systems).

Techniques: Activation Assay, Expressing, Control, Luciferase, Incubation, Concentration Assay, Enzyme-linked Immunosorbent Assay

Journal: Biology

Article Title: Engineering a Dual Specificity γδ T-Cell Receptor for Cancer Immunotherapy

doi: 10.3390/biology13030196

Figure Lengend Snippet: Evaluation of the anti-tumour activity of chimeric G115 γδ TCRs against BxPC3 pancreatic tumour cells. ( A ) Analysis of integrin expression on BxPC3 cells. Red–integrin; blue–isotype control. Data are representative of 3 independent replicates. Co-cultures were performed between BxPC3 tumour cells (No Zol; B ) or Zol-sensitised BxPC3 tumour cells (+Zol; C ) and untrans(duced) or transduced T-cell populations at an effector to target ratio of 1:1 for 72 h. Tumour cell viability was determined using an MTT assay (mean ± SEM of indicated replicates). Statistical analysis was performed using one-way ANOVA; **** p < 0.0001, * p < 0.05, N/S–not significant.

Article Snippet: Integrin expression was detected by flow cytometry using the following antibodies: αvβ3 integrin (FAB3050A, R&D Systems, Minneapolis, MN, USA), αvβ5 integrin (FAB2528A, R&D systems), αvβ6 integrin (FAB4155A, R&D Systems) and αvβ8 integrin (FAB4775A, R&D systems).

Techniques: Activity Assay, Expressing, Control, MTT Assay

Journal: Biology

Article Title: Engineering a Dual Specificity γδ T-Cell Receptor for Cancer Immunotherapy

doi: 10.3390/biology13030196

Figure Lengend Snippet: Evaluation of the anti-tumour activity of chimeric G115 γδ TCRs against Panc1 pancreatic tumour cells. ( A ) Analysis of integrin expression on Panc1 cells. Red–integrin; blue–isotype control. Data are representative of 3 independent replicates. Co-cultures were performed between Panc1 tumour cells (No Zol; B ) or Zol-sensitised Panc1 tumour cells (+ Zol; C ) and untransduced (untrans) or transduced T-cell populations at an effector to target ratio of 1:1 for 72 h. Tumour cell viability was determined using an MTT assay (mean ± SEM of indicated replicates). Statistical analysis was performed using one-way ANOVA; **** p < 0.0001, ** p < 0.01, N/S–not significant.

Article Snippet: Integrin expression was detected by flow cytometry using the following antibodies: αvβ3 integrin (FAB3050A, R&D Systems, Minneapolis, MN, USA), αvβ5 integrin (FAB2528A, R&D systems), αvβ6 integrin (FAB4155A, R&D Systems) and αvβ8 integrin (FAB4775A, R&D systems).

Techniques: Activity Assay, Expressing, Control, MTT Assay

Journal: Nature immunology

Article Title: Fungal sensing by dectin-1 directs the non-pathogenic polarization of T H 17 cells through balanced type I IFN responses in human DCs.

doi: 10.1038/s41590-022-01348-2

Figure Lengend Snippet: Fig. 6 | Dectin-1 induces αvβ8 expression via IFN-β, which is critical for TGF-β activation and non-pathogenic TH17 polarization. a,b, SEAP assay on supernatant of HEK-Blue TGF-β reporter cells for quantification of active TGF-β in the supernatant of unstimulated DCs or after stimulation of DCs with curdlan or C. albicans, in the presence of blocking αvβ1, αvβ3, αvβ5, αvβ6 and αvβ8 antibodies or isotype (IgG1 and IgG2a) control antibodies (a) or after transduction with non-targeting (control) or specific siRNAs to silence αv (ITGAV) or β8 (ITGB8) expression (b) at 24 h (a, IgG1, αvβ1, αvβ3, αvβ5, αvβ6 Ab n = 2, IgG2a, αvβ8 Ab n = 5; b, n = 4). Data in a,b represent mean ± s.d. of independent donors. **P < 0.01, *P < 0.05 (paired, two-tailed Student’s t-test), calculated between untreated and treated (a) or control and specific siRNA-transduced (b) samples that were likewise stimulated. c,d,g,h, Flow cytometry analyses by staining for αv (c) or β8 (d,g,h) expression (FI) in unstimulated DCs or after stimulation of DCs with curdlan or C. albicans (d), in the presence of blocking dectin-1 or IFN-α/ βR antibodies (g) or after silencing of IRF1 or IRF5 expression (h), at indicated times (c, n = 2; d,g, n = 3; h, n = 4). Isotype indicates negative control staining. Representative histograms for independent donors are shown. e,f, Real-time PCR analyses of ITGB8 relative mRNA levels in unstimulated DCs or after stimulation of

