Journal: Cell reports

Article Title: BIRC2 Expression Impairs Anti-Cancer Immunity and Immunotherapy Efficacy.

doi: 10.1016/j.celrep.2020.108073

Figure Lengend Snippet: Figure 4. BIRC2 Knockdown in Melanoma Cells Decreases Tumor Growth and Alters Inflammatory Cell Recruitment to the Tumor Micro- environment (A) B16F10 subclones expressing NTC or BIRC2 shRNA (sh3 or sh4) were implanted subcutaneously in female C57BL/6 mice, and tumor growth was monitored. (B–F) Tumors were harvested on day 35 and the percentage of CD8+/CD44+/CD69+ activated T cells (B), CD11b+/NK1.1+ NK cells (C), CD11b+/CD11c+/F4/80

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies Anti-mouse BIRC2 Novus Biologicals Cat# NB100-56889 Anti-mouse b-Actin Santa Cruz Biotechnology Cat# sc-47778 Anti-mouse CD3 BioLegend Cat# 102102 Anti-mouse CD3 Novus Biologicals Cat# FAB4841G-100 Anti-mouse CD4 Novus Biologicals Cat# FAB554A-100 Anti-mouse CD8A Novus Biologicals Cat# NBP1-49045PE Anti-mouse CD11b Novus Biologicals Cat# NB110-89474AF405 Anti-mouse CD11c Novus Biologicals Cat# NB110-40766AF488 Anti-mouse CD25 Novus Biologicals Cat# NBP2-27425AF488 Anti-mouse CD28 BioLegend Cat# 100223 Anti-mouse CD44 Novus Biologicals Cat# NBP1-47386APC Anti-mouse CD45 Novus Biologicals Cat# NB100-77417AF488 Anti-mouse CD45 Novus Biologicals Cat# NB100-77417AF405 Anti-mouse CD69 Novus Biologicals Cat# NBP1-28011AF488 Anti-mouse CD80 Novus Biologicals Cat# NBP1-43385AF488 Anti-mouse CD314 Novus Biologicals Cat# FAB1547V-100UG Anti-mouse F4/80 Novus Biologicals Cat# NB600-404APC Anti-mouse FoxP3 Novus Biologicals Cat# NB100-39002PE Anti-human HIF-1a Novus Biologicals Cat# NB100-479 Anti-human HIF-1b Novus Biologicals Cat# NB100-124 Anti-human HIF-2a Novus Biologicals Cat# NB100-122 Anti-mouse IFNG Novus Biologicals Cat# IC485V-100UG Anti-mouse Ly6c Novus Biologicals Cat# NBP1-28046AF488 Anti-mouse Ly6g Novus Biologicals Cat# FAB1037A-100 Anti-mouse NK1.1 Novus Biologicals Cat# NB100-77528APC Anti-mouse p50 Novus Biologicals Cat# NBP2-6735 Anti-mouse Rel A Novus Biologicals Cat# NB100-2176 Anti-mouse Rel B Novus Biologicals Cat# NBP2-20123 Anti-mouse a-Tubulin Novus Biologicals Cat# NB600-506 Armenian Hamster IgG, anti-mouse CXCL9 (MIG) Bio X Cell Cat# BE0309 Polyclonal Armenian hamster IgG Bio X Cell Cat# BE0091 Syrian Hamster IgG, anti-mouse CTLA-4 Bio X Cell Cat# BP0131 Rat IgG2a, k, anti-mouse PD-1 (CD279) Bio X Cell Cat# BP0146 Rat IgG2a isotype control Bio X Cell Cat# BE0089 Chemicals, Peptides, and Recombinant Proteins Acriflavine Sigma Aldrich SKU # A8126 TRIzol Reagent Invitrogen Cat# 15596026 Puromycin Dihydrochloride ThermoFisher Cat# A1113803 ECL Prime Western Blotting System GE Healthcare SKU# GERPN2232 PolyJet In Vitro DNA Transfection Reagent Signagen Cat # SL100688 Rabbit anti-mouse IgG-HRP Santa Cruz Biotech Cat# sc-358914 Rabbit IgG HRP Linked Whole Ab GE Healthcare SKU# GENA934 (Continued on next page) e1 Cell Reports 32, 108073, August 25, 2020

