Journal: International Journal of Molecular Sciences

Article Title: Disseminated Tumor Cells (DTCs) in Patients with Cervical Cancer Reveal Mesenchymal Properties and Potential Therapeutic Targets—A New Perspective?

doi: 10.3390/ijms27114875

Figure Lengend Snippet: Positive and negative control cells for both marker panels. ( A ) HeLa (white arrows): in the first panel positive for cytokeratin (CK), Vimentin (Vim), VEGF and in the second panel positive for p16INK4A, negative for PD-L1 and CD45. ( B ) T98G (red arrows): positive for Vim. ( C ) CaSki (yellow arrows): positive for CK, Vim, VEGF, PD-L1 and p16INK4A. ( D ) MCF-7 (green arrows): positive for CK. All cell lines were negative for CD45. Only hematopoietic cells showed CD45 positive staining. Scale bar = 10 μm. Images were processed using Zeiss ZEN 3.7 software and the figure was assembled using Microsoft PowerPoint.

Article Snippet: In the first round of staining, anti-human Pan-Cytokeratin APC-conjugated REAdye_lease antibody (Cat. No. 130-123-091, Miltenyi Biotech, Bergisch Gladbach, Germany), anti-human vimentin FITC-conjugated REAdye_lease antibody (Cat. No. 130-127-022, Miltenyi Biotech, Bergisch Gladbach, Germany) and anti-human VEGF PE-conjugated REAdye_lease antibody (Cat. No. 130-118-061, Miltenyi Biotech, Bergisch Gladbach, Germany) were applied.

Techniques: Negative Control, Marker, Staining, Software

Journal: International Journal of Molecular Sciences

Article Title: Disseminated Tumor Cells (DTCs) in Patients with Cervical Cancer Reveal Mesenchymal Properties and Potential Therapeutic Targets—A New Perspective?

doi: 10.3390/ijms27114875

Figure Lengend Snippet: Representative images of patient-derived DTCs with three different profiles. ( A ) DTC positive for Vim, VEGF and p16, negative for CK, PD-L1 and CD45. ( B ) CK and Vim positive while negative for VEGF, PD-L1, p16 and CD45. ( C ) DTC positive for Vim and PD-L1, negative for all other markers. Scale bar = 10 μm. Images were processed using Zeiss ZEN 3.7 software and figure was assembled using Microsoft PowerPoint.

Article Snippet: In the first round of staining, anti-human Pan-Cytokeratin APC-conjugated REAdye_lease antibody (Cat. No. 130-123-091, Miltenyi Biotech, Bergisch Gladbach, Germany), anti-human vimentin FITC-conjugated REAdye_lease antibody (Cat. No. 130-127-022, Miltenyi Biotech, Bergisch Gladbach, Germany) and anti-human VEGF PE-conjugated REAdye_lease antibody (Cat. No. 130-118-061, Miltenyi Biotech, Bergisch Gladbach, Germany) were applied.

Techniques: Derivative Assay, Software

Journal: International Journal of Molecular Sciences

Article Title: Disseminated Tumor Cells (DTCs) in Patients with Cervical Cancer Reveal Mesenchymal Properties and Potential Therapeutic Targets—A New Perspective?

doi: 10.3390/ijms27114875

Figure Lengend Snippet: Comparison of p16, VEGF and PD-L1 in matching tumor tissue. In four cases we found VEGF on DTCs but not in matching tumor tissue, indicating a potential therapeutic target. Further, we detected PD-L1 on DTCs in 11 patients that had PD-L1 negative tissue at diagnosis, suggesting a potential discordance between tumor tissue and DTC phenotype. Chart generated using Microsoft Excel.

Article Snippet: In the first round of staining, anti-human Pan-Cytokeratin APC-conjugated REAdye_lease antibody (Cat. No. 130-123-091, Miltenyi Biotech, Bergisch Gladbach, Germany), anti-human vimentin FITC-conjugated REAdye_lease antibody (Cat. No. 130-127-022, Miltenyi Biotech, Bergisch Gladbach, Germany) and anti-human VEGF PE-conjugated REAdye_lease antibody (Cat. No. 130-118-061, Miltenyi Biotech, Bergisch Gladbach, Germany) were applied.

Techniques: Comparison, Biomarker Discovery, Generated

Journal: International Journal of Molecular Sciences

Article Title: Disseminated Tumor Cells (DTCs) in Patients with Cervical Cancer Reveal Mesenchymal Properties and Potential Therapeutic Targets—A New Perspective?

doi: 10.3390/ijms27114875

Figure Lengend Snippet: Study design. At diagnosis a tissue biopsy was obtained to histologically confirm cervical cancer cases. The FFPE tissue samples were stained against p16, PD-L1 and VEGF for comparison with DTC profiles. One day prior to oncologic surgery, blood samples were collected for CTC detection using the standardized CK based brightfield method. Bone marrow aspirates were sampled during surgery and DTCs were analyzed using the CK based and the multi-parameter IF staining method. Post-operative follow-up blood samples were collected within two years after surgery during routine visits if applicable. Clinical Outcome was assessed up to February 2025. Created with BioRender.com.

