monoclonal anti-human/anti-mouse c5a antibodies  (R&D Systems)

Journal: Scientific Reports

Article Title: C5a–C5AR1 axis as a potential trigger of the rupture of intracranial aneurysms

doi: 10.1038/s41598-024-53651-7

Figure Lengend Snippet: The co-expression of C5AR1 and its ligand C5a in intracranial aneurysm lesions. Specimens of intracranial aneurysm (IA) lesions (Aneurysm) or control arterial walls (Superficial temporal artery in ( A ), Anterior cerebral artery-olfactory artery bifurcation in ( B )) from human cases ( A ) or rats ( B ) were harvested and examined in immunohistochemistry. The representative images of immunofluorescent staining of IA lesions for C5AR1 (green), C5a/C5a des-arg (red in ( A )), C5 (red in ( B )), and merged images with nuclear staining by DAPI (blue) are shown. A demagnified image of lower panels in ( B ) is shown on the right. The square in the demagnified image corresponds to the area shown in the magnified images. Scale bar: 50 μm.

Article Snippet: The incubation mixture was then subjected to a western blot analysis using mouse monoclonal anti-human C5a/C5a des-Arg antibody (clone 2952, #HM2079, Hycult Biotech).

Techniques: Expressing, Control, Immunohistochemistry, Staining

Journal: Scientific Reports

Article Title: C5a–C5AR1 axis as a potential trigger of the rupture of intracranial aneurysms

doi: 10.1038/s41598-024-53651-7

Figure Lengend Snippet: Tissue-type Plasminogen Activator as an up-stream factor to produce C5a from C5. ( A ) The expression of tissue-type Plasminogen Activator (tPA) and Plasminogen in intracranial aneurysm (IA) lesions. IA lesions from human cases or induced in rats were harvested subjected to immunohistochemical analyses. The images of immunofluorescent staining of IA lesions for tPA (green), Plasminogen (red), and merged images with nuclear staining by DAPI (blue) are shown. Scale bar: 50 μm. ( B ) Co-expression of tPA with C5a in IA lesions. Human IA lesions were harvested subjected to immunohistochemical analyses. The images of immunofluorescent staining of IA lesions for C5a/C5a des-arg which is the derivative of C5a (green), tPA (red), and merged images with nuclear staining by DAPI (blue) are shown. Scale bar: 50 μm. ( C ) Enzymatic cleavage of C5 into C5a by Plasmin. Recombinant C5 (100 μg/ml) was co-incubated with each dose of recombinant Plasmin as indicated (0–100 μg/ml) for 1.5 h in a cell-free system. The incubation mixture was subjected to western blot analyses. The representative images from the western blot analyses to detect C5 or C5a are shown. M; size marker. ( D ) The alternation in the coagulation-fibrinolysis system in IA lesions of rats. The heat map of genes related with the coagulation-fibrinolysis system from the comprehensive gene expression profile data set (GEO accession: GSE161044) is shown.

Article Snippet: The incubation mixture was then subjected to a western blot analysis using mouse monoclonal anti-human C5a/C5a des-Arg antibody (clone 2952, #HM2079, Hycult Biotech).

Techniques: Expressing, Immunohistochemical staining, Staining, Recombinant, Incubation, Western Blot, Marker, Coagulation

Journal: Scientific Reports

Article Title: C5a–C5AR1 axis as a potential trigger of the rupture of intracranial aneurysms

doi: 10.1038/s41598-024-53651-7

Figure Lengend Snippet: C5a–C5AR1 signaling cascade-dependent induction of pro-inflammatory factors. ( A – C ) The C5a–C5AR1 signaling cascade-dependent induction of pro-inflammatory genes in phorbol myristate acetate-primed cell line from human case with diffuse histiocytic lymphoma, U937 cell line. Cells were treated with recombinant C5a as indicated in the panels (0–10 h in ( A ), 0–100 nM in ( B ), 10 nM and 10 h in ( C ). Some cells were pre-treated with the selective C5AR1 inhibitor, W54011 (300 nM, 30 min) prior to the treatment with recombinant C5a. Total RNA was then purified from treated cells subjected to quantitative RT-PCR analyses to examine the induction of TNF (a gene for TNF-α), IL1B or PTGS2 (a gene for COX-2; cyclooxygenase-2). Data represents as mean ± s.e.m (( A ), n = 4; ( B , C ), n = 6). Statistical analysis was done by the Wilcoxon rank sum test in ( A ), the Kruskal–Wallis test followed by the Steel–Dwass test in ( B , C ). *p < 0.05. ( D ) Co-expression of C5a with pro-inflammatory factors in intracranial aneurysm (IA) lesions. IA lesions induced in rats were harvested subjected to immunohistochemical analyses. The images of immunofluorescent staining of IA lesions for C5ar1 (green), TNF-α (red), IL-1β (red), COX-2 (red), and merged images with nuclear staining by DAPI (blue) are shown. Scale bar: 50 μm.

Article Snippet: The incubation mixture was then subjected to a western blot analysis using mouse monoclonal anti-human C5a/C5a des-Arg antibody (clone 2952, #HM2079, Hycult Biotech).

Techniques: Recombinant, Purification, Quantitative RT-PCR, Expressing, Immunohistochemical staining, Staining