Techniques: Knockdown, Expressing, shRNA

Journal: Cell reports

Article Title: BIRC2 Expression Impairs Anti-Cancer Immunity and Immunotherapy Efficacy.

doi: 10.1016/j.celrep.2020.108073

Figure Lengend Snippet: Figure 5. BIRC2 Knockdown in Breast Can- cer Cells Decreases Tumor Growth and Al- ters Inflammatory Cell Recruitment to the Tumor Microenvironment (A) EMT6 subclones expressing NTC or either of two shRNAs targeting BIRC2 (sh4 and sh5) were cultured at 20% O2 and analyzed for expression of BIRC2 protein by immunoblot assay. (B) EMT6 subclones (NTC, sh4, and sh5) were im- planted into the mammary fat pad of female BALB/c mice, and tumor volumes were determined (mean ± SEM; n = 4); *p < 0.05 (Kruskal-Wallis test with Benjamini-Hochberg post-test). (C–F) Tumors were harvested on day 13, and the percentage of CD8+/CD44+/CD69+ activated T cells (C), CD3/NK1.1+ NK cells (D), CD11b+/F4/ 80/CD11c+ DCs (E), and CD11b+/Ly6C+ MDSCs (F) was determined (mean ± SEM; n = 4); *p < 0.05 for the indicated pairs (Kruskal-Wallis test with Benjamini-Hochberg post-test). All immune cell populations were calculated as a percentage of the total number of live cells (based on forward and side scatter). (G) EMT6 subclones were implanted into the mammary fat pad of female SCID mice, and tumor growth was monitored. See also Figure S3B.

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies Anti-mouse BIRC2 Novus Biologicals Cat# NB100-56889 Anti-mouse b-Actin Santa Cruz Biotechnology Cat# sc-47778 Anti-mouse CD3 BioLegend Cat# 102102 Anti-mouse CD3 Novus Biologicals Cat# FAB4841G-100 Anti-mouse CD4 Novus Biologicals Cat# FAB554A-100 Anti-mouse CD8A Novus Biologicals Cat# NBP1-49045PE Anti-mouse CD11b Novus Biologicals Cat# NB110-89474AF405 Anti-mouse CD11c Novus Biologicals Cat# NB110-40766AF488 Anti-mouse CD25 Novus Biologicals Cat# NBP2-27425AF488 Anti-mouse CD28 BioLegend Cat# 100223 Anti-mouse CD44 Novus Biologicals Cat# NBP1-47386APC Anti-mouse CD45 Novus Biologicals Cat# NB100-77417AF488 Anti-mouse CD45 Novus Biologicals Cat# NB100-77417AF405 Anti-mouse CD69 Novus Biologicals Cat# NBP1-28011AF488 Anti-mouse CD80 Novus Biologicals Cat# NBP1-43385AF488 Anti-mouse CD314 Novus Biologicals Cat# FAB1547V-100UG Anti-mouse F4/80 Novus Biologicals Cat# NB600-404APC Anti-mouse FoxP3 Novus Biologicals Cat# NB100-39002PE Anti-human HIF-1a Novus Biologicals Cat# NB100-479 Anti-human HIF-1b Novus Biologicals Cat# NB100-124 Anti-human HIF-2a Novus Biologicals Cat# NB100-122 Anti-mouse IFNG Novus Biologicals Cat# IC485V-100UG Anti-mouse Ly6c Novus Biologicals Cat# NBP1-28046AF488 Anti-mouse Ly6g Novus Biologicals Cat# FAB1037A-100 Anti-mouse NK1.1 Novus Biologicals Cat# NB100-77528APC Anti-mouse p50 Novus Biologicals Cat# NBP2-6735 Anti-mouse Rel A Novus Biologicals Cat# NB100-2176 Anti-mouse Rel B Novus Biologicals Cat# NBP2-20123 Anti-mouse a-Tubulin Novus Biologicals Cat# NB600-506 Armenian Hamster IgG, anti-mouse CXCL9 (MIG) Bio X Cell Cat# BE0309 Polyclonal Armenian hamster IgG Bio X Cell Cat# BE0091 Syrian Hamster IgG, anti-mouse CTLA-4 Bio X Cell Cat# BP0131 Rat IgG2a, k, anti-mouse PD-1 (CD279) Bio X Cell Cat# BP0146 Rat IgG2a isotype control Bio X Cell Cat# BE0089 Chemicals, Peptides, and Recombinant Proteins Acriflavine Sigma Aldrich SKU # A8126 TRIzol Reagent Invitrogen Cat# 15596026 Puromycin Dihydrochloride ThermoFisher Cat# A1113803 ECL Prime Western Blotting System GE Healthcare SKU# GERPN2232 PolyJet In Vitro DNA Transfection Reagent Signagen Cat # SL100688 Rabbit anti-mouse IgG-HRP Santa Cruz Biotech Cat# sc-358914 Rabbit IgG HRP Linked Whole Ab GE Healthcare SKU# GENA934 (Continued on next page) e1 Cell Reports 32, 108073, August 25, 2020

