Journal: Cancer Immunology Research

Article Title: Granulocyte–Macrophage Colony-Stimulating Factor Influence on Soluble and Membrane-Bound ICOS in Combination with Immune Checkpoint Blockade

doi: 10.1158/2326-6066.CIR-22-0702

Figure Lengend Snippet: Comparison between human ICOS full length and splice variant. A, Schematic representation of full length and spliced variant of human ICOS. The full length of ICOS consists of 5 exons. The gray boxes denote untranslated regions, and the black boxes denote coding sequences. The transmembrane domain (TM) is located in exon 3 and labeled with a green bar. The splicing region is shown in red. B, Comparison of cDNA nucleotide sequences, amino acid sequences, and 5′UTR sequences between ICOS-FL and the splice variant. Nucleotide sequences of cDNA for ICOS-FL (b1) and the splice variant of ICOS (b4) are represented; spliced region of ICOS is shown in red and delineated by red brackets; the nucleotides and amino acids of TM are shown in green. Asterisk indicates the stop codon. Amino acid sequences of full length of ICOS (b2) and the splice variant of ICOS (b5) are shown. The underline depicts additional differences in amino acid sequence compared with ICOS-FL. “Stop” represents a stop codon. 5′UTR sequences of ICOS-FL (b3) and the splice variant of ICOS (b6) are shown. The 5′UTR of the ICOS-SV lacks 32 nucleotides compared with the 5′UTR of ICOS-FL shown in red and delineated by red brackets. C, Schematic diagram of ICOS-FL and the ICOS-SV protein. Thirty-two out of 38 amino acids in the intracellular domain are spliced, as shown in red.

Article Snippet: To evaluate soluble ICOS-SVs in the sera of melanoma patients or cell culture supernatants, the Human ICOS DuoSet ELISA kit (DY169, R&D Systems) was used according to the manufacturer's protocol.

Techniques: Comparison, Variant Assay, Labeling, Sequencing

Journal: Cancer Immunology Research

Article Title: Granulocyte–Macrophage Colony-Stimulating Factor Influence on Soluble and Membrane-Bound ICOS in Combination with Immune Checkpoint Blockade

doi: 10.1158/2326-6066.CIR-22-0702

Figure Lengend Snippet: Secretion of the ICOS-SV from ICOS-SV–overexpressing cells. A, RT-PCR to detect the ICOS-SV in human T cells. Total RNA was isolated from purified T cells in PBMCs of a healthy donor. Expression of the ICOS variant was examined by RT-PCR with specific primers. RT-PCR of ICOS-FL is shown as a control. B, DNA sequencing of ICOS-SV cDNA from human T cells. The cDNA of the PCR product was cloned into a TOPO TA vector. The ICOS-SV was confirmed by DNA sequencing using an M13 forward primer. Splicing point is shown by the red arrow. C, DNA sequencing of ICOS-FL cDNA is shown as a control. No splicing occurs in the region corresponding to the position in ( B ), as shown in blue. D, Expression of ICOS-SV and ICOS-FL in 293T cells determined by immunoblot. 293T cells were transduced with recombinant GFP retroviral vector encoding either ICOS-SV or ICOS-FL as a positive control. The parental cells and the GFP-empty vector–transduced cells were used as negative controls. Sample loading was normalized to actin. E, Cell culture supernatants were collected from the cells described in ( A ). Secreted ICOS-SV was examined by ELISA. Two-tailed unpaired t test was used to compare ICOS secretion between the two groups. ***, P < 0.001. Standard deviation of the mean (SD) is shown. F, To validate the secreted ICOS variant protein in the cell supernatant, immunoprecipitation, SDS-PAGE, and immunoblotting assays were performed. Sample loading was normalized to cell numbers.

Article Snippet: To evaluate soluble ICOS-SVs in the sera of melanoma patients or cell culture supernatants, the Human ICOS DuoSet ELISA kit (DY169, R&D Systems) was used according to the manufacturer's protocol.

