rabbit anti human phospho her2 erbb2 tyr1221 1222  (Cell Signaling Technology Inc)

Journal: iScience

Article Title: From byproduct to biotherapeutic: Comparative study of buffy coats and leukoreduction system chambers for NK cell-based immunotherapies

doi: 10.1016/j.isci.2026.114907

Figure Lengend Snippet: Functional characterization of NK cells (A) Degranulation capacity of NK cells following a 2 h co-culture with K562 leukemia cells, measured by surface CD107a expression ( n = 5). (B and C) Europium-based cytotoxicity assay assessing the specific lysis of K562 target cells at E:T ratios of 10:1, 5:1, 3:1, 1:1, and 0.5:1 on day 7 (B) and day 14 (C) of culture to monitor functional decline over time ( n = 6 for both time points). Here, all comparisons were not significant and are summarized as such (n.s.: p > 0.05). (D) IFN-γ release following 16 h of co-culture of NK cells with K562 cells, quantified via Luminex assay ( n = 5). (E) Kinetic live-cell killing assay using the Incucyte monitoring of Nuclight Red signal loss in K562 cells over 72 h at an E:T ratio of 1:1 ( n = 6). (F) Proportion of donors reaching 25%, 50%, and 75% cumulative lysis thresholds during Incucyte co-culture with K562 cells. (G) CD107a degranulation assay of NK cells with or without pre-incubation with trastuzumab (anti-HER2 antibody) during a 2 h co-culture with MDA-MB-453 cells ( n = 5). (H) Incucyte-based cytotoxicity assay shows the real-time lysis of MDA-MB-453 cells by trastuzumab-loaded NK cells at an E:T ratio of 1:1 over 72 h ( n = 6). (I) Proportion of donors reaching 25%, 50%, and 75% cumulative lysis during the ADCC assay. (n varies as not all samples reached the benchmark values within the observation period). All ADCC experiments have been performed after 14 days of in vitro NK cell expansion. N/A indicates that statistical analysis was not possible for this group. 9J) CD107a degranulation of unmodified NK cells and ErbB2-CAR-NK cells after 2 h co-culture with MDA-MB-453 cells ( n = 5). (K) Incucyte-based cytotoxicity assay showing killing dynamics of ErbB2-CAR-NK cells against MDA-MB-453 cells at an E:T ratio of 1:1 over 72 h ( n = 6). (L) Proportion of donors reaching 25%, 50%, and 75% cumulative lysis during the CAR-mediated cytotoxicity assay. (n varies as not all samples reached the benchmark values within the observation period). All CAR-killing experiments have been performed after 14 days of in vitro NK cell expansion. Unless otherwise stated, n refers to biologically independent donor samples and is indicated per group. Data are shown as mean ± SEM. p-values were determined by Student’s t test with Welsh correction (A, D, F, G, I, J, and L) or by one-way ANOVA (B and C) with Tukey post-test (K and L).

Article Snippet: Mouse Monoclonal anti-CD340 (Her2) , Miltenyi Biotec , Cat# 130-124-474.

Techniques: Functional Assay, Co-Culture Assay, Expressing, Cytotoxicity Assay, Lysis, Luminex, Degranulation Assay, Incubation, ADCC Assay, In Vitro

Journal: Cancers

Article Title: Characterization of HER2-Positive Murine Breast Cancer Models for Investigating HER2-Targeted Therapy and Immunotherapy

doi: 10.3390/cancers18060997

Figure Lengend Snippet: Validation of HER2 expression in the newly established HER2-positive mouse tumors. ( A ) Western blotting analysis of HER2 and phosphor-HER2 in transduced 4T1 and EO771 cell lines. BT474 included as HER2+ control. ( B ) Representative immunofluorescence staining of HER2 expression in transduced EO771 cells. ( C ) Tumor volume curve of 4T1 ( top ) and EO771 ( bottom ) HER2-OE lines (n = 4–5 per group). Data is mean ± SD. ( D ) Representative immunohistochemistry staining of HER2 expression on tumor slices. The uncropped blots are included in .

Article Snippet: Briefly, for IF, cells grown in 96-well plates were fixed with 10% formalin for 30 min at room temperature and blocked with 1% BSA in PBS for 1 h. Cells were incubated overnight at 4 °C with primary rabbit anti-human HER2 antibody (Cell Signaling Technology, 2242S, 1:100 dilution), followed by incubation with Cy5-conjugated goat anti-rabbit secondary antibody (Jackson ImmunoResearch, West Grove, PA, USA, 711-175-152, 1:200) for 1 h at room temperature in the dark.

