c5ar1 mab 18 41 6 based f ab 2 fragments  (TargetMol)

Journal: Journal of Innate Immunity

Article Title: Functional Analysis of a Novel Complement C5a Receptor 1-Blocking Monoclonal Antibody

doi: 10.1159/000535084

Figure Lengend Snippet: Screening of individual hybridoma supernatants on whole blood. Corresponding immunization peptide regions (1–36, 90–109, 175–188, and 266–282) of individual clones are presented below. Negative controls (IgG1k isotype mAb, IgG2ak isotype mAb) and a positive control (C5aR1 detection: mAb S5/1) were included. Data are presented as fold increase based on the secondary antibody control (secondary control). The experiment was repeated three times with the blood of a different anonymous healthy donor per repeat; data points were acquired in singlets. Data are presented as mean ± SD.

Article Snippet: A volume of 390 μL blood was combined with 45 μL of either DPBS (Merck, cat. no.: D8662), IgG1k isotype mAb (BioLegend, cat. no.: 401408), C5aR1 mAb 18-41-6, C5aR1 mAb 18-41-6-based F(ab’) 2 fragments, or avacopan (TargetMol Chemicals, cat. no.: T8223) in sterile 1.8 mL cryogenic tubes (Thermo Fisher Scientific, cat. no.: 363401) at a final concentration of 400 n m and/or 100 μ m (after addition of bacterial stimulant).

Techniques: Clone Assay, Positive Control, Control

Journal: Journal of Innate Immunity

Article Title: Functional Analysis of a Novel Complement C5a Receptor 1-Blocking Monoclonal Antibody

doi: 10.1159/000535084

Figure Lengend Snippet: Screening of selected, purified C5aR1 mAbs on whole blood. Negative controls (IgG1ak isotype mAb and IgG2ak isotype mAb) and a positive control (C5aR1 detection: mAb S5/1) were included. Data are presented as a fold increase based on the secondary antibody control (secondary control). The experiment was repeated three times with the blood of a different anonymous healthy donor per repeat; data points were acquired in singlets. Data are presented as mean ± SD.

Article Snippet: A volume of 390 μL blood was combined with 45 μL of either DPBS (Merck, cat. no.: D8662), IgG1k isotype mAb (BioLegend, cat. no.: 401408), C5aR1 mAb 18-41-6, C5aR1 mAb 18-41-6-based F(ab’) 2 fragments, or avacopan (TargetMol Chemicals, cat. no.: T8223) in sterile 1.8 mL cryogenic tubes (Thermo Fisher Scientific, cat. no.: 363401) at a final concentration of 400 n m and/or 100 μ m (after addition of bacterial stimulant).

Techniques: Purification, Positive Control, Control

Journal: Journal of Innate Immunity

Article Title: Functional Analysis of a Novel Complement C5a Receptor 1-Blocking Monoclonal Antibody

doi: 10.1159/000535084

Figure Lengend Snippet: Assay of C5aR1 inhibitors on iLite ® C5a Assay Ready Cells, displaying commercial mAb and small-molecule C5aR1 inhibitors ( a ) and in-house C5aR1 mAbs ( b ). Data were normalized to a stimulation control (added C5a) and analyzed by nonlinear curve fitting. Data points were collected in duplicates across multiple assay plates; each assay plate was repeated three times. Data are presented as mean ± SD.

Article Snippet: A volume of 390 μL blood was combined with 45 μL of either DPBS (Merck, cat. no.: D8662), IgG1k isotype mAb (BioLegend, cat. no.: 401408), C5aR1 mAb 18-41-6, C5aR1 mAb 18-41-6-based F(ab’) 2 fragments, or avacopan (TargetMol Chemicals, cat. no.: T8223) in sterile 1.8 mL cryogenic tubes (Thermo Fisher Scientific, cat. no.: 363401) at a final concentration of 400 n m and/or 100 μ m (after addition of bacterial stimulant).

Techniques: Control

Journal: Journal of Innate Immunity

Article Title: Functional Analysis of a Novel Complement C5a Receptor 1-Blocking Monoclonal Antibody

doi: 10.1159/000535084

Figure Lengend Snippet: Assessment of mAb 18-41-6 on non-transfected Flp-In™-CHO cells, C5aR1his-transfected Flp-In™-CHO cells, and C5aR2his-transfected Flp-In™-CHO cells. Negative controls (IgG1k isotype mAb and IgG2ak isotype mAb) and positive controls (C5aR1 detection: mAb S5/1; C5aR2 detection: mAb 1D9-M12) were included. Data are presented as fold change based on secondary antibody control (secondary control). The experiment was repeated three times; data points were obtained in singlets. Data are presented as mean ± SD.

Article Snippet: A volume of 390 μL blood was combined with 45 μL of either DPBS (Merck, cat. no.: D8662), IgG1k isotype mAb (BioLegend, cat. no.: 401408), C5aR1 mAb 18-41-6, C5aR1 mAb 18-41-6-based F(ab’) 2 fragments, or avacopan (TargetMol Chemicals, cat. no.: T8223) in sterile 1.8 mL cryogenic tubes (Thermo Fisher Scientific, cat. no.: 363401) at a final concentration of 400 n m and/or 100 μ m (after addition of bacterial stimulant).