Techniques: Reverse Transcription Polymerase Chain Reaction, Isolation, Purification, Expressing, Variant Assay, Control, DNA Sequencing, Clone Assay, Plasmid Preparation, Western Blot, Transduction, Recombinant, Retroviral, Positive Control, Cell Culture, Enzyme-linked Immunosorbent Assay, Two Tailed Test, Standard Deviation, Immunoprecipitation, SDS Page

Journal: Cancer Immunology Research

Article Title: Granulocyte–Macrophage Colony-Stimulating Factor Influence on Soluble and Membrane-Bound ICOS in Combination with Immune Checkpoint Blockade

doi: 10.1158/2326-6066.CIR-22-0702

Figure Lengend Snippet: Suppressive effects of sICOS-SV on the costimulation of T cells. A, CHO-K1 cells were transduced with either ICOSL-expressing lentiviral vector or GFP-empty vector (CHO-ICOSL − ) as a negative control. Expression of ICOSL on the cell surface of CHO-K1 was examined by flow cytometry using anti-ICOSL (open histogram with solid line) or lentivirus GFP-empty vector (open histogram with dotted line). Isotype antibody was used as a negative control (filled histogram). B, ICOSL-expressing CHO-K1 cells were stained with ICOS-SV Ig (red line), control IgG Ig (blue line), or PD-1 Ig (orange line) and analyzed by flow cytometry. Both soluble control Ig and PD-1 Ig were used as negative controls. GFP-empty vector-expression CHO-K1 cells stained with ICOS-SV Ig (green line) were used as another negative control for the binding assay. Isotype antibody control was used as a negative control (gray filled histogram). C, CD154 expression was determined by FACS analysis. Isotype control antibody is shown in gray. Activated T cells treated with a suboptimal dose of anti-CD3 were incubated with control Ig + CHO-ICOSL − cells (CHO cells transduced with GFP-empty vector, blue), control Ig + CHO-ICOSL + cells (green), PD-1 Ig + CHO-ICOSL + cells (red), or ICOS-SV Ig + CHO-ICOSL + cells (yellow). Both control Ig and PD-1 Ig were used as negative controls. D, Two-tailed unpaired t test was used to compare expression of CD154 between two groups. **, P < 0.01; ***, P < 0.001. Standard deviation of the mean (SD) is shown. E, CD69 expression was determined by FACS analysis. Isotype control antibody is shown in gray. Activated T cells treated with a suboptimal dose of anti-CD3 were incubated with control Ig + CHO-ICOSL − cells (CHO cells transduced with GFP-empty vector, blue), control Ig + CHO-ICOSL + cells (green), PD-1 Ig + CHO-ICOSL + cells (red), or ICOS-SV Ig + CHO-ICOSL + cells (yellow). Both control Ig and PD-1 Ig were used as negative controls. F, Two-tailed unpaired t test was used to compare expression of CD69 between the two groups. **, P < 0.01; ***, P < 0.001. Standard deviation of the mean (SD) is shown. G, CHO cells transduced with GFP-empty vector (CHO-ICOSL − ) were used as negative controls. Sample loading was normalized to total pan-Akt. H, T-cell proliferation was determined by a [ 3 H]-TdR thymidine incorporation assay. CHO cells transduced with empty vector (CHO-ICOSL − ) were used as negative controls. Two-tailed unpaired t test was used to compare 3 H uptake between the two groups, respectively. *, P < 0.1. Standard deviation of the mean (SD) is shown. I, Schematic diagram of ICOS/ICOSL costimulatory T-cell proliferation, as well as the blocking function of the sICOS-SV. Depicted are the cytoplasmic tail sequences of ICOS-FL and sICOS-SV isoforms. The YMFM Src Homology 2 (SH2) binding motif in the cytoplasmic tail of the ICOS-FL is highlighted in pink. Upon ICOS engagement by the ICOSL on CHO cells, the unique YMFM motif recruits a p85a and a p50a subunits of PI3K, resulting in the elevated phosphorylation of Akt, thereby inducing PI3K activity. In contrast, the ICOS-SV, a truncated isoform lacking the YMFM motif in its cytoplasmic tail, cannot elicit phosphorylation of Akt. Consequently, it fails to promote T-cell proliferation. The secreted ICOS-SV (red) competes with membrane-bound ICOS for binding to ICOSL, thereby blocking the interaction between ICOSL and membrane ICOS. As a result, the sICOS-SV suppresses phosphorylation of Akt and T-cell proliferation, leading to the inhibition of T-cell immunity. The diagram was created with BioRender.com. All data shown are representative of at least 2 independent experiments.

