mouse anti human c9 ae11 antibody  (Hycult Biotech)

Journal: Scientific Reports

Article Title: Inflammatory transcriptomic signatures in a human cellular NMOSD model reveal upregulation of NF-κB and IL6 pathways

doi: 10.1038/s41598-025-27335-9

Figure Lengend Snippet: Complement component deposition on the surface of different cell lines after application of the monoclonal AQP4 antibody E5415A (10 µg/mL) and active or heat-inactivated human complement. ( A ) Terminal complement complex (TCC) formation on U-87MG-AQP4-ECFP cells. After the treatments, cells were washed with PBS with 10% heat-inactivated FCS, and mouse anti-human C5b-C9 neo (aE11) AlexaFluor594 (Novus Biologicals) was applied with 5 µg/mL to visualize TCC deposition on the cell surface. ( B–D ) Complement component C3/C3b/iC3b deposition. After the treatments, cells were washed with PBS with 10% heat-inactivated FCS, and mouse anti-human/mouse C3/C3b/iC3b purified (6C9) (Cedarlane) was applied with 5 µg/mL, followed by 2 µg/mL goat-anti mouse IgG1 Cross-Adsorbed Secondary Antibody, AlexaFluor594 (Invitrogen, Thermo Fisher Scientific) to visualize opsonization by C3/C3b/iC3b on the cell surface of ( B ) U-87MG-AQP4-ECFP cells, ( C ) U-87MG-ECFP cells and ( D ) primary human astrocytes. Scale bar = 20 μm. AF , Alexa Fluor; AQP4, aquaporin-4; DAPI, 4′,6-diamidino-2-phenylindole; ECFP, enhanced cyan fluorescent protein.

Article Snippet: After the treatments, cells were washed with PBS with 10% heat-inactivated FCS, and mouse anti-human C5b-C9 neo (aE11) AlexaFluor594 (Novus Biologicals) was applied with 5 μg/mL to visualize TCC deposition on the cell surface. ( B–D ) Complement component C3/C3b/iC3b deposition.

Techniques: Purification

Journal: Scientific Reports

Article Title: Inflammatory transcriptomic signatures in a human cellular NMOSD model reveal upregulation of NF-κB and IL6 pathways

doi: 10.1038/s41598-025-27335-9

Figure Lengend Snippet: Immunocytochemistry of U-87MG-AQP4-ECFP cells after treatment with NMOSD patient sera and human complement. U-87MG-AQP4-ECFP cells were treated with 10% NMOSD patient sera (with or without AQP4-IgG) in combination with active or heat-inactivated human complement. Then, the cells were washed with PBS with 10% heat-inactivated fetal calf serum, and immunocytochemistry was performed. ( A ) Terminal complement complex (TCC) staining. The cellular surface was stained for TCC deposition by applying mouse anti-C5b-C9 neo (aE11) AlexaFluor594 (Novus Biologicals) with 5 µg/mL. ( B ) NF-κB component p65 staining. Translocation of p65 from the cytosol into the nucleus was visualized by intracellular immunocytochemistry. Cells were fixed, permeabilized, and blocked. Then, 2 µg/mL rabbit anti-human p65 (D14E12) (Cell Signaling) served as the primary antibody, followed by goat anti-rabbit AlexaFluor647 (Invitrogen, Thermo Fisher Scientific). White arrows indicate p65 translocation into the nucleus. Scale bar = 20 μm. AF, Alexa Fluor; DAPI, 4′,6-diamidino-2-phenylindole; TCC, terminal complement complex.

Article Snippet: After the treatments, cells were washed with PBS with 10% heat-inactivated FCS, and mouse anti-human C5b-C9 neo (aE11) AlexaFluor594 (Novus Biologicals) was applied with 5 μg/mL to visualize TCC deposition on the cell surface. ( B–D ) Complement component C3/C3b/iC3b deposition.

Techniques: Immunocytochemistry, Staining, Translocation Assay