rabbit polyclonal anti cd133  (Proteintech)

Journal: Frontiers in Immunology

Article Title: TREM1 is essential for maintaining stemness of liver cancer stem-like cells in hepatocellular carcinoma

doi: 10.3389/fimmu.2025.1618342

Figure Lengend Snippet: TREM1 promotes tumorigenicity, clonogenic potential and spheroid formation in CD133 + EpCAM + liver cancer stem-like cells. (A) Flow cytometry dot plots reveal significant TREM1 expression in CD133 + EpCAM + cells from Huh7, and HepG2 cell lines and HCC P1 patient sample. (B) RT-PCR analysis shows significant expression of stem cell factors in MACS-purified Huh7 CD133 + EpCAM + cells in comparison to CD133 - EpCAM - fractions (n=3 per group) (C) Flow cytometry dot plots depict reduction in CD133 + Epcam + LCSLCs during TREM1 silencing in Huh7 and HepG2 cells (n=4 per group). (D) RT-PCR analysis of MACS-purified LCSLCs reveals a significant decrease in stem cell factor expression during TREM1 ablation (n=3 per group). (E) Western blot analysis shows significant expression of stem cell proteins in MACS purified Huh7 and HepG2 control CD133 + EpCAM + cells in comparison to TREM1 KO CD133 + EpCAM + cells. (F) Spheroid formation assay shows TREM1 abrogation significantly limits spheroid formation and overall proliferation of LCSLCs. Keyence microscope was used for the acquisition of bright field images. Scale bars = 50 μm. Spheroids were counted using ImageJ (n=3 per group) (G) Colony formation assay demonstrates TREM1-positive LCSLCs form significantly more colonies than their KO counterparts. Representative images show TREM1 KO and Control Huh7 and HepG2 LCSLC colonies stained using crystal violet after 14 days. Data were plotted using GraphPad Prism (n=3 per group). (H) Representative images of tumors from NSG CDX models. 5000 Huh7 CD133 + EpCAM + LCSLCs from Control and TREM1 KO groups were injected subcutaneously (n=6 mice per group). The experiment was independently repeated three times for statistical analysis. ***p<0.001, ****p<0.0001.

Article Snippet: Followed by blocking with SuperBlock blocking buffer (PI37580, Thermo Fisher Scientific) for 1h, membranes were incubated with primary antibodies - TREM1 monoclonal (FERMA548755, Invitrogen,1:1000), CD133 Rabbit polyclonal (A0219, Abclonal, 1:1000), Epcam Rabbit polyclonal (A23654, Abclonal, 1:1000), OCT4 Rabbit polyclonal (A7920, Abclonal, 1:1000), Nanog Rabbit polyclonal (A3232, Abclonal, 1:1000), Sox2 Rabbit polyclonal (A19118, Abclonal, 1:1000), β Actin (MAB8929SP, R&D Systems,1:1000), α tubulin (sc-53029, Santa Cruz Biotechnology, 1:1000) at 4°C overnight.

Techniques: Flow Cytometry, Expressing, Reverse Transcription Polymerase Chain Reaction, Purification, Comparison, Western Blot, Control, Tube Formation Assay, Microscopy, Colony Assay, Staining, Injection

Journal: Frontiers in Immunology

Article Title: TREM1 is essential for maintaining stemness of liver cancer stem-like cells in hepatocellular carcinoma

doi: 10.3389/fimmu.2025.1618342

Figure Lengend Snippet: Bulk RNA sequencing and pathway analysis of CD133 + EpCAM + cells from Huh7 control and TREM1 KO cell lines. (A) Heat map created using K-means clustering of the top 10,000 differentially expressed genes (DEGs) reveals that TREM1 KO in Huh7 LCSLCs significantly impacts cell proliferation pathways. Each sample was analyzed in triplicate. (B) Pathway analysis using Gene Set Enrichment Analysis (GSEA). The lollipop plot highlights the downregulated Gene Ontology (GO) Biological Process pathways in TREM1 KO Huh7 LCSLCs. (C) The lollipop plot shows the GO cellular components inhibited by the TREM1 KO, emphasizing the nuclear structures and complexes impacted. (D) Volcano plot displays the DEGs between Huh7 Ctrl and Huh7 TREM1 KO CD133 + EpCAM + cells. Genes with a log2 fold change > 1 and adjusted p- value < 0.05 are highlighted in red, indicating significant upregulation, while those with a log2 fold change < -1 and adjusted p -value < 0.05 are highlighted in blue, indicating significant downregulation. This plot underscores the downregulation of TREM1 and cancer stem cell-associated genes in the Huh7 CD133 + EpCAM + TREM1 KO cells.