Article Snippet: To evaluate soluble ICOS-SVs in the sera of melanoma patients or cell culture supernatants, the Human ICOS DuoSet ELISA kit (DY169, R&D Systems) was used according to the manufacturer's protocol.

Techniques: Transduction, Expressing, Plasmid Preparation, Negative Control, Flow Cytometry, Staining, Control, Binding Assay, Incubation, Two Tailed Test, Standard Deviation, Thymidine Incorporation Assay, Blocking Assay, Phospho-proteomics, Activity Assay, Membrane, Inhibition

Journal: Cancer Immunology Research

Article Title: Granulocyte–Macrophage Colony-Stimulating Factor Influence on Soluble and Membrane-Bound ICOS in Combination with Immune Checkpoint Blockade

doi: 10.1158/2326-6066.CIR-22-0702

Figure Lengend Snippet: sICOS from melanoma patients inhibits T-cell activation and proliferation induced by GM-CSF–driven DCs in MLRs. A, CD4 + T cells (responders) were stimulated by allogeneic GM-CSF–driven MoDCs (stimulators: negative control DCs, DCs generated by GM-CSF/IL4 + anti-IgG, or DCs generated by GM-CSF/IL4 + anti-CD116) in MLRs. T cells were assessed for activation via flow cytometry using anti-ICOS (blue), anti-GITR (green), or anti-CD25 (gray). Isotype control antibodies were used as negative controls. B, Statistical analysis of the percentage of CD4 + ICOS + , CD4 + GITR + , and CD4 + CD25 + T-cell populations from different groups as described above. C, Soluble ICOS levels were determined by ELISA using supernatants from the MLRs described above. D – E, Serum from sICOS-high and sICOS-negative patients were added to the MLRs, and T-cell proliferation ( D ) and CD69 expression ( E ) were evaluated. Additionally, sICOS was depleted from sICOS-high serum to assess its effects on T cells. F, Statistical analysis of the percentage of CD4 + CD69 + T-cell populations from different groups as described above. All data shown are representative of at least 2 independent experiments. Two-tailed unpaired t test was used between the two groups. **, P < 0.01; ***, P < 0.001; ****, P < 0.0001; ns, not significant ( P > 0.05). Standard deviation of the mean (SD) is shown.

Article Snippet: To evaluate soluble ICOS-SVs in the sera of melanoma patients or cell culture supernatants, the Human ICOS DuoSet ELISA kit (DY169, R&D Systems) was used according to the manufacturer's protocol.

Techniques: Activation Assay, Negative Control, Generated, Flow Cytometry, Control, Enzyme-linked Immunosorbent Assay, Expressing, Two Tailed Test, Standard Deviation

Journal: Frontiers in immunology

Article Title: Involvement of CD40-CD40L and ICOS-ICOSL in the development of chronic rhinosinusitis by targeting eosinophils.

doi: 10.3389/fimmu.2023.1171308

Figure Lengend Snippet: FIGURE 1 The expression of CD40, CD40L, ICOS, and ICOSL in nasal tissues of patients with ECRS and non-eCRS. (A) The representative immunohistochemistry stainings of CD40, CD40L, ICOS, and ICOSL. Original magnification, ×400. (B) The mean numbers of CD40+ (non-eCRS, n = 19; ECRS, n = 9), CD40L+ (non-eCRS, n = 15; ECRS, n = 9), ICOS+ (non-eCRS, n = 12; ECRS, n = 8), and ICOSL+ (non-eCRS, n = 15; ECRS, n = 10) cells in nasal tissues.

Article Snippet: Blood was treated with red blood cell lysis buffer and then incubated for 24 h at 37°C with either recombinant human CD40L protein (rhCD40L, 5 μg/ml; R&D Systems, Minneapolis, MN, USA, 6420-CL-025) or recombinant human ICOS protein (rhICOS, 10 μg/ml; R&D Systems, Minneapolis, MN, USA, 169-CS-050).

