ccl5 rabbit primary antibody  (Beijing Solarbio Science)

Journal: Frontiers in Chemistry

Article Title: Activated platelet membrane nanovesicles recruit neutrophils to exert the antitumor efficiency

doi: 10.3389/fchem.2022.955995

Figure Lengend Snippet: Characteristics of PLTm nanovesicles. (A) PLTm nanovesicles observed under TEM, scale bar: 100 nm. (B) TEM size distribution of PLTm nanovesicles. (C) DLS of PLTm nanovesicles. (D) Zeta potential of PLTm nanovesicles. (E) SDS-PAGE protein analysis of PLTs and activated PLTm nanovesicles. M: Marker, P: PLTs, aPNs: activated PLTm nanovesicles. (F) The expression of proteins (P-selectin, ICAM-2 and CCL5) in PLTs and activated PLTm nanovesicles were analyzed by Western blot. aPNs: activated PLTm nanovesicles.

Article Snippet: Bovine serum albumin (BSA) was procured from Yeasen Biological Technology Co., Ltd. BCA Protein Quantitation Kit and Coomassie Blue Fast Staining Solution were from Beyotime Biology Co., Ltd. Polyvinylidene fluoride (PVDF) membrane was produced by Cell Signaling Technology, Co., Ltd. P-selectin rabbit primary antibody was provided by Shanghai Lianshuo Biological Technology Co., Ltd. ICAM-2 (F-5) mouse primary antibody was purchased from Santa Cruz Biotechnology Co., Ltd. CCL5 rabbit primary antibody was provided by Solarbio Biotechnology Co., Ltd. β-Actin (60008-1-Ig) mouse primary antibody was provided by Proteintech Group, Inc. Servicebio Biological Technology Co., Ltd. provided hematoxylin and eosin (HE), Ki-67 immunofluorescence detection kit, and terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) immunofluorescence detection kit.

Techniques: Zeta Potential Analyzer, SDS Page, Marker, Expressing, Western Blot

Journal: International Journal of Biological Sciences

Article Title: Tumor-associated macrophage-derived chemokine CCL5 facilitates the progression and immunosuppressive tumor microenvironment of clear cell renal cell carcinoma

doi: 10.7150/ijbs.74647

Figure Lengend Snippet: The overall depiction of differential expression levels of CCL5 in ccRCC and normal tissues, and its correlation with clinicopathological characteristics and outcomes in TCGA and FUSCC cohorts. ( A ) Representative immunohistochemistry images detecting CCL5 expression in paired ccRCC and normal tissues from FUSCC cohort (n=20). ( B ) Cumulative results of CCL5 IHC score from FUSCC cohort using the Students' t test. ( C ) Differential CCL5 expression in 533 tumor and 72 adjacent normal kidney samples from TCGA cohort using the Students' t test. ( D ) Distribution of paired Tumor/Normal ratio using RT-qPCR analysis from FUSCC cohort. ( E ) Western blotting assay of CCL5 expression in 4 paired fresh ccRCC and para-tumor kidney tissues. ( F ) Differential CCL5 expression in paired tumor, normal and metastatic lymph node (mlymph node) using the Students' t test (n=17). ( G ) Differential CCL5 mRNA expression with clinical AJCC stage and pathological WHO/ISUP grade using one-way ANOV test. ( H ) Multivariate Cox regression analysis enrolling clinicopathological indicators and T/N ratio in predicting overall survival displayed in a forest plot.

Article Snippet: IHC staining was performed to assess the expression levels of CCL5 using primary antibodies against CCL5 (No.36467; CST) and peroxidase-conjugated goat anti-rat IgG as previously described .

Techniques: Quantitative Proteomics, Immunohistochemistry, Expressing, Quantitative RT-PCR, Western Blot

Journal: International Journal of Biological Sciences

Article Title: Tumor-associated macrophage-derived chemokine CCL5 facilitates the progression and immunosuppressive tumor microenvironment of clear cell renal cell carcinoma

doi: 10.7150/ijbs.74647

Figure Lengend Snippet: Prognostic implications of CCL5 expression patterns in large-scale patients with ccRCC from the FUSCC and TCGA cohorts. ( A-B ) Kaplan-Meier and log-rank tests identify predictive value of CCL5 expression in prognostic prognosis in discovery set (n=290) from FUSCC cohort. ( C-D ) Kaplan-Meier and log-rank tests identify prognostic value of CCL5 expression in testing set (n=102) from FUSCC cohort. ( E-F ) Kaplan-Meier and log-rank tests identify prognostic value of CCL5 expression in validation set (n=530) from TCGA cohort.