Article Snippet: Followed by blocking with SuperBlock blocking buffer (PI37580, Thermo Fisher Scientific) for 1h, membranes were incubated with primary antibodies - TREM1 monoclonal (FERMA548755, Invitrogen,1:1000), CD133 Rabbit polyclonal (A0219, Abclonal, 1:1000), Epcam Rabbit polyclonal (A23654, Abclonal, 1:1000), OCT4 Rabbit polyclonal (A7920, Abclonal, 1:1000), Nanog Rabbit polyclonal (A3232, Abclonal, 1:1000), Sox2 Rabbit polyclonal (A19118, Abclonal, 1:1000), β Actin (MAB8929SP, R&D Systems,1:1000), α tubulin (sc-53029, Santa Cruz Biotechnology, 1:1000) at 4°C overnight.

Techniques: RNA Sequencing, Control

Journal: Frontiers in Immunology

Article Title: TREM1 is essential for maintaining stemness of liver cancer stem-like cells in hepatocellular carcinoma

doi: 10.3389/fimmu.2025.1618342

Figure Lengend Snippet: Bulk RNA sequencing and pathway analysis of CD133 + EpCAM + cells from HepG2 control and TREM1 KO cell lines. (A) Heat map created using K-means clustering of the top 10,000 differentially expressed genes (DEGs) reveals that TREM1 KO in HepG2 LCSLCs predominantly affects extracellular pathways. Each sample was analyzed in triplicate. (B) Pathway analysis using GSEA. The lollipop plot highlights the downregulated pathways in TREM1 KO HepG2 LCSLCs, focusing on significant extracellular pathways affected by the knockout. (C) The lollipop plot shows the GO cellular components inhibited by the TREM1 KO, emphasizing the extracellular structures and complexes impacted. (D) Volcano plot displays the DEGs between HepG2 Control and HepG2 KO CD133 + EpCAM + cells. Genes with a log2 fold change > 1 and adjusted p -value < 0.05 are highlighted in red, indicating significant upregulation, while those with a log2 fold change < -1 and adjusted p -value < 0.05 are highlighted in blue, indicating significant downregulation. This plot highlights the downregulation of TREM1 and cancer stem cell-associated genes in the HepG2 CD133 + EpCAM + TREM1 KO cells.

Article Snippet: Followed by blocking with SuperBlock blocking buffer (PI37580, Thermo Fisher Scientific) for 1h, membranes were incubated with primary antibodies - TREM1 monoclonal (FERMA548755, Invitrogen,1:1000), CD133 Rabbit polyclonal (A0219, Abclonal, 1:1000), Epcam Rabbit polyclonal (A23654, Abclonal, 1:1000), OCT4 Rabbit polyclonal (A7920, Abclonal, 1:1000), Nanog Rabbit polyclonal (A3232, Abclonal, 1:1000), Sox2 Rabbit polyclonal (A19118, Abclonal, 1:1000), β Actin (MAB8929SP, R&D Systems,1:1000), α tubulin (sc-53029, Santa Cruz Biotechnology, 1:1000) at 4°C overnight.

Techniques: RNA Sequencing, Control, Knock-Out

Journal: Frontiers in Immunology

Article Title: TREM1 is essential for maintaining stemness of liver cancer stem-like cells in hepatocellular carcinoma

doi: 10.3389/fimmu.2025.1618342

Figure Lengend Snippet: TREM1 inhibition via VJDT depletes LCSLCs, reduces tumor size, and decreases spheroid formation. (A) Tumor growth curves for Huh7 vehicle and VJDT treated mice (mean ± SEM, n=5 mice/group). Representative images of tumors from indicated groups on day 22. (B) Flow cytometry analysis shows a significant reduction in CD133 + EpCAM + LCSLCs in VJDT-treated tumors compared to the vehicle (n=4 per group). (C) Western blot analysis of two vehicle-treated and two VJDT-treated tumors shows reduced expression of stem cell-related proteins in VJDT-treated tumors. (D) Representative images from the spheroid formation assay demonstrate reduced spheroid formation following VJDT treatment. Scale bar = 50 µm. Spheroids were counted using ImageJ. **p<0.01, ***p<0.001, ns-not significant.

Article Snippet: Followed by blocking with SuperBlock blocking buffer (PI37580, Thermo Fisher Scientific) for 1h, membranes were incubated with primary antibodies - TREM1 monoclonal (FERMA548755, Invitrogen,1:1000), CD133 Rabbit polyclonal (A0219, Abclonal, 1:1000), Epcam Rabbit polyclonal (A23654, Abclonal, 1:1000), OCT4 Rabbit polyclonal (A7920, Abclonal, 1:1000), Nanog Rabbit polyclonal (A3232, Abclonal, 1:1000), Sox2 Rabbit polyclonal (A19118, Abclonal, 1:1000), β Actin (MAB8929SP, R&D Systems,1:1000), α tubulin (sc-53029, Santa Cruz Biotechnology, 1:1000) at 4°C overnight.

Techniques: Inhibition, Flow Cytometry, Western Blot, Expressing, Tube Formation Assay