Techniques: Expressing, Immunohistochemistry

Journal: Frontiers in immunology

Article Title: Involvement of CD40-CD40L and ICOS-ICOSL in the development of chronic rhinosinusitis by targeting eosinophils.

doi: 10.3389/fimmu.2023.1171308

Figure Lengend Snippet: FIGURE 2 The correlation among the levels of CD40, CD40L, ICOS, and ICOSL in nasal tissues of patients with CRS. N = 21, 26, 25, 20, 25, and 21 in (A–F).

Article Snippet: Blood was treated with red blood cell lysis buffer and then incubated for 24 h at 37°C with either recombinant human CD40L protein (rhCD40L, 5 μg/ml; R&D Systems, Minneapolis, MN, USA, 6420-CL-025) or recombinant human ICOS protein (rhICOS, 10 μg/ml; R&D Systems, Minneapolis, MN, USA, 169-CS-050).

Techniques:

Journal: Frontiers in immunology

Article Title: Involvement of CD40-CD40L and ICOS-ICOSL in the development of chronic rhinosinusitis by targeting eosinophils.

doi: 10.3389/fimmu.2023.1171308

Figure Lengend Snippet: FIGURE 4 The correlation among the levels of CD40, CD40L, ICOS, and ICOSL nasal polyp of patients with edematous CRS. N = 13, 15, 15, 13, 15, and 13 in (A–F).

Article Snippet: Blood was treated with red blood cell lysis buffer and then incubated for 24 h at 37°C with either recombinant human CD40L protein (rhCD40L, 5 μg/ml; R&D Systems, Minneapolis, MN, USA, 6420-CL-025) or recombinant human ICOS protein (rhICOS, 10 μg/ml; R&D Systems, Minneapolis, MN, USA, 169-CS-050).

Techniques:

Journal: Frontiers in immunology

Article Title: Involvement of CD40-CD40L and ICOS-ICOSL in the development of chronic rhinosinusitis by targeting eosinophils.

doi: 10.3389/fimmu.2023.1171308

Figure Lengend Snippet: FIGURE 3 The expression of CD40-CD40L and ICOS-ICOSL in different histological patterns of patients with CRS. (A) The percentage of six different histological patterns in ECRS (n = 10) and non-eCRS (n = 21), respectively. (B) The percentage of three histological patterns (edematous: edematous, edematous + fibrotic, and edematous + hyperplasia; fibrotic: fibrotic and edematous + fibrotic; hyperplasia: hyperplasia and edematous + hyperplasia) in ECRS (n = 10) and non-eCRS (n = 21). (C–F) Expression levels of ICOSL (non-edematous, n = 7; edematous, n = 12), ICOS (non- edematous, n = 10; edematous, n = 16), CD40 (non-edematous, n = 13; edematous, n = 15) and CD40L (non-edematous, n = 10; edematous, n = 15) in nasal tissues of patients with edematous and non-edematous CRS.

Article Snippet: Blood was treated with red blood cell lysis buffer and then incubated for 24 h at 37°C with either recombinant human CD40L protein (rhCD40L, 5 μg/ml; R&D Systems, Minneapolis, MN, USA, 6420-CL-025) or recombinant human ICOS protein (rhICOS, 10 μg/ml; R&D Systems, Minneapolis, MN, USA, 169-CS-050).

Techniques: Expressing

Journal: Frontiers in immunology

Article Title: Involvement of CD40-CD40L and ICOS-ICOSL in the development of chronic rhinosinusitis by targeting eosinophils.

doi: 10.3389/fimmu.2023.1171308

Figure Lengend Snippet: FIGURE 6 (A–D) The correlation between expression levels of nasal tissues CD40 (n = 28), CD40L (n = 25), ICOS (n = 21), ICOSL (n = 27), and blood eosinophil count. (E, F) The correlation between expression levels of ICOS (n = 19) and ICOSL (n = 24) in nasal tissues and Lund–Mackay score.

Article Snippet: Blood was treated with red blood cell lysis buffer and then incubated for 24 h at 37°C with either recombinant human CD40L protein (rhCD40L, 5 μg/ml; R&D Systems, Minneapolis, MN, USA, 6420-CL-025) or recombinant human ICOS protein (rhICOS, 10 μg/ml; R&D Systems, Minneapolis, MN, USA, 169-CS-050).

