Journal: BMC Immunology

Article Title: Natural killer cells are crucial for the efficacy of Icon (factor VII/human IgG1 Fc) immunotherapy in human tongue cancer

doi: 10.1186/1471-2172-11-49

Figure Lengend Snippet: Lysis of TCA8113 cancer cells by Icon-dependent NK cell-mediated ADCC and complement-mediated CDC in vitro . a . Isolation of murine NK cells from a single cell suspension of splenocytes from SCID/CB-17 mice. b . Icon-dependent NK cell-mediated ADCC. c . Icon-dependent complement-mediated cytotoxicity. Note that % specific lysis was derived from the % of cytotoxicity of Icon + NK cell or Icon + complement subtracted by % of cytotoxicity of NK cell alone or complement alone. P values and no significance (ns) were analysed by unpaired t test (b) or one way ANOVA with Tukey's Multiple Comparison Test (c). Data are representative of two experiments.

Article Snippet: 1:4 diluted rabbit complement MA (Cedarlane Laboratories) in HBSS was added to each well that had been incubated with mouse Icon protein (Icon-CDC experimental wells) or HBSS buffer (complement alone wells) and incubated at 37°C for 60 min. For the ADCC assay, freshly isolated mouse NK and non-NK cells were added to the plates and incubated at 37°C for 4 hrs.

Techniques: Lysis, In Vitro, Isolation, Suspension, Flow Cytometry Complement-Mediated Cytotoxicity Assay, Derivative Assay, Comparison

Journal: BMC Immunology

Article Title: Natural killer cells are crucial for the efficacy of Icon (factor VII/human IgG1 Fc) immunotherapy in human tongue cancer

doi: 10.1186/1471-2172-11-49

Figure Lengend Snippet: Icon protein has no effect on cancer cell proliferation in the absence of NK cell and complement . After incubation with mouse Icon protein for 1 hr followed by overnight growth, cancer cell proliferation was determined by CellTiter One Solution reagent, as described in Methods. P values obtained were of no significance as determined by one-way ANOVA with Tukey's Multiple Comparison Test. Data are representative of two experiments.

Article Snippet: 1:4 diluted rabbit complement MA (Cedarlane Laboratories) in HBSS was added to each well that had been incubated with mouse Icon protein (Icon-CDC experimental wells) or HBSS buffer (complement alone wells) and incubated at 37°C for 60 min. For the ADCC assay, freshly isolated mouse NK and non-NK cells were added to the plates and incubated at 37°C for 4 hrs.

Techniques: Incubation, Comparison

Journal: Scientific reports

Article Title: A nonadjuvanted HLA-restricted peptide vaccine induced both T and B cell immunity against SARS-CoV-2 spike protein.

doi: 10.1038/s41598-024-71663-1

Figure Lengend Snippet: Fig. 4. In vivo immune responses of injected UC peptides and complement activity of immunized serum. (a) BALB/c mice were intramuscularly administered 3 doses of 50 µg of UC-100 (n = 3) or 400 µg of UC-152 (n = 4) 14 days apart. (b–d) The serum of UC-100 mice was collected pre-(Day-1) and posttreatment (Day41) and tested in duplicate and represented as the mean for the levels of (b) total IgG; (c) specific IgG against UC-100 peptide; (d) specific IgG against SARS-CoV-2 spike trimer protein. (e–g) The serum of UC-152 mice was collected pre-(Day-1) and posttreatment (Day41) and tested in duplicate and represented as the mean for the levels of (e) total IgG; (f) specific IgG against UC-152 peptide; (g) specific IgG against SARS-CoV-2 spike trimer protein. (h,i) Antibody-mediated complement activity was tested in duplicate and is represented as the mean for pre- and postserum of mice vaccinated with UC-100 and UC-152 peptides, respectively. The complement activity was expressed as the IC50 of cytotoxicity of the (h) UC-100-vaccinated group and (i) UC-152-vaccinated group. Significant differences are indicated by asterisks (unpaired one-tailed Student’s t test; *p < 0.05, **p < 0.01, ***p < 0.001).

Article Snippet: A final concentration of 10% young rabbit complement (CL3441-S50-R, Cedarlane, Ontario, Canada) was added to each well and incubated for an additional 30 min at 37 °C and 5% CO2.

