Human Midkine Antibody

Detection of Human Midkine by Western Blot. Western blot shows lysates of SH-SY5Y human neuroblastoma cell line. PVDF membrane was probed with 2 µg/mL of Goat Anti-Human Midkine Antigen Affinity-purified Polyclonal Antibody (Catalog # AF-258-PB) followed by HRP-conjugated Anti-Goat IgG Secondary Antibody (HAF017). A specific band was detected for Midkine at approximately 17 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.

Midkine in Human Prostate. Midkine was detected in immersion fixed paraffin-embedded sections of human prostate using Goat Anti-Human Midkine Antigen Affinity-purified Polyclonal Antibody (Catalog # AF-258-PB) at 10 µg/mL overnight at 4 °C. Before incubation with the primary antibody, tissue was subjected to heat-induced epitope retrieval using Antigen Retrieval Reagent-Basic (CTS013). Tissue was stained using the Anti-Goat HRP-DAB Cell & Tissue Staining Kit (brown CTS008) and counterstained with hematoxylin (blue). Specific staining was localized to stromal cell cytoplasm. View our protocol for Chromogenic IHC Staining of Paraffin-embedded Tissue Sections.

Midkine in Human Ovarian Array. Midkine was detected in immersion fixed paraffin-embedded sections of human ovarian array using Goat Anti-Human Midkine Antigen Affinity-purified Polyclonal Antibody (Catalog # AF-258-PB) at 10 µg/mL overnight at 4 °C. Tissue was stained using the Anti-Goat HRP-DAB Cell & Tissue Staining Kit (brown; CTS008) and counterstained with hematoxylin (blue). View our protocol for Chromogenic IHC Staining of Paraffin-embedded Tissue Sections.

Detection of Human Midkine by Simple Western^TM. Simple Western lane view shows lysates of SH-SY5Y human neuroblastoma cell line, loaded at 0.2 mg/mL. A specific band was detected for Midkine at approximately 26 kDa (as indicated) using 100 µg/mL of Goat Anti-Human Midkine Antigen Affinity-purified Polyclonal Antibody (Catalog # AF-258-PB) followed by 1:50 dilution of HRP-conjugated Anti-Goat IgG Secondary Antibody (HAF109). This experiment was conducted under reducing conditions and using the 12-230 kDa separation system.

Western Blot Shows Human Midkine Specificity Using Knockout Cell Line. Western blot shows lysates of HAP1 human near-haploid cell line untreated (-) or treated (+) with 3.0 ug/ml Brefeldin A for 18 hr and Midkine knockout HAP1 cell line (KO). Nitrocellulose membrane was probed with 2 µg/mL of Goat Anti-Human Midkine Antigen Affinity-purified Polyclonal Antibody (Catalog # AF-258-PB) followed by HRP-conjugated Anti-Goat IgG Secondary Antibody. A specific band was detected for Midkine at approximately 11 kDa (as indicated) in the parental HAP1 cell line, but is not detectable in knockout HAP1 cell line. The Ponceau stained transfer of the blot is shown. This experiment was conducted under reducing conditions. Image, protocol, and testing courtesy of YCharOS Inc. See ycharos.com for additional details.

Detection of Midkine by Immunoprecipitation. Immunoprecipitation was performed on concentrated culture media of HAP1 human near-haploid cell line using 2.0 μg of Mouse Anti-Human Midkine Monoclonal Antibody (Catalog # AF-258-PB) pre-coupled to protein G or protein A beads. Immunoprecipitated Midkine was detected with Rabbit Anti-Human Midkine Monoclonal Antibody (NBP2-66948 ) at 1/500. The Ponceau stained transfers of each blot are shown. SM=10% starting material; UB=10% unbound fraction; IP=immunoprecipitated. Image, protocol, and testing courtesy of YCharOS Inc. (ycharos.com).