Techniques: Knockdown, Expressing, Cell Culture, Western Blot

Journal: Cell reports

Article Title: BIRC2 Expression Impairs Anti-Cancer Immunity and Immunotherapy Efficacy.

doi: 10.1016/j.celrep.2020.108073

Figure Lengend Snippet: Figure 6. BIRC2 Knockdown in B16F10 Cells Increases Anti-tumor Immunity by Increasing CXCL9 Expression (A) NTC and BIRC2-KD subclones were implanted into C57BL/6 mice. When BIRC2-KD tumors became palpable, mice were treated with anti-CXCL9 or IgG every 3 days. Tumor volumes were determined (mean ± SEM; n = 4); *p < 0.05 (Kruskal-Wallis test with Benjamini-Hochberg post-test). (B–E) Tumors were harvested on day 35, and the percentage of CD8+ T cells (relative to CD45+ population) (B), CD8+/CD44+/CD69+ T cells (C), CD3/NK1.1+ NK cells (D), and CD11b+/CD11c+/F4/80 DCs (E) was determined (mean ± SEM; n = 4); *p < 0.05 (Kruskal-Wallis test with Benjamini-Hochberg post-test). All immune cell populations (except B) were calculated as a percentage of the total live cells (based on forward and side scatter). (F–H) The Pearson correlation test was performed to compare CXCL9 mRNA expression with CD8+ T cell score (F), NK cell score (G), and DC score (H), using TCGA data from 481 human melanomas. See also Figures S3C and S4.