Techniques: Expressing

Journal: Frontiers in immunology

Article Title: Involvement of CD40-CD40L and ICOS-ICOSL in the development of chronic rhinosinusitis by targeting eosinophils.

doi: 10.3389/fimmu.2023.1171308

Figure Lengend Snippet: FIGURE 5 Association between levels of CD40-CD40L, ICOS-ICOSL, and eosinophil in nasal tissues of CRS. (A) The correlation analysis between the number of CD40+ (n = 28), CD40L+ (n = 25), ICOS+ (n = 21), ICOSL+ (n = 27) cells, and tissue eosinophils levels in CRS. (B, C) The co-localization of eosinophils (PRG2, 488) and CD40 (CY3) as well as ICOSL (CY3) assessed by immunofluorescence in patients with ECRS. Original magnification, ×400.

Article Snippet: Blood was treated with red blood cell lysis buffer and then incubated for 24 h at 37°C with either recombinant human CD40L protein (rhCD40L, 5 μg/ml; R&D Systems, Minneapolis, MN, USA, 6420-CL-025) or recombinant human ICOS protein (rhICOS, 10 μg/ml; R&D Systems, Minneapolis, MN, USA, 169-CS-050).

Techniques:

Journal: Frontiers in immunology

Article Title: Involvement of CD40-CD40L and ICOS-ICOSL in the development of chronic rhinosinusitis by targeting eosinophils.

doi: 10.3389/fimmu.2023.1171308

Figure Lengend Snippet: FIGURE 7 The effect of CD40-CD40L and ICOS-ICOSL pathways on eosinophil activation. Peripheral blood of patients with ECRS was stimulated with medium alone (Control, circle, n = 5), IgG (5 µg/ml, rectangle, n = 7), (A) rhCD40L (5 µg/ml, triangle, n = 10) or (B) rhICOS (10 µg/ml, triangle) for 24 h. Then, CD69 expression on eosinophils was detected by flow cytometry. Activated eosinophils were defined as CD45+CD16−CD69+ cells. The percentage of activated eosinophils after different stimulation.

Article Snippet: Blood was treated with red blood cell lysis buffer and then incubated for 24 h at 37°C with either recombinant human CD40L protein (rhCD40L, 5 μg/ml; R&D Systems, Minneapolis, MN, USA, 6420-CL-025) or recombinant human ICOS protein (rhICOS, 10 μg/ml; R&D Systems, Minneapolis, MN, USA, 169-CS-050).

Techniques: Activation Assay, Control, Expressing, Cytometry

Journal: JCI Insight

Article Title: Protective role of tissue-resident Tregs in a murine model of beryllium-induced disease

doi: 10.1172/jci.insight.156098

Figure Lengend Snippet: Frequency (left panel) and number (right panel) of CD4 + CD25 + FoxP3 + T cells in the lungs of HLA-DP2 Tg mice sensitized with 1 (1×) or 3 (3×) doses of BeO and harvested at day 12 ( A ) or sensitized with 1 (1×), 3 (3×), or 7 (7×, sensitization/boost) BeO exposures and examined at day 21 ( B ). ( C ) Representative dot plots show CD103 and CD69 expression on CD4 + CD25 + FoxP3 + Tregs derived from the spleen (top panels) and lung (bottom panels) of mice exposed to BeO on 1 (1×), 3 (3×), or 7 (7×) occasions and examined at day 21. ( D and E ) Number of resident effector (RE, CD103 – CD69 + ) and resident memory (RM, CD103 + CD69 + ) T cells among tissue-resident Tregs in HLA-DP2 Tg mice exposed to 1, 3, and 7 doses of BeO. ( F ) Representative histograms show expression of ICOS, Nrp-1, GITR, and CTLA-4 on tissue-specific CD25 + FoxP3 + CD4 + Tregs in mice exposed to 1 (1×, red) or 3 (3×, green) doses of BeO and analyzed at day 12 or exposed to 1 (1×, blue), 3 (3×, purple), or 7 (7×, brown) doses of BeO and analyzed at day 21. ( G – I ) T cell homeostatic chemokines CXCL-10 ( G ), CCL19 ( H ), and CCL21 ( I ) were measured in the lung tissue lysates by ELISA. Data (mean ± SEM) are representative of 3 individual experiments (2–5 mice per group). Significance was determined by 1-way ANOVA ( A , B , D , and E ) and Mann-Whitney U test ( G – I ). * P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001.

