mouse rat wisp 1 quantikine elisa kit  (Bio-Techne corporation)

Journal: bioRxiv

Article Title: Cell Communication Network factor 4 promotes tumor-induced immunosuppression in breast cancer

doi: 10.64898/2026.01.07.698253

Figure Lengend Snippet: CCN4 concentration as quantified by ELISA in media conditioned for 48 hours by WT Py230 and Py8119 cells and two independent CCN4 KO variants for each parental cell line. Cell-free media was used as a negative control and media conditioned by B16F0 cells was used as a positive control. Individual data points represent technical replicates and values reported below quantification threshold are indicated as n.d.

Article Snippet: To measure CCN4 secretion from each WT cell line and KO variant, cells were grown for 48 hours to reach about 80% confluence and the media was filtered for ELISA (enzyme-linked immunosorbent assay) analysis using a Mouse CCN4 Quantikine ELISA Development Kit (R&D Systems, Minneapolis, MN).

Techniques: Concentration Assay, Enzyme-linked Immunosorbent Assay, Negative Control, Positive Control

Journal: bioRxiv

Article Title: Cell Communication Network factor 4 promotes tumor-induced immunosuppression in breast cancer

doi: 10.64898/2026.01.07.698253

Figure Lengend Snippet: Kaplan-Meier time-to-event survival curves for immunocompetent C57BL/6 mice (B and D) and severely immunocompromised NSG mice (A and C) challenged subcutaneously with wild-type (black curves) and two CCN4 KO variants (KO1 in blue and KO2 in dotted red curves) derived from Py8119 (A and B) and Py230 (C and D) cell lines. The tumor size exceeding 100 mm 3 was considered the triggering event. The number of animals that experienced a triggering event among the total number of animals challenged with tumor cells is indicated in the legend. Statistical difference among the three curves was assessed using a Peto & Peto Modification the Gehan-Wilcoxon test with the resulting Chi-squared, degrees of freedom and resulting P-values are indicated. As this was an exploratory study, the p-values indicate the extent to which these three Kaplan-Meier survival curves are different.

Article Snippet: To measure CCN4 secretion from each WT cell line and KO variant, cells were grown for 48 hours to reach about 80% confluence and the media was filtered for ELISA (enzyme-linked immunosorbent assay) analysis using a Mouse CCN4 Quantikine ELISA Development Kit (R&D Systems, Minneapolis, MN).

Techniques: Derivative Assay, Modification

Journal: bioRxiv

Article Title: Cell Communication Network factor 4 promotes tumor-induced immunosuppression in breast cancer

doi: 10.64898/2026.01.07.698253

Figure Lengend Snippet: (A) Box-and-whisker plots for spleen weights obtained from C57BL/6 mice challenged with WT Py230 cells and CCN4 KO variants In a time-matched experimental design with the end point at 69 days (n = 5 for each group). (B) Spleen weights and corresponding excised tumor weights obtained from C57BL/6 mice challenged with WT Py8119 cells and CCN4 KO variants in a size-matched experimental design (n = 5 for each group). As there was no statistical difference between CCN4 KO variants, results were combined for analysis. A homoscedastic two-sided Student’s t-test was used to assess statistical significance.

Article Snippet: To measure CCN4 secretion from each WT cell line and KO variant, cells were grown for 48 hours to reach about 80% confluence and the media was filtered for ELISA (enzyme-linked immunosorbent assay) analysis using a Mouse CCN4 Quantikine ELISA Development Kit (R&D Systems, Minneapolis, MN).

Techniques: Whisker Assay

Journal: bioRxiv

Article Title: Cell Communication Network factor 4 promotes tumor-induced immunosuppression in breast cancer

doi: 10.64898/2026.01.07.698253

Figure Lengend Snippet: Following enzymatic dissociation of tumor tissues, the immune cell compartment was quantified by full spectrum flow cytometry. A self-organizing map shows the distribution of negative Live/Dead violet and positive CD45 cells in size-matched WT (left) and CCN4 KO1 (center) tumors (n = 5 for both WT and CCN4 KO1 Py230). Right panel shows the percentage of events associated with each cluster that relate to cells identified by manual gating.

Article Snippet: To measure CCN4 secretion from each WT cell line and KO variant, cells were grown for 48 hours to reach about 80% confluence and the media was filtered for ELISA (enzyme-linked immunosorbent assay) analysis using a Mouse CCN4 Quantikine ELISA Development Kit (R&D Systems, Minneapolis, MN).

Techniques: Flow Cytometry

Journal: bioRxiv

Article Title: Cell Communication Network factor 4 promotes tumor-induced immunosuppression in breast cancer

doi: 10.64898/2026.01.07.698253

Figure Lengend Snippet: In a size-matched experimental design, the tumor-infiltrating lymphocytes were quantified by flow cytometry using antibody panels focused on the T cell (A, B, and C) and the myeloid (D, E, and F) compartments. Mice received either WT Py8119 cells (black circles) or one of two independently generated CCN4 KO variants (red + and blue x, n = 5 for all groups). Results highlight the CD45+ fraction among total Live events measured using the T cell (A) and myeloid (D) panels. The proportion of infiltrating CD4+ T cells within the live CD45+ compartment (B) was not different but the CD25+ fraction of live CD4+ T cells (C) was increased in CCN4 KO tumors. The MDSC compartment was shifted from PMN-MDSC (E, Live CD45+ CD11b+ Ly6G+ Ly6C int events) to Mo-MDSC (F, Live CD45+ CD11b+ Ly6G-Ly6C+ events) upon CCN4 KO. p-values calculated between WT and pooled CCN4 KOs using two-sided Student’s t test.