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies Anti-mouse BIRC2 Novus Biologicals Cat# NB100-56889 Anti-mouse b-Actin Santa Cruz Biotechnology Cat# sc-47778 Anti-mouse CD3 BioLegend Cat# 102102 Anti-mouse CD3 Novus Biologicals Cat# FAB4841G-100 Anti-mouse CD4 Novus Biologicals Cat# FAB554A-100 Anti-mouse CD8A Novus Biologicals Cat# NBP1-49045PE Anti-mouse CD11b Novus Biologicals Cat# NB110-89474AF405 Anti-mouse CD11c Novus Biologicals Cat# NB110-40766AF488 Anti-mouse CD25 Novus Biologicals Cat# NBP2-27425AF488 Anti-mouse CD28 BioLegend Cat# 100223 Anti-mouse CD44 Novus Biologicals Cat# NBP1-47386APC Anti-mouse CD45 Novus Biologicals Cat# NB100-77417AF488 Anti-mouse CD45 Novus Biologicals Cat# NB100-77417AF405 Anti-mouse CD69 Novus Biologicals Cat# NBP1-28011AF488 Anti-mouse CD80 Novus Biologicals Cat# NBP1-43385AF488 Anti-mouse CD314 Novus Biologicals Cat# FAB1547V-100UG Anti-mouse F4/80 Novus Biologicals Cat# NB600-404APC Anti-mouse FoxP3 Novus Biologicals Cat# NB100-39002PE Anti-human HIF-1a Novus Biologicals Cat# NB100-479 Anti-human HIF-1b Novus Biologicals Cat# NB100-124 Anti-human HIF-2a Novus Biologicals Cat# NB100-122 Anti-mouse IFNG Novus Biologicals Cat# IC485V-100UG Anti-mouse Ly6c Novus Biologicals Cat# NBP1-28046AF488 Anti-mouse Ly6g Novus Biologicals Cat# FAB1037A-100 Anti-mouse NK1.1 Novus Biologicals Cat# NB100-77528APC Anti-mouse p50 Novus Biologicals Cat# NBP2-6735 Anti-mouse Rel A Novus Biologicals Cat# NB100-2176 Anti-mouse Rel B Novus Biologicals Cat# NBP2-20123 Anti-mouse a-Tubulin Novus Biologicals Cat# NB600-506 Armenian Hamster IgG, anti-mouse CXCL9 (MIG) Bio X Cell Cat# BE0309 Polyclonal Armenian hamster IgG Bio X Cell Cat# BE0091 Syrian Hamster IgG, anti-mouse CTLA-4 Bio X Cell Cat# BP0131 Rat IgG2a, k, anti-mouse PD-1 (CD279) Bio X Cell Cat# BP0146 Rat IgG2a isotype control Bio X Cell Cat# BE0089 Chemicals, Peptides, and Recombinant Proteins Acriflavine Sigma Aldrich SKU # A8126 TRIzol Reagent Invitrogen Cat# 15596026 Puromycin Dihydrochloride ThermoFisher Cat# A1113803 ECL Prime Western Blotting System GE Healthcare SKU# GERPN2232 PolyJet In Vitro DNA Transfection Reagent Signagen Cat # SL100688 Rabbit anti-mouse IgG-HRP Santa Cruz Biotech Cat# sc-358914 Rabbit IgG HRP Linked Whole Ab GE Healthcare SKU# GENA934 (Continued on next page) e1 Cell Reports 32, 108073, August 25, 2020

Techniques: Knockdown, Expressing

Journal: Immunity

Article Title: Notch4 signaling limits regulatory T-cell-mediated tissue repair and promotes severe lung inflammation in viral infections

doi: 10.1016/j.immuni.2021.04.002

Figure Lengend Snippet:

Article Snippet: Antibodies against the following human antigens were used: CD3 (HIT3a, catalog no: 300318, 1:200,Biolegend), CD4 (RPA-T4, catalog no: 300530, 1:200,Biolegend), Foxp3 (PCH-101,catalog no: 48-4776-42,56-4716-41, 1:100 Thermofisher), Helios (22F6, catalog no: 47-9883-42 1:100, Thermofisher), Notch1 (HMN1-519, catalog no: 566023, 1:300, BD PharMingen), Notch2 (HMN2-25, catalog no: 742291, 1:300, BD PharMingen), Notch3 (HMN3-21, catalog no: 744828, 1:300, BD PharMingen), Notch4 (HMN4-2, Catalogue no: 563269, 1:300, BD PharMingen), Yap1 (D8H1X, Catalogue no: 14729S, 1:100, Cell Signaling Technology), beta-Catenin (196624, Catalogue no: IC13292V, 1:200, R&D system), CD25 (BC96, Catalogue no: 51-025-941 1:300, Thermofisher), IL-4 (MP4-25D2, Catalogue no: 500828 1:200, Biolegend), IL-13 (JES10-5A2, Catalogue no: 501912, 1:200, Biolegend), CCR4 (L291H4, Catalogue no: 359417 1:300, Biolegend), CXCR3 (G025H7, Catalogue no: 353708, 1:300 Biolegend), CRTH2 (BM16, Catalogue no: 350108 1:300, Biolegend), CD127 (A019D5, Catalogue no: 351320 1:300, Biolegend), IFNγ (4S.B3, Catalogue no: 560741 1:200, BD Biosciences) and Amphiregulin polyclonal antibody (orb7378 1:200, Biorbyt), CD196 (G034E3, Catalogue no: 353423, 1:300 Biolegend).