Techniques: Transfection, Control

Journal: Journal of Innate Immunity

Article Title: Functional Analysis of a Novel Complement C5a Receptor 1-Blocking Monoclonal Antibody

doi: 10.1159/000535084

Figure Lengend Snippet: Assay of C5aR1 inhibitors within a C5a-driven PMN calcium flux assay. A negative inhibition control (IgG1k isotype mAb) and a positive inhibition control (20 μ m avacopan) were included. After first recording the baseline, either C5a or buffer was added, followed by the addition of either ionomycin or buffer. Data are presented as fold increase based on the initially recorded baseline (mean FITC detector MFI). The experiment was repeated three times with the blood of a different anonymous healthy donor per repeat; data points were obtained in singlets. Data are presented as mean + SD (one-sided) for better visualization.

Article Snippet: A volume of 390 μL blood was combined with 45 μL of either DPBS (Merck, cat. no.: D8662), IgG1k isotype mAb (BioLegend, cat. no.: 401408), C5aR1 mAb 18-41-6, C5aR1 mAb 18-41-6-based F(ab’) 2 fragments, or avacopan (TargetMol Chemicals, cat. no.: T8223) in sterile 1.8 mL cryogenic tubes (Thermo Fisher Scientific, cat. no.: 363401) at a final concentration of 400 n m and/or 100 μ m (after addition of bacterial stimulant).

Techniques: Calcium Flux Assay, Inhibition, Control

Journal: Journal of Innate Immunity

Article Title: Functional Analysis of a Novel Complement C5a Receptor 1-Blocking Monoclonal Antibody

doi: 10.1159/000535084

Figure Lengend Snippet: Assay of C5aR1 inhibitors on C5a-stimulated PMNs ( a–c ), followed by titration of mAb 18-41-6-based F(ab’) 2 fragments versus equimolar concentrations of avacopan ( d , e ). a–c A negative control for C5aR1 inhibition (IgG1k isotype mAb) and a positive control (75 μ m avacopan) were included. Data points were normalized to a stimulation control sample (T10stim) with added C5a. a A baseline control (without incubation and without stimulation, Tb) and an incubation control (incubation and without stimulation, T10) are depicted. Titration curves of mAb 18-41-6-based F(ab’) 2 fragments and avacopan were analyzed by nonlinear curve fitting. The experiments were repeated three times with the blood of two different anonymous healthy donors ( a–c ) or one different anonymous healthy donor per repeat ( d , e ); data points were acquired in singlets. Data are presented as mean ± SD.

Article Snippet: A volume of 390 μL blood was combined with 45 μL of either DPBS (Merck, cat. no.: D8662), IgG1k isotype mAb (BioLegend, cat. no.: 401408), C5aR1 mAb 18-41-6, C5aR1 mAb 18-41-6-based F(ab’) 2 fragments, or avacopan (TargetMol Chemicals, cat. no.: T8223) in sterile 1.8 mL cryogenic tubes (Thermo Fisher Scientific, cat. no.: 363401) at a final concentration of 400 n m and/or 100 μ m (after addition of bacterial stimulant).

Techniques: Titration, Negative Control, Inhibition, Positive Control, Control, Incubation

Journal: Journal of Innate Immunity

Article Title: Functional Analysis of a Novel Complement C5a Receptor 1-Blocking Monoclonal Antibody

doi: 10.1159/000535084

Figure Lengend Snippet: Assay of C5aR1 inhibitors in whole blood stimulated by HI E. coli . a–d A negative control (IgG1k isotype mAb) and a positive control (100 μ m avacopan) for C5aR1 inhibition were included. Data are normalized to a stimulation control (T15stim) with added HI E. coli . a A baseline control (without incubation and without stimulation, Tb) and an incubation control (incubation and without stimulation, T15) are depicted. e–g Nonlinear curve fitting was employed to analyze mAb 18-41-6-based F(ab’) 2 fragments and avacopan titration curves. Experiments were repeated three times, with the blood of two different anonymous healthy donors ( a–d ) or one different anonymous healthy donor per repeat ( e–g ); data points were obtained in singlets. Data are presented as mean ± SD.

Article Snippet: A volume of 390 μL blood was combined with 45 μL of either DPBS (Merck, cat. no.: D8662), IgG1k isotype mAb (BioLegend, cat. no.: 401408), C5aR1 mAb 18-41-6, C5aR1 mAb 18-41-6-based F(ab’) 2 fragments, or avacopan (TargetMol Chemicals, cat. no.: T8223) in sterile 1.8 mL cryogenic tubes (Thermo Fisher Scientific, cat. no.: 363401) at a final concentration of 400 n m and/or 100 μ m (after addition of bacterial stimulant).

Techniques: Negative Control, Positive Control, Inhibition, Control, Incubation, Titration