Article Snippet: DCs were preincubated for 2 h with MMP14 inhibitor NSC405020 (100 μM; Tocris) or blocking antibodies, anti-dectin-1 (20 μg ml−1; clone 259931, MAB1859, R&D Systems), anti-IFN-α/βR2 (20 μg ml−1; clone MMHAR-2, PBL Assay Science), anti-αvβ1 (10 μg ml−1; clone P5D2, MAB17781, R&D Systems), anti-αvβ3 (10 μg ml−1; clone 23C6, MAB3050, R&D Systems), anti-αvβ5 (10 μg ml−1; clone P5H9, MAB2528, R&D Systems), anti-αvβ6 (10 μg ml−1; clone 10D5, ab77906, Abcam), anti-αvβ8 (10 μg ml−1; kind gift from S.L.

Techniques: Expressing, Activation Assay, SEAP Assay, Blocking Assay, Control, Transduction, Two Tailed Test, Flow Cytometry, Staining, Negative Control, Real-time Polymerase Chain Reaction

Journal: Radiation Oncology (London, England)

Article Title: Targeting α ν β 3 and α ν β 5 inhibits photon-induced hypermigration of malignant glioma cells

doi: 10.1186/1748-717X-6-132

Figure Lengend Snippet: Photon-induced stimulation of integrin expression . A: FACS analysis of α ν β 3 (upper row) and α ν β 5 (lower row) expression with (red) and without (black) photon irradiation with single doses of 2 Gy (left) and 10 Gy (right). B: Graphical analysis of integrin α ν β 3 (gray bars) and α ν β 5 (black bars) expression with (bars) and without (red line) irradiation with single photon doses of 2, 5, and 10 Gy in U87 (upper chart) and Ln229 (lower chart) glioma cells. Display of mean value ± standard deviation (SD); statistical analysis using Student's t -test (*, indicating significance p < 0.05)

Article Snippet: Integrin blockade was performed using monoclonal antibodies directed against α ν β 3 - (MAB3050, R&D) and α ν β 5 -integrins (MAB 2528, R&D).

Techniques: Expressing, Irradiation, Standard Deviation

Journal: Radiation Oncology (London, England)

Article Title: Targeting α ν β 3 and α ν β 5 inhibits photon-induced hypermigration of malignant glioma cells

doi: 10.1186/1748-717X-6-132

Figure Lengend Snippet: Inhibiton of Ln229 glioma cell migration by integrin inhibition . Quantitative analysis of Vn-based transmigration of Ln229 glioma cells following single photon doses of 0, 2, and 10 Gy without and with the addition of 50 ng/ml anti-α ν β 3 - and -α ν β 5 -antibodies and corresponding isotype controls. Display of mean value ± standard deviation (SD); statistical analysis using Student's t -test (*, indicating significance p < 0.05)

Article Snippet: Integrin blockade was performed using monoclonal antibodies directed against α ν β 3 - (MAB3050, R&D) and α ν β 5 -integrins (MAB 2528, R&D).

Techniques: Migration, Inhibition, Transmigration Assay, Standard Deviation

Journal: Radiation Oncology (London, England)

Article Title: Targeting α ν β 3 and α ν β 5 inhibits photon-induced hypermigration of malignant glioma cells

doi: 10.1186/1748-717X-6-132

Figure Lengend Snippet: Inhibiton of U87 glioma cell migration by integrin inhibition . Quantitative analysis of Vn-based transmigration of U87 glioma cells following single photon doses of 0, 2, and 10 Gy without and with the addition of 50 ng/ml anti-α ν β 3 - and -α ν β 5 -antibodies and corresponding isotype controls.

Article Snippet: Integrin blockade was performed using monoclonal antibodies directed against α ν β 3 - (MAB3050, R&D) and α ν β 5 -integrins (MAB 2528, R&D).

Techniques: Migration, Inhibition, Transmigration Assay