Article Snippet: In ICOS or CTLA-4 blocking experiments, HLA-DP2 Tg mice were injected with 300 μg of an anti–CTLA-4 antibody (BioXcell; clone UC10-4F10-11) or an anti-ICOS antibody (BioXcell; clone 7E.17G9) on day –1.

Techniques: Expressing, Derivative Assay, Enzyme-linked Immunosorbent Assay, MANN-WHITNEY

Journal: JCI Insight

Article Title: Protective role of tissue-resident Tregs in a murine model of beryllium-induced disease

doi: 10.1172/jci.insight.156098

Figure Lengend Snippet: ( A and B ) Plots show the number of CD4 + T cells (left panel) and CD44 + T effector cells (right panel) in the lungs of BeO-sensitized HLA-DP2 Tg mice on day 12 treated i.p. with either an isotype control antibody and anti–CTLA-4 ( A ) or anti-ICOS ( B ) blocking antibodies at day –1. ( C ) Percentage of FoxP3 + Tregs in the lungs of isotype and anti–CTLA-4 (left) or anti-ICOS (right) blocking antibody–treated, BeO-exposed HLA-DP2 Tg mice on day 12. ( D ) At day 12, Teff/Treg ratio was calculated by dividing the total number of tissue-specific Teffs (CD44 + ) by total Tregs (CD25 + FoxP3 + ) in anti–CTLA-4 (left) and anti-ICOS (right) antibody–treated HLA-DP2 Tg mice. ( E ) IFN-γ secretion by CD4 + T cells purified from the lungs of BeO-exposed and anti–CTLA-4 (left) or anti-ICOS (right) blocking antibody–treated HLA-DP2 Tg mice and stimulated with BeSO 4 (100 μM) in the presence of irradiated naive splenocytes. ( F ) Protein in the BALF of mice exposed with BeO (3×) and treated with anti–CTLA-4 (left) or anti-ICOS (right) blocking antibody was examined by ELISA. Data (mean ± SEM) are representative of 2 individual experiments (4–5 mice per group). Significance was determined by Mann-Whitney U test. * P < 0.05.

Article Snippet: In ICOS or CTLA-4 blocking experiments, HLA-DP2 Tg mice were injected with 300 μg of an anti–CTLA-4 antibody (BioXcell; clone UC10-4F10-11) or an anti-ICOS antibody (BioXcell; clone 7E.17G9) on day –1.

Techniques: Control, Blocking Assay, Purification, Irradiation, Enzyme-linked Immunosorbent Assay, MANN-WHITNEY

Journal: Nanomaterials (Basel, Switzerland)

Article Title: Sr-Containing Mesoporous Bioactive Glasses Bio-Functionalized with Recombinant ICOS-Fc: An In Vitro Study.

doi: 10.3390/nano11020321

Figure Lengend Snippet: Figure 1. Schematic illustration of Sr-containing mesoporous bioactive glasses (MBGs) functionalized with ICOS-Fc on osteoblast and osteoclast cells.

Article Snippet: After the indicated time, samples were centrifuged at 10,000 rpm for 5 min, re-suspended in PBS 1X+ 1% NGS and stained with an allophycocyanin (APC)-conjugated anti-human ICOS (R&D System) monoclonal antibody for 20 min at 4 ◦C.

Techniques:

Journal: Nanomaterials (Basel, Switzerland)

Article Title: Sr-Containing Mesoporous Bioactive Glasses Bio-Functionalized with Recombinant ICOS-Fc: An In Vitro Study.

doi: 10.3390/nano11020321

Figure Lengend Snippet: Figure 3. FE-SEM images of SG-Sr (a), SD-Sr (b), SG-Sr-ICOS-Fc (c), and SD-Sr-ICOS-Fc (d).

Article Snippet: After the indicated time, samples were centrifuged at 10,000 rpm for 5 min, re-suspended in PBS 1X+ 1% NGS and stained with an allophycocyanin (APC)-conjugated anti-human ICOS (R&D System) monoclonal antibody for 20 min at 4 ◦C.