Techniques: In Vivo, Injection, Activity Assay, One-tailed Test

Journal: Scientific reports

Article Title: A nonadjuvanted HLA-restricted peptide vaccine induced both T and B cell immunity against SARS-CoV-2 spike protein.

doi: 10.1038/s41598-024-71663-1

Figure Lengend Snippet: Fig. 5. In vivo immune responses of oral UC peptides and complement activity of immunized serum. (a) BALB/c mice (n = 5) were orally administered 3 doses of 200 µg UC-100 14 days apart. (b–d) The serum of UC-100 mice was collected pre-(Day-1) and posttreatment (Day 41) and tested in duplicate and represented as the mean for the levels of (b) total IgG; (c) specific IgG against UC-100 peptide; (d) specific IgG against SARS-CoV-2 spike trimer protein; (e) The antibody-mediated complement activity was tested in duplicate and represented as the mean for pre- and postserum of mice vaccinated with UC-100. The complement activity was expressed as the IC50 of cytotoxicity. (f,g) The serum of UC-100 mice was collected pre-(Day-1) and posttreatment (Day 41) and tested in duplicate and represented as the mean for the levels of (f) total IgA and (g) specific IgA against UC-100 peptide. Significant differences are indicated by asterisks (unpaired one-tailed Student’s t test; *p < 0.05, **p < 0.01, ***p < 0.001).

Article Snippet: A final concentration of 10% young rabbit complement (CL3441-S50-R, Cedarlane, Ontario, Canada) was added to each well and incubated for an additional 30 min at 37 °C and 5% CO2.

Techniques: In Vivo, Activity Assay, One-tailed Test

Journal: Scientific reports

Article Title: A nonadjuvanted HLA-restricted peptide vaccine induced both T and B cell immunity against SARS-CoV-2 spike protein.

doi: 10.1038/s41598-024-71663-1

Figure Lengend Snippet: Fig. 6. Summary of the SARS-CoV-2 HLA-restricted peptide T-cell vaccine. The possible mechanistic immune response to HLA-restricted peptides has been elucidated. The 9-mer UC peptides can be primed onto both HLA class I and II molecules, and the immune response is triggered for both humoral and cellular immunity. Through HLA class I molecule binding, CTLs can be activated, and an increase in IFN-γ cytokines and specific lytic activity have been observed. Through HLA class II molecule binding, B-cells are activated, and specific antibodies are produced in accordance with complement cytotoxicity activity.

Article Snippet: A final concentration of 10% young rabbit complement (CL3441-S50-R, Cedarlane, Ontario, Canada) was added to each well and incubated for an additional 30 min at 37 °C and 5% CO2.

Techniques: Binding Assay, Activity Assay, Produced

Journal: Nature Communications

Article Title: Helicobacter pylori initiates successful gastric colonization by utilizing L-lactate to promote complement resistance

doi: 10.1038/s41467-023-37160-1

Figure Lengend Snippet: a Schematic diagram of C4 primary activation and secondary degradation, adapted from . b Western blot of C4 protein and products after activation in the presence of WT and ∆ lctP H. pylori over 60 min, using non-reducing gels. A positive control of complement activation and C4c formation was triggered by heated human IgG. Positions of C4, C4b, and C4c are shown at the left, and molecular weight is indicated on the right. The result is representative of three independent experiments with triplicate biological samples. c H. pylori PMSS1 WT was grown ± L-lactate and then treated by 10% NHS for the indicated time periods. Each sample was split into three: (1) treated by sample buffer only (total C4b); (2) centrifuged at 3000 × g for 3 min before adding sample buffer (bound C4b); (3) treated by sample buffer plus the reducing agent beta-mercaptoethanol (BME) (C4 β-chain). All protein samples were run on 5-7% SDS PAGE and blotted with anti-C4 antibody. d Quantification of the experiment shown in (c), from three independent experiments. Bands were quantified by normalizing each band to the amount of total C4b or bound C4b at 3 min (leftmost band in each treatment group) using the Bio-Rad Image Lab software. Each point represents a biological sample for an independent experiment. The data were combined from the three independent experiments (represented as mean ± SD). The p values were obtained from unpaired two-tailed t -tests. The significance is indicated as * ( p < 0.05), *** ( p < 0.001), **** ( p < 0.0001), or n.s. (not significant). Source data and exact p- values are provided in the Source Data file.

Article Snippet: Primary antibody was added and incubated for 16–18 h at 4 °C using the 1:1000 dilution for Rabbit anti-human C4 antibody (mAb #60059) (Cell Signaling).

Techniques: Activation Assay, Western Blot, Positive Control, Molecular Weight, SDS Page, Software, Two Tailed Test