Human MELK Antibody

Detection of Human MELK by Western Blot. Western blot shows lysates of Huh-7 human hepatoma cell line, MO7e human megakaryocytic leukemic cell line, and THP-1 human acute monocytic leukemia cell line. PVDF membrane was probed with 1 µg/mL of Sheep Anti-Human MELK Antigen Affinity-purified Polyclonal Antibody (Catalog # AF4820) followed by HRP-conjugated Anti-Sheep IgG Secondary Antibody (Catalog # HAF016). A specific band was detected for MELK at approximately 74 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.

MELK in Human Breast Cancer Tissue. MELK was detected in immersion fixed paraffin-embedded sections of human breast cancer tissue using Sheep Anti-Human MELK Antigen Affinity-purified Polyclonal Antibody (Catalog # AF4820) at 10 µg/mL overnight at 4 °C. Tissue was stained using the Anti-Sheep HRP-DAB Cell & Tissue Staining Kit (brown; Catalog # CTS019) and counterstained with hematoxylin (blue). Specific staining was localized to plasma membranes of glandular epithelial cells. View our protocol for Chromogenic IHC Staining of Paraffin-embedded Tissue Sections.

Detection of MELK by Western Blot Effect of inhibition of MELK signaling on tumor growth and lung metastasis in TNBC xenograft mouse models. A, MELK expression in Cas9-p15 (P15) and MELK KO MDA-MB-231 (C3 and C28) cells. alpha -Tubulin was used as a loading control. B–F, MELK KO significantly suppressed lung metastases in an MDA-MB-231 xenograft mouse model. Female athymic nude mice were injected intravenously with luciferase-transfected Cas9-p15 or MELK KO MDA-MB-231-Luc-GFP stable cells. Metastatic tumors were measured weekly for 7 weeks by whole-body luciferase imaging using an IVIS 100 Imaging System. Shown are mouse whole-body luciferase images (B), photon counts per area (C), lung weight per mouse measured at week 7 following cell inoculation (D), mouse OS over a period of 80 days following cell inoculation (E), and IHC staining for hematoxylin and eosin (H&E), MELK, proliferation (Ki67), and mesenchymal markers (vimentin and fibronectin; F) in mice inoculated with Cas9-p15 or MELK KO MDA-MB-231 cells. Images were taken under 200 × magnification. G, MELK-In-17 significantly suppressed tumor growth in a 4T1 xenograft mouse model (P < 0.05). Murine 4T1 TNBC cells were injected into the mammary fat pads of female BALB/c mice. When tumor size reached 75–150 mm3, grouped mice were treated with vehicle or MELK-In-17 at 5 or 10 mg/kg via intraperitoneal injections daily for 25 days. In C–G, data are presented as mean ± SD. *, P < 0.05; **, P < 0.01. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/37377604), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of MELK by Western Blot Clinical relevance of MELK in breast cancer and expression of MELK in breast cancer cell lines. A,MELK mRNA levels in HR−HER2+, HR+HER2−, HR+HER2+, and TNBC tumors in the World Consortium IBC dataset, analyzed by multiple comparison using a general linear model. Data are presented as mean ± SD. – represents median, X represents mean. B, IHC images (200× magnification) showing MELK protein expression in normal epithelial tissue and luminal, HER2+, and TNBC breast tumors. Correlation between MELK mRNA levels (<7 or ≥7) and OS, PFS, and DMFS in patients in the overall cohort (C) and among those with TNBC (D), determined by the Kaplan–Meier method. MELK mRNA levels (E) and protein levels (F) in TNBC and non-TNBC cells. Inset, correlation between MELK mRNA and protein levels in TNBC cell lines. In F, alpha -tubulin was used as a loading control. MELK mRNA levels were determined using quantitative reverse transcriptase PCR. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/37377604), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of MELK by Western Blot Effect of inhibition of MELK signaling on tumor growth and lung metastasis in TNBC xenograft mouse models. A, MELK expression in Cas9-p15 (P15) and MELK KO MDA-MB-231 (C3 and C28) cells. alpha -Tubulin was used as a loading control. B–F, MELK KO significantly suppressed lung metastases in an MDA-MB-231 xenograft mouse model. Female athymic nude mice were injected intravenously with luciferase-transfected Cas9-p15 or MELK KO MDA-MB-231-Luc-GFP stable cells. Metastatic tumors were measured weekly for 7 weeks by whole-body luciferase imaging using an IVIS 100 Imaging System. Shown are mouse whole-body luciferase images (B), photon counts per area (C), lung weight per mouse measured at week 7 following cell inoculation (D), mouse OS over a period of 80 days following cell inoculation (E), and IHC staining for hematoxylin and eosin (H&E), MELK, proliferation (Ki67), and mesenchymal markers (vimentin and fibronectin; F) in mice inoculated with Cas9-p15 or MELK KO MDA-MB-231 cells. Images were taken under 200 × magnification. G, MELK-In-17 significantly suppressed tumor growth in a 4T1 xenograft mouse model (P < 0.05). Murine 4T1 TNBC cells were injected into the mammary fat pads of female BALB/c mice. When tumor size reached 75–150 mm3, grouped mice were treated with vehicle or MELK-In-17 at 5 or 10 mg/kg via intraperitoneal injections daily for 25 days. In C–G, data are presented as mean ± SD. *, P < 0.05; **, P < 0.01. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/37377604), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of MELK by Western Blot Clinical relevance of MELK in breast cancer and expression of MELK in breast cancer cell lines. A,MELK mRNA levels in HR−HER2+, HR+HER2−, HR+HER2+, and TNBC tumors in the World Consortium IBC dataset, analyzed by multiple comparison using a general linear model. Data are presented as mean ± SD. – represents median, X represents mean. B, IHC images (200× magnification) showing MELK protein expression in normal epithelial tissue and luminal, HER2+, and TNBC breast tumors. Correlation between MELK mRNA levels (<7 or ≥7) and OS, PFS, and DMFS in patients in the overall cohort (C) and among those with TNBC (D), determined by the Kaplan–Meier method. MELK mRNA levels (E) and protein levels (F) in TNBC and non-TNBC cells. Inset, correlation between MELK mRNA and protein levels in TNBC cell lines. In F, alpha -tubulin was used as a loading control. MELK mRNA levels were determined using quantitative reverse transcriptase PCR. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/37377604), licensed under a CC-BY license. Not internally tested by R&D Systems.