Article Snippet: To measure CCN4 secretion from each WT cell line and KO variant, cells were grown for 48 hours to reach about 80% confluence and the media was filtered for ELISA (enzyme-linked immunosorbent assay) analysis using a Mouse CCN4 Quantikine ELISA Development Kit (R&D Systems, Minneapolis, MN).

Techniques: Flow Cytometry, Generated

Journal: bioRxiv

Article Title: Cell Communication Network factor 4 promotes tumor-induced immunosuppression in breast cancer

doi: 10.64898/2026.01.07.698253

Figure Lengend Snippet: Results from R&D Systems’ Mouse XL Cytokine Array Kit of cytokine, chemokine, and growth factor expression by WT versus CCN4 KO cells derived from Py8119 (A) and Py230 (B) lines in vitro . (C) Compares the WT profiles between the Py8119 and Py230 cell lines. In presenting the combined results for assaying each cell line’s secretome using a single biological replicate, open circles represent results for specific cytokine probes, which are spotted in duplicate on the array, and filled circles represent positive and negative controls. Dotted red lines indicate Z-scores of 3 and −3. Particular secreted factors that are differentially expressed are annotated with their respective names.

Article Snippet: To measure CCN4 secretion from each WT cell line and KO variant, cells were grown for 48 hours to reach about 80% confluence and the media was filtered for ELISA (enzyme-linked immunosorbent assay) analysis using a Mouse CCN4 Quantikine ELISA Development Kit (R&D Systems, Minneapolis, MN).

Techniques: Expressing, Derivative Assay, In Vitro

Journal: bioRxiv

Article Title: Cell Communication Network factor 4 promotes tumor-induced immunosuppression in breast cancer

doi: 10.64898/2026.01.07.698253

Figure Lengend Snippet: (A) Projections of WT versus CCN4 KO variants along PC1 and PC2 axes for B16F0, YUMM1.7, Py230, and Py8119 cell lines. The location of duplicates of each cell line label correspond to technical replicates on the array. The percentage of variance explained by each principal component is indicated in the axis label. (B) Projection of the measured proteins based on the loading scores associated with PC1 (y-axis) listed from left to right in alphabetical order. The curves on the right represent the density distribution of the PC1 loading scores (black) and a normal distribution centered at −0.091 with a standard deviation of 0.00074 (red).

Article Snippet: To measure CCN4 secretion from each WT cell line and KO variant, cells were grown for 48 hours to reach about 80% confluence and the media was filtered for ELISA (enzyme-linked immunosorbent assay) analysis using a Mouse CCN4 Quantikine ELISA Development Kit (R&D Systems, Minneapolis, MN).

Techniques: Standard Deviation

Journal: bioRxiv

Article Title: Cell Communication Network factor 4 promotes tumor-induced immunosuppression in breast cancer

doi: 10.64898/2026.01.07.698253

Figure Lengend Snippet: (A) Schematic diagram of the network topology associated with the adherens pathway activity of beta-catenin. Adherens junctions are formed by homotypic interactions between the extracellular cadherin domains of a single pass transmembrane protein E-cadherin (E-cad: blue double oval). The cytoplasmic tail of E-cadherin provides a scaffold for a multi-protein complex that includes beta-catenin ( β -cat: red oval). (B) Upon the proteolytic cleavage of adherens junctions, a cytoplasmic fragment of E-cadherin and associated catenins (tBE) are transported to the cytoplasm. In the cytoplasm, the cytoplasmic fragment of E-cadherin and associated catenins can either enter the nucleus or undergo proteasomal degradation. In the nucleus, this multi-protein complex promotes the transcription and translation of CCN4, beta-catenin, and E-cadherin, among other factors. mRNA is represented by light green parallelogram. Once synthesized, CCN4 (light blue circle) is secreted. Newly synthesized beta-catenin and E-cadherin reform the multi-protein complex and are transported to the cell membrane to re-establish adherens junctions. If no E-cadherin binding sites are present, the multi-protein complex is internalized and degraded. (C) Graphical summary of mutations that include both changes in single nucleotides and in copy numbers for breast invasive carcinoma (TCGA, Cell 2015) samples. Of the 816 samples contained in the dataset obtained through cBioPortal (retrieved 11/10/2025), the queried genes are altered in 336 (41%) of the samples.

Article Snippet: To measure CCN4 secretion from each WT cell line and KO variant, cells were grown for 48 hours to reach about 80% confluence and the media was filtered for ELISA (enzyme-linked immunosorbent assay) analysis using a Mouse CCN4 Quantikine ELISA Development Kit (R&D Systems, Minneapolis, MN).

Techniques: Activity Assay, Synthesized, Membrane, Binding Assay