Techniques: Recombinant, Adjuvant, Enzyme-linked Immunosorbent Assay, Control, Blocking Assay, Mutagenesis, Software

Journal: Frontiers in veterinary science

Article Title: Allogeneic and xenogeneic lymphoid reconstitution in a RAG2 -/- IL2RG y /- severe combined immunodeficient pig: A preclinical model for intrauterine hematopoietic transplantation.

doi: 10.3389/fvets.2022.965316

Figure Lengend Snippet: FIGURE 2 Lymphoid cell analysis of peripheral blood and lymphoid tissue from immunodeficient DKO pigs. (A) Representative peripheral blood flow cytometry analysis showing CD79α+ cells, CD335+ cells, CD3+ CD4+ cells and CD3+ CD8+ cells for wild type (n = 3) and DKO (n = 4) pigs at 1 week of age. CD79α+, CD335+, CD3+ CD4+, and CD3+ CD8+ are nearly undetectable in the peripheral blood of DKO pigs. (B) Histogram plot with bar of percentage from gated mononuclear cells for CD79α+, CD335+, CD3+, CD3+ CD4+ and CD3+ CD8+ cells. Data represents mean, bars indicate standard deviation, and adjusted p values are represented for Tukey’s multiple comparisons tests one-way ANOVA with a significance value of 0.05. (C) Representative immunohistochemistry (40x magnification) showing spleen staining for CD79α + (B cells), CD335+

Article Snippet: For T-cells, mouse anti pig CD3 epsilon Alexa Fluor 405 labeled (Novus #NBP1-28225AF405), mouse anti pig CD8 PE labeled (BD #559584) and mouse anti pig CD4 PE-Cy7 labeled (BD #561473).

Techniques: Cell Analysis, Cytometry, Standard Deviation, Immunohistochemistry, Staining

Journal: Frontiers in veterinary science

Article Title: Allogeneic and xenogeneic lymphoid reconstitution in a RAG2 -/- IL2RG y /- severe combined immunodeficient pig: A preclinical model for intrauterine hematopoietic transplantation.

doi: 10.3389/fvets.2022.965316

Figure Lengend Snippet: FIGURE 3 In utero allogeneic transplantation of pH2B-eGFP fetal liver cells restored T-cells and TCR-β recombination in the thymus of 1-week-old DKO pigs. (A) Immunofluorescence staining (20x magnification) for CD3 (blue), and pH2B-eGFP (GFP-green). Wild-type thymus contains CD3 positive cells, but not pH2B-eGFP (GFP) positive cells. Thymus of non-injected DKO pigs is negative for both CD3 and pH2B-eGFP cells. The thymus of allografted DKO pigs contains CD3 cells, all overlapping with pH2B-GFP positive cells. (B) flow cytometry plots from the thymi of three allografted DKO pigs, showing expression of CD4 and CD8 (gated on CD3+ cells). Also shown are representative plots of GFP+ CD3+, CD3+CD8+, CD3+CD4+CD8+ (DP) and CD3+CD4+ cells. (C–F) Histogram showing thymic percentage and chimerism of CD3+ cells (C), CD3+CD8+ cells (D), DP cells (E) and CD3+CD4+ cells (F) from thymi of all three DKO allografted pigs, while a wild type (n = 3) serves as positive control. The green histogram represents the levels of pH2B-eGFP chimerism, indicating the percentage of allograft donor-origin cells from the respective population. (G) PCR assay for the identification of rearranged TCR-β locus from thymus gDNA. Non-injected DKO pig thymus served as a negative control and a wild type as the positive control. All three allografted pig thymi display rearranged TCR-β locus.