Techniques:

Journal: Nanomaterials (Basel, Switzerland)

Article Title: Sr-Containing Mesoporous Bioactive Glasses Bio-Functionalized with Recombinant ICOS-Fc: An In Vitro Study.

doi: 10.3390/nano11020321

Figure Lengend Snippet: Figure 4. N2 adsorption-desorption isotherm of SG-Sr, Amino-SG_Sr and SG-Sr-ICOS-Fc (a), SD-Sr, Amino-SD_Sr and SD-Sr-ICOS-Fc (b). Pore size distribution of SG-Sr, Amino-SG_Sr and SG-Sr-ICOS-Fc (c), SD-Sr, Amino-SD_Sr, and SD-Sr- ICOS-Fc (d).

Article Snippet: After the indicated time, samples were centrifuged at 10,000 rpm for 5 min, re-suspended in PBS 1X+ 1% NGS and stained with an allophycocyanin (APC)-conjugated anti-human ICOS (R&D System) monoclonal antibody for 20 min at 4 ◦C.

Techniques: Adsorption, Pore Size

Journal: Nanomaterials (Basel, Switzerland)

Article Title: Sr-Containing Mesoporous Bioactive Glasses Bio-Functionalized with Recombinant ICOS-Fc: An In Vitro Study.

doi: 10.3390/nano11020321

Figure Lengend Snippet: Figure 6. FESEM images of SG-Sr-ICOS-Fc bioactivity after 1 day (a) and 7 days (b) of soaking in SBF, SD-Sr-ICOS-Fc after 1 day (c) and 7 days (d) of soaking in SBF.

Article Snippet: After the indicated time, samples were centrifuged at 10,000 rpm for 5 min, re-suspended in PBS 1X+ 1% NGS and stained with an allophycocyanin (APC)-conjugated anti-human ICOS (R&D System) monoclonal antibody for 20 min at 4 ◦C.

Techniques:

Journal: Nanomaterials (Basel, Switzerland)

Article Title: Sr-Containing Mesoporous Bioactive Glasses Bio-Functionalized with Recombinant ICOS-Fc: An In Vitro Study.

doi: 10.3390/nano11020321

Figure Lengend Snippet: Figure 7. XRD at different time steps of soaking in SBF of SG-Sr-ICOS-Fc (a) and SD-Sr-ICOS-Fc (b).

Article Snippet: After the indicated time, samples were centrifuged at 10,000 rpm for 5 min, re-suspended in PBS 1X+ 1% NGS and stained with an allophycocyanin (APC)-conjugated anti-human ICOS (R&D System) monoclonal antibody for 20 min at 4 ◦C.

Techniques:

Journal: Nanomaterials (Basel, Switzerland)

Article Title: Sr-Containing Mesoporous Bioactive Glasses Bio-Functionalized with Recombinant ICOS-Fc: An In Vitro Study.

doi: 10.3390/nano11020321

Figure Lengend Snippet: Figure 10. Cytofluorimetric analysis and MFI-R results for SG-Sr-ICOS-Fc, SD-Sr-ICOS-Fc, SG-Sr and SD-Sr. Dot plots and cytofluorimetric histograms of ICOS-Fc for each sample tested are shown. C-: unstained sample; α-ICOS: sample stained with the antibody. FSC: forward scatter; SSC: side scatter.

Article Snippet: After the indicated time, samples were centrifuged at 10,000 rpm for 5 min, re-suspended in PBS 1X+ 1% NGS and stained with an allophycocyanin (APC)-conjugated anti-human ICOS (R&D System) monoclonal antibody for 20 min at 4 ◦C.

Techniques: Staining

Journal: Nanomaterials (Basel, Switzerland)

Article Title: Sr-Containing Mesoporous Bioactive Glasses Bio-Functionalized with Recombinant ICOS-Fc: An In Vitro Study.

doi: 10.3390/nano11020321

Figure Lengend Snippet: Figure 11. ELISA-like assay conducted on SG-Sr-ICOS-Fc and SD-Sr-ICOS-Fc post soaking in DMEM collected at different time steps (3, 7, 14 and 21 days). The presence and the binding of ICOS-Fc were detected by ELISA-like assay using the human ICOSL-His as capture protein and anti-human-Hrpas detection antibody. The graph shows the Optical Density (OD) values (mean and standard error were obtained from three separate samples).