Article Snippet: IHC staining was performed to assess the expression levels of CCL5 using primary antibodies against CCL5 (No.36467; CST) and peroxidase-conjugated goat anti-rat IgG as previously described .

Techniques: Expressing, Biomarker Discovery

Journal: International Journal of Biological Sciences

Article Title: Tumor-associated macrophage-derived chemokine CCL5 facilitates the progression and immunosuppressive tumor microenvironment of clear cell renal cell carcinoma

doi: 10.7150/ijbs.74647

Figure Lengend Snippet: Inhibition of CCL5 restrains proliferation, migration capacities of ccRCC cells and chemotaxis of macrophages. ( A ) Western blotting assay of CCL5 expression in transfected A498 and 786O cell lines. The data represent three independent experiments. ( B ) Cell viability of negative control and transfected ccRCC cells using CCK-8 assay. ( C ) Effect of CCL5-siRNA transfection on cell migration was determined using wound-healing assay. Histograms and Students's t test were used to evaluate the significance between two groups. ( D ) Correlation of CCL5 and epithelial-mesenchymal transition (EMT)-related markers in 533 patients with ccRCC from TCGA were detected by Spearson's correlation analysis. ( E ) The linear relationship of CCL5 expression with PI3K/AKT signaling pathway and tumor proliferation signature by Spearson's correlation. ( F ) Using Western blotting assay, expression of crucial markers in the EMT and PI3K/AKT signaling pathways were assessed.

Article Snippet: IHC staining was performed to assess the expression levels of CCL5 using primary antibodies against CCL5 (No.36467; CST) and peroxidase-conjugated goat anti-rat IgG as previously described .

Techniques: Inhibition, Migration, Chemotaxis Assay, Western Blot, Expressing, Transfection, Negative Control, CCK-8 Assay, Wound Healing Assay, Protein-Protein interactions

Journal: International Journal of Biological Sciences

Article Title: Tumor-associated macrophage-derived chemokine CCL5 facilitates the progression and immunosuppressive tumor microenvironment of clear cell renal cell carcinoma

doi: 10.7150/ijbs.74647

Figure Lengend Snippet: Inhibition of CCL5 restrains proliferation and chemotaxis of THP1-derived TAMs. ( A ) THP1 cells were induced differentiation into macrophages (M0) using PMA treatment and further transformed into M2 phenotype macrophages (specifically THP1-derived TAMs) induced by IL-4. ( B ) Using Western blotting assay, expression of iNOS, Arg-1, TGF-β, and CCL5 were assessed in PMA induced THP1 cells and THP1-derived TAMs. ( C ) CCK8 assay was supplemented to explore the effect of CCL5 on the proliferation ability of THP1-derived TAMs. ( D ) Representative macrophage migration images of the control group, conditioned medium (CM) of tumor cells group and the CM of CCL5-knockdown ccRCC cells. Histograms and Students's t test were used to evaluate the significance between two groups.

Article Snippet: IHC staining was performed to assess the expression levels of CCL5 using primary antibodies against CCL5 (No.36467; CST) and peroxidase-conjugated goat anti-rat IgG as previously described .

Techniques: Inhibition, Chemotaxis Assay, Derivative Assay, Transformation Assay, Western Blot, Expressing, CCK-8 Assay, Migration, Control, Knockdown

Journal: International Journal of Biological Sciences

Article Title: Tumor-associated macrophage-derived chemokine CCL5 facilitates the progression and immunosuppressive tumor microenvironment of clear cell renal cell carcinoma