Article Snippet: For T-cells, mouse anti pig CD3 epsilon Alexa Fluor 405 labeled (Novus #NBP1-28225AF405), mouse anti pig CD8 PE labeled (BD #559584) and mouse anti pig CD4 PE-Cy7 labeled (BD #561473).

Techniques: In Utero, Transplantation Assay, Staining, Injection, Cytometry, Expressing, Positive Control, Negative Control

Journal: Frontiers in veterinary science

Article Title: Allogeneic and xenogeneic lymphoid reconstitution in a RAG2 -/- IL2RG y /- severe combined immunodeficient pig: A preclinical model for intrauterine hematopoietic transplantation.

doi: 10.3389/fvets.2022.965316

Figure Lengend Snippet: FIGURE 6 Cell number and percentage quantification of cord blood flow cytometry data from DKO xenografted pigs. (A–F) Scatter plot showing number of human cells/100 µL and percentage from mononuclear cell (from CD3+ for T subsets) or pig cord blood for human CD45, human CD3, human CD4, human CD8, human double positive (hDP) and human double negative (hDN) T-cells. Human blood served as positive control while the blood of non-injected DKO pigs served as a negative control. The line represents the mean, and adjusted p values are represented for Tukey’s multiple comparisons tests one-way ANOVA with a significance value of P < 0.05. (G,H) Scatter plot showing human CD335 and CD19 percentage from mononuclear cells. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001.

Article Snippet: For T-cells, mouse anti pig CD3 epsilon Alexa Fluor 405 labeled (Novus #NBP1-28225AF405), mouse anti pig CD8 PE labeled (BD #559584) and mouse anti pig CD4 PE-Cy7 labeled (BD #561473).

Techniques: Cytometry, Positive Control, Injection, Negative Control

Journal: Frontiers in veterinary science

Article Title: Allogeneic and xenogeneic lymphoid reconstitution in a RAG2 -/- IL2RG y /- severe combined immunodeficient pig: A preclinical model for intrauterine hematopoietic transplantation.

doi: 10.3389/fvets.2022.965316

Figure Lengend Snippet: FIGURE 9 Sustained human engraftment of the thymus in DKO xenografted pigs. (A) Flow cytometry analysis from xenografted DKO pig thymi for hCD3, hCD4 and hCD8 from birth to 3 weeks of age. In contrast to peripheral blood and spleen, where human cells were undetectable by 2 weeks of age, in the thymus, robust engraftment could still be detected at 3 weeks of age. There were no changes in the distribution of SP hCD4, SP hCD8 with age. (B) Flow cytometry plots of wild-type pig thymi showing pig CD4 and pig CD8 expression in age-matched wild-type pig thymi. (C–F) Scatter plot with individual values for percentages (from CD3+) of pig (WT) and human (xenografted) CD3+ CD8+ (C), CD3+ CD4+ (D), DP (E) and DN (F) from wild type and xenografted thymi, respectively. Line represents the mean, and adjusted p values are represented for unpaired two-sample t-test with a significance value of 0.05. (G) PCR assay with human primers specific for identification of human TREC (∼400 bp). Pig IL2RG amplicon served as internal primer control (∼1 Kb) (Top black arrow). Each PCR reaction contains 50 ng of gDNA. Human peripheral blood mobilized stem cells gDNA served as positive control (+), while gDNA from the thymus of a non-injected DKO pig served as negative control (–). Samples tested include thymus from xenografted pigs (birth, 2 weeks, and 3 weeks), and gDNA from cord blood of xenografted DKO pigs at birth (when human cells were detected via flow cytometry).

