monoclonal antibodies for hfpr1  (R&D Systems)

Journal: The Journal of biological chemistry

Article Title: Amyloid beta and its naturally occurring N-terminal variants are potent activators of human and mouse formyl peptide receptor 1.

doi: 10.1016/j.jbc.2022.102642

Figure Lengend Snippet: Figure 4. N-abridged Aβ peptides induce hFPR1-dependent responses in glial U87 cells. A, mean Ca2+ peak responses of U87 cells after stimulation with different concentrations of Aβ11-40 or Aβ17-40. Striped bars indicate the response towards 10 μM of the positive control WKWVm-NH2, light gray and dark gray bars indicate negative controls; n = 3, N = 1. B, comparison of the Ca2+ responses upon stimulation with either 5 μM Aβ11-40 or Aβ17-40 alone (green bars) or in the presence of 10 μM of the competitive FPR-antagonist tBoc2 (black bars); n = 3, N = 1. C, dose-dependent chemotaxis of U87 cells upon stimulation with either Aβ11-40 or Aβ17-40. Each bar represents the number of cells that migrated through a porous membrane towards the respective stimuli. Green bars indicate migration towards N-abridged fragments, striped bars display migration towards the positive control 1 μM fMLF, light gray bars show migration without stimuli, and dark gray bars represent the response to 0.1% DMSO, n = 3, N = 1. D, migration of U87 cells that were either untreated (green bars) or treated with 10 μM tBoc2 (black bars) towards 1 μM of Aβ11-40 or Aβ17-40; n = 3, N = 1. All Error bars, S.D. One-way ANOVA test, Dunnett post hoc test for A and C and t test for B and D; *p ≤0.05; **p ≤0.01; ***p ≤0.001; ns, no significance. Aβ, amyloid beta; FPR, Formyl peptide receptor.

Article Snippet: After blocking with 5% [v/v] FCS in PBS for 30 min at RT, primary antibodies diluted in blocking solution were applied to the cells and incubated over night at 4 C. Hereby, monoclonal antibodies for hFPR1 (R&D Systems, MAB3744, 1 μg/ml), hFPR2 (Santa Cruz Biotechnology, sc-57141, 0.2 μg/ml), and hFPR3 (R&D Systems, MAB3896, 1 μg/ml) were used.

Techniques: Positive Control, Comparison, Chemotaxis Assay, Membrane, Migration

Journal: The Journal of biological chemistry

Article Title: Amyloid beta and its naturally occurring N-terminal variants are potent activators of human and mouse formyl peptide receptor 1.

doi: 10.1016/j.jbc.2022.102642

Figure Lengend Snippet: Figure 5. The activation of U87 cells by N-abridged Aβ peptides depends on hFPR1. A, PCR experiments show that U87 cells contain high mRNA levels of hFPR1 but only low amounts of FPR2 and FPR3. Left: Representative image of gel electrophoresis after reverse transcription polymerase chain reaction with primers for all hFPRs. Right: Quantification of complementary DNA for all FPRs obtained through reverse transcription quantitative polymerase chain reaction, n = 5, N = 2. Details on the primer efficiency, specificity, and linearity are given in Figures S6 and S7. One-way ANOVA test, Dunnett post hoc test. B, left: representative immunocytochemistry staining of U87 cells and transfected HEK293T cells with FPR subtype–specific antibodies (red) and nuclei staining (blue). For visibility, brightness and contrast were adjusted for U87 cells and HEK293T cells differently; for absolute intensity comparison pictures with equal settings for exposure time, brightness and contrast pictures are shown in Figure S5. Scale bars indicate 100 μm. Right: Quantification of the mean FPR staining in U87 cells (green) in comparison to FPR-transfected HEK293T cells (red). Analysis was performed on images acquired with the same settings. n = 2,

Article Snippet: After blocking with 5% [v/v] FCS in PBS for 30 min at RT, primary antibodies diluted in blocking solution were applied to the cells and incubated over night at 4 C. Hereby, monoclonal antibodies for hFPR1 (R&D Systems, MAB3744, 1 μg/ml), hFPR2 (Santa Cruz Biotechnology, sc-57141, 0.2 μg/ml), and hFPR3 (R&D Systems, MAB3896, 1 μg/ml) were used.

Techniques: Activation Assay, Nucleic Acid Electrophoresis, Reverse Transcription, Polymerase Chain Reaction, Real-time Polymerase Chain Reaction, Immunocytochemistry, Staining, Transfection, Comparison