Techniques: Biomarker Discovery, Expressing, Western Blot, Control, Immunofluorescence, Staining, Immunohistochemistry

Journal: Cancers

Article Title: Characterization of HER2-Positive Murine Breast Cancer Models for Investigating HER2-Targeted Therapy and Immunotherapy

doi: 10.3390/cancers18060997

Figure Lengend Snippet: Trastuzumab alone was not able to reduce primary tumor size but decreased tumor brain metastasis in 4T1-HER2 models. ( A ) Tumor volume curve of 4T1-GFP, -HER2 WT , and -HER2 YVMA tumors treated with either saline or trastuzumab (15 mg/kg, IV). Filled shapes represent saline-treated groups; open shapes represent trastuzumab-treated groups. No statistically significant difference was observed between groups (repeated measures ANOVA p = 0.56). ( B ) Plot of average radiance from GFP fluorescence in the brains of 4T1 tumor-bearing mice. Symbols represent individual mice. ( C ) Fluorescence images of the brains of 4T1 tumor-bearing mice. TRA, trastuzumab. Data is shown as mean ± SD. Two-way ANOVA with Tukey’s HSD test was used in ( B ). *, p < 0.05; **, p < 0.01; ***, p < 0.001.

Article Snippet: Briefly, for IF, cells grown in 96-well plates were fixed with 10% formalin for 30 min at room temperature and blocked with 1% BSA in PBS for 1 h. Cells were incubated overnight at 4 °C with primary rabbit anti-human HER2 antibody (Cell Signaling Technology, 2242S, 1:100 dilution), followed by incubation with Cy5-conjugated goat anti-rabbit secondary antibody (Jackson ImmunoResearch, West Grove, PA, USA, 711-175-152, 1:200) for 1 h at room temperature in the dark.

Techniques: Saline, Fluorescence

Journal: Cancers

Article Title: Characterization of HER2-Positive Murine Breast Cancer Models for Investigating HER2-Targeted Therapy and Immunotherapy

doi: 10.3390/cancers18060997

Figure Lengend Snippet: HER2-targeted therapy synergizes with immunotherapy in HER2-overexpressing (OE) tumors. ( A ) Tumor volume ( left ) and survival ( right ) curves for EO771-HER2 YVMA and 4T1-HER2 YVMA models treated with saline, anti-HER2 agents (TRA + TUC), ICB (αPD-1 and αCTLA-4), or combination therapy (anti-HER2 + ICB) as indicated by arrows. ( B , C ) Flow cytometry quantification of CD4+ ( B ) and CD8+ ( C ) T cells in EO771-HER2 YVMA tumors on day 5. Mantel–Cox logrank test was used for survival analysis in ( A ). Two-way ANOVA with Bonferroni correction for multiple comparisons was used for ( B , C ). All data are shown as mean ± SD; * p < 0.05; ** p < 0.01.

Article Snippet: Briefly, for IF, cells grown in 96-well plates were fixed with 10% formalin for 30 min at room temperature and blocked with 1% BSA in PBS for 1 h. Cells were incubated overnight at 4 °C with primary rabbit anti-human HER2 antibody (Cell Signaling Technology, 2242S, 1:100 dilution), followed by incubation with Cy5-conjugated goat anti-rabbit secondary antibody (Jackson ImmunoResearch, West Grove, PA, USA, 711-175-152, 1:200) for 1 h at room temperature in the dark.

Techniques: Saline, Flow Cytometry

Journal: Cancers

Article Title: Characterization of HER2-Positive Murine Breast Cancer Models for Investigating HER2-Targeted Therapy and Immunotherapy

doi: 10.3390/cancers18060997

Figure Lengend Snippet: HER2-overexpressing models show differential responses to T-DM1 and T-Dxd. Tumor volume curves for BT474 (human HER2+) ( A ), HER2− MDA-MB-231 (human TNBC) ( B ), 4T1-HER2 WT ( C ), and EO771-HER2 WT ( D ) treated with saline (black line), T-DM1 (blue line), or T-Dxd (red line). n = 3 per group. * p < 0.05.

Article Snippet: Briefly, for IF, cells grown in 96-well plates were fixed with 10% formalin for 30 min at room temperature and blocked with 1% BSA in PBS for 1 h. Cells were incubated overnight at 4 °C with primary rabbit anti-human HER2 antibody (Cell Signaling Technology, 2242S, 1:100 dilution), followed by incubation with Cy5-conjugated goat anti-rabbit secondary antibody (Jackson ImmunoResearch, West Grove, PA, USA, 711-175-152, 1:200) for 1 h at room temperature in the dark.

Techniques: Saline