Article Snippet: For T-cells, mouse anti pig CD3 epsilon Alexa Fluor 405 labeled (Novus #NBP1-28225AF405), mouse anti pig CD8 PE labeled (BD #559584) and mouse anti pig CD4 PE-Cy7 labeled (BD #561473).

Techniques: Flow Cytometry, Expressing, Amplification, Control, Positive Control, Injection, Negative Control, Cytometry

Journal: Frontiers in veterinary science

Article Title: Allogeneic and xenogeneic lymphoid reconstitution in a RAG2 -/- IL2RG y /- severe combined immunodeficient pig: A preclinical model for intrauterine hematopoietic transplantation.

doi: 10.3389/fvets.2022.965316

Figure Lengend Snippet: FIGURE 10 Postnatal clearance of human cells from the spleen and bone marrow of xenografted DKO pigs. Flow cytometry analysis for the presence of human CD45+, human CD3+, human CD19+ and human CD335 (versus side scatter) in the spleen (A) or bone marrow (B) of xenografted DKO pigs from birth to 3 weeks of age. Human CD45+ and human CD3+ cells were identified in the bone marrow and spleen of a DKO pig at birth but became undetectable between 2 and 3 weeks of age. Few, if any, hCD19 or hCD335 were detected at any stage of analysis. These data parallels that seen in circulating cells. (C) Detection and the overall organization of CD45+ cells in the spleen (20x magnification) of WT (pCD45) (n = 1) and xenografted DKO (hCD45) (n = 1) at day 1, and week 2 (n = 1). Human CD45+ cells were detected at day 1 but not at week 2 in DKO xenografted pigs, consistent with the clearance observed via flow cytometry. Immune-competent WT pig spleen staining with pig-specific CD45 antibodies showed multifocal distribution of pig CD45, as expected. In contrast, xenografted DKO day 1 spleen showed a different pattern, with human CD45 cells surrounding areas that resemble large vascular structures.

Article Snippet: For T-cells, mouse anti pig CD3 epsilon Alexa Fluor 405 labeled (Novus #NBP1-28225AF405), mouse anti pig CD8 PE labeled (BD #559584) and mouse anti pig CD4 PE-Cy7 labeled (BD #561473).

Techniques: Flow Cytometry, Cytometry, Staining

Journal: iScience

Article Title: Nanosphere pharmacodynamics improves safety of immunostimulatory cytokine therapy

doi: 10.1016/j.isci.2024.108836

Figure Lengend Snippet:

Article Snippet: Arginase 1/ARG1/liver Arginase Antibody [Alexa Fluor® 405], polyclonal , Novus , NBP1-32731AF405.

Techniques: Recombinant, Staining, Cell Culture, Red Blood Cell Lysis, Enzyme-linked Immunosorbent Assay, Isolation, Multiplex Assay, Software, Sterility

Journal: Nature Biomedical Engineering

Article Title: Antibody-displaying extracellular vesicles for targeted cancer therapy

doi: 10.1038/s41551-024-01214-6

Figure Lengend Snippet: a , Flow cytometry measurement of uptake by MFI of mNG + EVs (Fc-EVs versus ctrl-EVs versus no EVs) in HER2 positive breast cancer cells (SKBR-3 cells) when incubated with HER2-Ab (trastuzumab), control-Ab (IgG ctrl) or no Ab, showing significant increase in uptake of Fc-EVs by trastuzumab. b , Flow cytometry measurement of uptake (by MFI) of mNG + Fc-EVs decorated with the PD-L1-Ab atezolizumab in PD-L1 expression stimulated (IFNγ) or unstimulated malignant melanoma (B16F10) cells. c , Fluorescence microscopy images of B16F10 cells stained with DAPI (blue), untreated (UT) or treated with mNG + (green) Fc-EVs alone or with the PD-L1-Ab atezolizumab, showing increased uptake of the Fc-EVs when decorated with PD-L1-Ab. Panels a and b are shown as mean ± s.d. n = 3 biological replicates. Statistical significance was calculated using one-way ( b ) or two-way ( a ) ANOVA with Tukey’s post-test compared with each value; P values are indicated above the plots throughout.