Article Snippet: After the indicated time, samples were centrifuged at 10,000 rpm for 5 min, re-suspended in PBS 1X+ 1% NGS and stained with an allophycocyanin (APC)-conjugated anti-human ICOS (R&D System) monoclonal antibody for 20 min at 4 ◦C.

Techniques: Enzyme-linked Immunosorbent Assay, Binding Assay

Journal: Nanomaterials (Basel, Switzerland)

Article Title: Sr-Containing Mesoporous Bioactive Glasses Bio-Functionalized with Recombinant ICOS-Fc: An In Vitro Study.

doi: 10.3390/nano11020321

Figure Lengend Snippet: Figure 12. Cell viability studies performed on SG-Sr-ICOS-Fc (white bars) and SD-Sr-ICOS-Fc (black bars) samples with MC3T3-E1 cell line at different exposure times (a) 2 d, (b) 4 d, and (c) 7 d considering different particle concentrations. ** p < 0.01 vs. untreated cells (CTR) (Dunnett’ s test). The graphs show cell viability (%) as mean and standard error obtained from three independent experiments. Cell viability was calculated with the following formula: cell viability = absorbance of sample/absorbance of control (untreated cells) × 100.

Article Snippet: After the indicated time, samples were centrifuged at 10,000 rpm for 5 min, re-suspended in PBS 1X+ 1% NGS and stained with an allophycocyanin (APC)-conjugated anti-human ICOS (R&D System) monoclonal antibody for 20 min at 4 ◦C.

Techniques: Control

Journal: Nanomaterials (Basel, Switzerland)

Article Title: Sr-Containing Mesoporous Bioactive Glasses Bio-Functionalized with Recombinant ICOS-Fc: An In Vitro Study.

doi: 10.3390/nano11020321

Figure Lengend Snippet: Figure 14. Clonogenic assay: (a) U2OS; (b) HOS. Cells were treated with ICOS-Fc, SD-Sr-ICOS-Fc, SD-Sr, SG-Sr-ICOS-Fc, and SG-Sr at 2 and 0.2 µg/mL concentration for 72 h. Then, the cell medium was changed, and the cells were cultured for additional 7 days in a free medium (c), (d) Colonies were then photographed. Then, the cells were treated with acetic acid to induce a completely dissolution of the crystal violet and absorbance was evaluated. Three different experiments were performed. Data are shown as mean ± SEM.

Article Snippet: After the indicated time, samples were centrifuged at 10,000 rpm for 5 min, re-suspended in PBS 1X+ 1% NGS and stained with an allophycocyanin (APC)-conjugated anti-human ICOS (R&D System) monoclonal antibody for 20 min at 4 ◦C.

Techniques: Clonogenic Assay, Concentration Assay, Cell Culture, Dissolution

Journal: Cell Reports Medicine

Article Title: Response and recurrence correlates in individuals treated with neoadjuvant anti-PD-1 therapy for resectable oral cavity squamous cell carcinoma

doi: 10.1016/j.xcrm.2021.100411

Figure Lengend Snippet:

Article Snippet: ICOS (C398.4A) , Fluidigm , Cat# 3168024B, RRID: AB_2858237.

Techniques: Recombinant, Isolation, Mass Cytometry, Software

Journal: Cell reports

Article Title: Molecular Signatures of Dengue Virus-Specific IL-10/IFN-γ Co-producing CD4 T Cells and Their Association with Dengue Disease

doi: 10.1016/j.celrep.2019.11.098

Figure Lengend Snippet:

Article Snippet: CD278/ICOS, 148Nd , Fluidigm , Cat#3148019B; Clone C398.4A; RRID: AB_2756435.

Techniques: Recombinant, Staining, Activation Assay, Software

Journal: Cell Reports

Article Title: COX2 regulates senescence secretome composition and senescence surveillance through PGE 2

doi: 10.1016/j.celrep.2021.108860

Figure Lengend Snippet:

Article Snippet: Rat anti-CD278-176Yb , Fluidigm , # 3176014B.

Techniques: Recombinant, Enzyme-linked Immunosorbent Assay, Plasmid Preparation, Software, Sequencing