Human Perforin Antibody

Perforin-1 Antibody in Human Spleen via seqIF™ staining on COMET™​ Perforin-1 was detected in immersion fixed paraffin-embedded sections of human Spleen using Mouse Anti-Human Perforin-1, Monoclonal Antibody (Catalog #MAB8011) at a 1ug/mL concentration at 37 ° Celsius for 4 minutes. Before incubation with the primary antibody, tissue underwent an all-in-one dewaxing and antigen retrieval preprocessing using PreTreatment Module (PT Module) and Dewax and HIER Buffer H (pH 9; Epredia Catalog # TA-999-DHBH). Tissue was stained using the Alexa Fluor™ 647 Goat anti-Mouse IgG Secondary Antibody at 1:200 at 37 ° Celsius for 2 minutes. (Yellow; Lunaphore Catalog # DR647MS) and counterstained with DAPI (blue; Lunaphore Catalog # DR100). Specific staining was localized to the membrane and cytoplasm. Protocol available in COMET™ Panel Builder.

Detection of Human Perforin by Western Blot. Western blot shows lysates of human NK cells and HDLM-2 human Hodgkin's lymphoma cell line. PVDF membrane was probed with 2 µg/mL of Mouse Anti-Human Perforin Monoclonal Antibody (Catalog # MAB8011) followed by HRP-conjugated Anti-Mouse IgG Secondary Antibody (Catalog # HAF018). A specific band was detected for Perforin at approximately 70 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.

Perforin in Human Spleen. Perforin was detected in immersion fixed paraffin-embedded sections of human spleen using Mouse Anti-Human Perforin Monoclonal Antibody (Catalog # MAB8011) at 5 µg/mL for 1 hour at room temperature followed by incubation with the Anti-Mouse IgG VisUCyte™ HRP Polymer Antibody (Catalog # VC001). Before incubation with the primary antibody, tissue was subjected to heat-induced epitope retrieval using Antigen Retrieval Reagent-Basic (Catalog # CTS013). Tissue was stained using DAB (brown) and counterstained with hematoxylin (blue). Specific staining was localized to cell surfaces and cytoplasm in splenocytes. View our protocol for IHC Staining with VisUCyte HRP Polymer Detection Reagents.

Detection of Human Perforin by Simple Western Patient derived NK cells can be expanded in vitro and NKEVs contain protein and nucleic acid cargo reflecting the cells of origin. NK cells were isolated from patient PBMCs via negative selection and EVs were isolated with size exclusion chromatography from the NK cell culture medium (n=20, 10 pre and 10 post) (a). Representative image of cytokine profiling (n=12) of the secretome showed the presence of classic chemokines (CCL1, -5) and cytokines (IFNy, GM-CSF) (b). NKEVs were 100-200 nm in diameter post SEC (n=20) (c) and express canonical EV markers and cytotoxic NK proteins (d). Whole transcriptome sequencing (n=21; 20 patient NKEV and 1 control pool NKEV) found the majority of NKEV RNA cargo is protein coding and long noncoding transcripts (e). Differential gene expression (DE) analysis of LUAD/LUSC NKEVs identified a small number of significantly DE transcripts between LUAD and LUSC, such as ERAP2, which is upregulated in LUAD NKEVs (f). Mass spectrometry proteomics analysis (n=21; 20 patient NKEV and 1 control pool NKEV) of NKEV cargo identified over 4000 proteins (g), and modestly differentially expressed proteins between LUAD and LUSC groups (h). (b) is one representative dot blot, PT007-post and (d) is one representative Western blot, PT007-post. (c) shows NTA analysis for all individual samples in grey, with the mean distribution in red and SEM error bars. Image collected and cropped by CiteAb from the following publication (https://pubmed.ncbi.nlm.nih.gov/40821787/), licensed under a CC-BY license. Not internally tested by R&D Systems.