Article Snippet: The tissues were processed as described in ref. , with lysed tissue and nLuc + EV input being analysed for nanoluc luminescence or for single-cell suspension for flow cytometry, as described above, but with the HER2 Ab (R&D Systems, FAB9896V-100UG) used for flow cytometry applications.

Techniques: Flow Cytometry, Incubation, Control, Expressing, Fluorescence, Microscopy, Staining

Journal: Nature Biomedical Engineering

Article Title: Antibody-displaying extracellular vesicles for targeted cancer therapy

doi: 10.1038/s41551-024-01214-6

Figure Lengend Snippet: IV injection of Fc-EVs with HER2-Ab (trastuzumab, Fc-EV + HER2-Ab) compared to control-Ab (Fc-EV + IgG-ctrl) in HER2 + breast cancer (SKBR-3) tumour-bearing Swiss nude mice. a , The experimental set-up with inoculation of SKBR-3 cells followed by tumour formation for 2 months before IV injection of nLuc + Fc-EVs with Abs, followed by tissue collection 30 min post injection. b , Fold change in detected EVs (based on luminescence) per gram tumour tissue compared to Fc-EV + IgG-ctrl. c , d , Accumulation of Fc-EV + HER2-Ab (based on luminescence) compared to Fc-EV + IgG-ctrl per gram spleen ( c ) and liver ( d ). All data are shown as mean ± s.d. n = 10 mice. Statistical significance was calculated using two-tailed unpaired t -test analysis compared with each value; P values are indicated above the plots throughout.

Article Snippet: The tissues were processed as described in ref. , with lysed tissue and nLuc + EV input being analysed for nanoluc luminescence or for single-cell suspension for flow cytometry, as described above, but with the HER2 Ab (R&D Systems, FAB9896V-100UG) used for flow cytometry applications.

Techniques: IV Injection, Control, Injection, Two Tailed Test

Journal: Translational Oncology

Article Title: HER2 as a potential therapeutic target on quiescent prostate cancer cells

doi: 10.1016/j.tranon.2023.101642

Figure Lengend Snippet: Impact of T-DM1 treatment on metastasis free survival in a prostate cancer left ventricle injection xenograft model. (A) Experimental design for the in vivo experiment. GFP/luciferase-expressing PC3 cells were injected into male SCID mice by left ventricle intracardiac (i.c.) injection. Human IgG (15 mg/kg) or T-DM1 (15 mg/kg) were injected by intraperitoneal (i.p.) injection every 3 days until 12 days after tumor injection IgG: n = 12. T-DM1: n = 14. (B) Kaplan-Meier analysis of time to formation of metastases visible by bioluminescence imaging or death. (C) Representative bioluminescence images of control mice (human IgG injected) and T-DM1 treated mice 124 days after tumor injection. (D) Antibody dependent cellular cytotoxicity (ADCC) assays using flow cytometry. PC3 cells were labeled with DiD for subsequent identification, and were treated with T-DM1 before addition of splenocytes from SCID mice. PC3 cells were identified as DiD positive and non-viable cells were identified as positive for DAPI. (E) Dead PC3 cells (%) quantified from data in panel (D) Data represent mean ± S.D. (N=3).

Article Snippet: Monoclonal mouse IgG2B (R&D systems, Cat #: IC0041V) was used as isotype control.

Techniques: Injection, In Vivo, Luciferase, Expressing, Imaging, Control, Flow Cytometry, Labeling