Article Snippet: In the first round of staining, anti-human Pan-Cytokeratin APC-conjugated REAdye_lease antibody (Cat. No. 130-123-091, Miltenyi Biotech, Bergisch Gladbach, Germany), anti-human vimentin FITC-conjugated REAdye_lease antibody (Cat. No. 130-127-022, Miltenyi Biotech, Bergisch Gladbach, Germany) and anti-human VEGF PE-conjugated REAdye_lease antibody (Cat. No. 130-118-061, Miltenyi Biotech, Bergisch Gladbach, Germany) were applied.

Techniques: Biomarker Discovery, Staining, Comparison

Journal: International Journal of Molecular Sciences

Article Title: Disseminated Tumor Cells (DTCs) in Patients with Cervical Cancer Reveal Mesenchymal Properties and Potential Therapeutic Targets—A New Perspective?

doi: 10.3390/ijms27114875

Figure Lengend Snippet: Sequential multi-parameter immunofluorescent staining. After applying the first antibody panel (CK, Vim and VEGF), the slides were scanned and DTC profiles were assessed. Subsequently, releasable fluorochrome-conjugates were digested and the slides were subjected to the second antibody panel (PD-L1, p16, CD45) followed by scanning and DTC detection. Created with BioRender.com.

Article Snippet: In the first round of staining, anti-human Pan-Cytokeratin APC-conjugated REAdye_lease antibody (Cat. No. 130-123-091, Miltenyi Biotech, Bergisch Gladbach, Germany), anti-human vimentin FITC-conjugated REAdye_lease antibody (Cat. No. 130-127-022, Miltenyi Biotech, Bergisch Gladbach, Germany) and anti-human VEGF PE-conjugated REAdye_lease antibody (Cat. No. 130-118-061, Miltenyi Biotech, Bergisch Gladbach, Germany) were applied.

Techniques: Staining

Journal: International Journal of Molecular Sciences

Article Title: Disseminated Tumor Cells (DTCs) in Patients with Cervical Cancer Reveal Mesenchymal Properties and Potential Therapeutic Targets—A New Perspective?

doi: 10.3390/ijms27114875

Figure Lengend Snippet: Box plot showing the residual fluorescence signal after the release step (%) in 295 CaSki cells following treatment with a VEGF-PE conjugated antibody. The release efficiency was evaluated by signal quantification in the cells before and after treatment with the release reagent. The median fluorescence signal was 24.5% (mean 26.7%), corresponding to a signal reduction of 75.5%. The range was 1.75% to 87.78%. Statistical graph was generated using IBM SPSS Statistics Version 29.0.

Article Snippet: In the first round of staining, anti-human Pan-Cytokeratin APC-conjugated REAdye_lease antibody (Cat. No. 130-123-091, Miltenyi Biotech, Bergisch Gladbach, Germany), anti-human vimentin FITC-conjugated REAdye_lease antibody (Cat. No. 130-127-022, Miltenyi Biotech, Bergisch Gladbach, Germany) and anti-human VEGF PE-conjugated REAdye_lease antibody (Cat. No. 130-118-061, Miltenyi Biotech, Bergisch Gladbach, Germany) were applied.

Techniques: Fluorescence, Generated

Journal: International Journal of Molecular Sciences

Article Title: Disseminated Tumor Cells (DTCs) in Patients with Cervical Cancer Reveal Mesenchymal Properties and Potential Therapeutic Targets—A New Perspective?

doi: 10.3390/ijms27114875

Figure Lengend Snippet: CaSki cells before and after the release step. Representative immunofluorescence image of two CaSki cells spiked into bone marrow cells ( A ) before and ( B ) after the release step. The PE channel shows VEGF staining before staining with CD45, while nuclei were stained with DAPI (blue). Images were processed using Zeiss ZEN software and figure was assembled using Microsoft PowerPoint.

Article Snippet: In the first round of staining, anti-human Pan-Cytokeratin APC-conjugated REAdye_lease antibody (Cat. No. 130-123-091, Miltenyi Biotech, Bergisch Gladbach, Germany), anti-human vimentin FITC-conjugated REAdye_lease antibody (Cat. No. 130-127-022, Miltenyi Biotech, Bergisch Gladbach, Germany) and anti-human VEGF PE-conjugated REAdye_lease antibody (Cat. No. 130-118-061, Miltenyi Biotech, Bergisch Gladbach, Germany) were applied.

Techniques: Immunofluorescence, Staining, Software