doi: 10.7150/ijbs.74647

Figure Lengend Snippet: Perturbation of CCL5 in immune microenvironment of ccRCC and its distribution on TAMs. ( A ) Differential CCL5 expression distinguished the individual with ccRCC into two immunophenotypes using unsupervised clustering xCell algorithm for patients with ccRCC from TCGA database. ( B ) Cumulative results showing subpopulations gated on CCL5 high CD45 + leucocytes from fresh human ccRCC samples (n=20). ( C ) Representative immunohistochemistry images of CCL5 expressed macrophages. ( D ) Co-labeled with macrophage marker CD68 (red) and CCL5 (green) in ccRCC tissues. Nuclei were counterstained blue with DAPI. The right panel was merged by the aforementioned three images. ( E ) Opal multiplex immunofluorescence (mIF) was implemented to determine presence of tertiary lymphoid structures (TLS), abundance of tumor associated lymphocytes (TILs) and other cells, and PD-L1 expression in accordance to CCL5 expression on a multispectral imaging system in ccRCC tissues. ( F ) Histogram showing the aggregation of TILs and presence of TLS in ccRCC samples with different CCL5 expression levels. The bar graphs means the number of cases with TILs aggregation, immature TLS or mature TLS. The percentages written on top of each group means the proportion of this group of samples in the CCL5high or CCL5high group. ( G ) Histogram showing the percent of CD68 + CD163 + macrophage M2 in stromal and tumor samples with different CCL5 expression levels using unpaired t test. ( H ) Histogram showing the percent of PD-L1 + macrophages in ccRCC samples with different CCL5 expression levels using unpaired t test.

Article Snippet: IHC staining was performed to assess the expression levels of CCL5 using primary antibodies against CCL5 (No.36467; CST) and peroxidase-conjugated goat anti-rat IgG as previously described .

Techniques: Expressing, Immunohistochemistry, Labeling, Marker, Multiplex Assay, Immunofluorescence, Imaging

Journal: International Journal of Biological Sciences

Article Title: Tumor-associated macrophage-derived chemokine CCL5 facilitates the progression and immunosuppressive tumor microenvironment of clear cell renal cell carcinoma

doi: 10.7150/ijbs.74647

Figure Lengend Snippet: Intra-tumoral CCL5+ TAMs exposed a distinct subset with pro-tumorigenic exhausted features and impaired total CD8 + T cell function of ccRCC ( A ) We next evaluated the Relationship between transcriptomic CCL5 expression and cells infiltration level in the TME among 33 cancers in the TCGA database using Spearson's correlation analysis. ( B-C ) Representative images and histogram of flow cytometry showed infiltration of CCL5 high CD68 + cells in total CD45 + CD68 + cells in tumor and peritumor tissues (n=15). Data were analyzed using the Students' t test. ( D ) Differential level of immune checkpoints activation, exhaustion signatures and CXCL13 expression were compared based on Cibersort algorithm between CCL5 high and CCL5 low ccRCC samples from TCGA cohort using unpaired t test. ( E ) Differential proliferation biomarker Ki-67 expression in CD8 + T cells within CCL5 + or CCL5 - TAMs ccRCC fresh tumor tissues using flow cytometry analysis and unpaired t test. ( F ) Differential immunity stimulatory effectors in CD8 + T cells within CCL5 + or CCL5 - TAMs ccRCC fresh tumor tissues using flow cytometry analysis and unpaired t test. ( G ) Differential immune checkpoints molecules within CCL5 + or CCL5 - TAMs ccRCC fresh tumor tissues using flow cytometry analysis and unpaired t test.

Article Snippet: IHC staining was performed to assess the expression levels of CCL5 using primary antibodies against CCL5 (No.36467; CST) and peroxidase-conjugated goat anti-rat IgG as previously described .

Techniques: Cell Function Assay, Expressing, Flow Cytometry, Activation Assay, Biomarker Discovery

Journal: International Journal of Biological Sciences

Article Title: Tumor-associated macrophage-derived chemokine CCL5 facilitates the progression and immunosuppressive tumor microenvironment of clear cell renal cell carcinoma

doi: 10.7150/ijbs.74647

Figure Lengend Snippet: Accumulation of CCL5 + TAMs distinguished clinical outcomes in patients with ccRCC. ( A ) The necessary transcription signatures composing CCL5 + TAMs were enrolled in Lasso regression analysis to establish the CCL5 + TAMs model for 530 patients with ccRCC from validation TCGA cohort. ( B-C ) Kaplan-Meier and log-rank tests identify prognostic value of TAMs infiltration alone in validation set (n=530) from TCGA cohort. ( D-E ) Kaplan-Meier and log-rank tests identify prognostic value of CCL5 + TAMs infiltration in validation set (n=530) from TCGA cohort.

Article Snippet: IHC staining was performed to assess the expression levels of CCL5 using primary antibodies against CCL5 (No.36467; CST) and peroxidase-conjugated goat anti-rat IgG as previously described .

Techniques: Biomarker Discovery