e coli atcc 25922gfp  (ATCC)

Journal: The Journal of Immunology Author Choice

Article Title: Harnessing endogenous anti-glycan antibodies using a novel, bifunctional immunotherapy to treat gram-negative bacterial infections

doi: 10.1093/jimmun/vkaf055

Figure Lengend Snippet: Recruitment of anti-αGal antibodies to multiple Gram-negative bacterial strains by CTX-09 compared to control compound, Polymyxin B. (A) Chemical structure of CTX-09. (B) Schematic outlining the recruitment of anti-αGal antibodies to a Gram-negative bacterium via the effector moiety of CTX-09 compared to Polymyxin B. Created in Biorender.com. (C) Anti-αGal IgM recruitment mediated by 20 µM CTX-09 or Polymyxin B to non-MDR and MDR Gram-negative bacterial strains. The fold change in recruitment over vehicle control is presented. (D) Anti-αGal IgG recruitment was measured to E. coli ATCC 700973 in the presence of increasing concentrations of CTX-09 and Polymyxin B. Distribution of data displayed as bars representing geometric mean ± SD with individual points shown, or line with points representing geometric mean ± SD. * P < 0.05; ** P < 0.01; *** P < 0.001: **** P < 0.0001 using unpaired t -test ( n = 3 independent experiments) or 2-way ANOVA with Šídák's multiple comparisons ( n = 3).

Article Snippet: E. coli ATCC 700973, E. coli ATCC 25922–GFP, and P. aeruginosa ATCC 27853 were purchased from LGC standards; the remaining strains for in vitro and in vivo testing were purchased from UK Health Security Agency culture collections (see for more details on the above strains).

Techniques: Control

Journal: The Journal of Immunology Author Choice

Article Title: Harnessing endogenous anti-glycan antibodies using a novel, bifunctional immunotherapy to treat gram-negative bacterial infections

doi: 10.1093/jimmun/vkaf055

Figure Lengend Snippet: Assessment of complement deposition and complement mediated killing induced by CTX-09. (A) C3b deposition was measured by incubating E. coli ATCC 25922 expressing green fluorescent protein (GFP) with 50% human serum (HS) and the indicated concentrations of CTX-09. The fold change relative to vehicle control was calculated for each CTX-09 concentration. (B) Fluorescence microscopy images comparing C3b deposition using PE-labeled anti-C3b antibody, for bacteria + 50% HS or 50% HS + 2.5 µM CTX-09. GFP= E. coli ATCC 25922; PE= C3b. Scale bar (white) = 100 µm. (C) Antibody-mediated complement deposition was assessed by measuring C1q and C3b deposition on E. coli ATCC 700973 after incubation with 20% HS + CTX-09 or Polymyxin B (PMB). Baseline = Bacteria incubated in 20% HS only. (D) Flow cytometry dot plots showing a CTX-09-mediated increase in C1q+C3b+ E. coli ATCC 700973 cells (upper right quadrant) vs 20% HS alone and 20% HS supplemented with 20 µM CTX-09. (E) Complement dependent cytotoxicity (CDC) was determined by measuring E. coli ATCC 700973 CFU concentrations following incubation with 20% HS and indicated concentrations of CTX-09. LOD, limit of detection. (F) Antibody dependent CDC was confirmed by comparing reductions in bacterial counts following incubation with either 20% HS, 20 µM CTX-09, or 20% HS + 20 µM CTX-09. LOD, limit of detection. Distribution of data displayed as bars representing mean or geometric mean ± SD with individual points plotted, or as line with points representing geometric mean ± SD. ns, not significant; * P < 0.05; ** P < 0.01; *** P < 0.001; **** P < 0.0001 using 2-way ANOVA with Šídák's multiple comparisons ( n = 4) or 1-way ANOVA with Holm-Šídák's multiple comparisons ( n = 3–6).

Article Snippet: E. coli ATCC 700973, E. coli ATCC 25922–GFP, and P. aeruginosa ATCC 27853 were purchased from LGC standards; the remaining strains for in vitro and in vivo testing were purchased from UK Health Security Agency culture collections (see for more details on the above strains).

Techniques: Expressing, Control, Concentration Assay, Fluorescence, Microscopy, Labeling, Bacteria, Incubation, Flow Cytometry

Journal: The Journal of Immunology Author Choice

Article Title: Harnessing endogenous anti-glycan antibodies using a novel, bifunctional immunotherapy to treat gram-negative bacterial infections

doi: 10.1093/jimmun/vkaf055

Figure Lengend Snippet: Demonstrating CTX-09 dependent increases in opsonophagocytic killing of E. coli . (A) Schematic outlining the process of antibody dependent cellular phagocytosis facilitated by CTX-09. Created in Biorender.com. (B) Comparison of E. coli ATCC 25922 opsonophagocytic killing in no human serum (HS) and 5% HS at increasing concentrations of CTX-09 with HL-60 cells differentiated to neutrophil-like cells with phagocytic capacity (dHL-60). LOD, limit of detection. (C) To establish the effect of dHL-60 presence specifically, the reduction in E. coli ATCC 25922 CFU was determined in the presence of HS at indicated concentrations of CTX-09 ± dHL-60 cells. (D) Confirmation of phagocytic cell enhanced clearance of bacterial burden was determined by comparison of combinations of 0.4 µM CTX-09, 5% HS, and dHL-60 cells. (E) Comparison of ADCP in the presence of CTX-09 and Polymyxin B to determine anti-αGal-mediated activity. (F) Representative fluorescence microscopy images comparing phagocytosis of E. coli by dHL-60 cells in the presence or absence of HS and/or CTX-09. Hoechst 33342= Nucleus; PE= CD11b; GFP= E. coli ATCC 25922. Scale bar (white)= 150 µm. Distribution of data displayed as bars representing geometric mean ± SD with individual points shown. ns, not significant; * P < 0.05; ** P < 0.01; *** P < 0.001; **** P < 0.0001 using Kruskal-Wallis with Dunn's multiple comparisons ( n = 3), One-way ANOVA with Holm-Šídák multiple comparisons ( n = 3), Mann-Whitney U test ( n = 3), and unpaired t -test ( n = 3). Holm-Šídák method multiple unpaired t -test with Welch's correction was used for (C) and (E), with a significance threshold of α < 0.05.

Article Snippet: E. coli ATCC 700973, E. coli ATCC 25922–GFP, and P. aeruginosa ATCC 27853 were purchased from LGC standards; the remaining strains for in vitro and in vivo testing were purchased from UK Health Security Agency culture collections (see for more details on the above strains).

Techniques: Comparison, Activity Assay, Fluorescence, Microscopy, MANN-WHITNEY

Journal: The Journal of Immunology Author Choice

Article Title: Harnessing endogenous anti-glycan antibodies using a novel, bifunctional immunotherapy to treat gram-negative bacterial infections

doi: 10.1093/jimmun/vkaf055

Figure Lengend Snippet: Assessment of CTX-09 efficacy in a neutropenic murine thigh infection model. (A) Pharmacokinetics of CTX-09 and human anti-αGal IgG in mouse plasma over 12 h. LLOQ, lower limit of quantification. (B) Study design outlining timings of cyclophosphamide administration, passive immunization with anti-αGal, infection with E. coli ATCC 700973, and CTX-09 or mock dosing in female GGTA1 −/− mice. s.c., sub-cutaneous; i.m., intra-muscular. (C) Comparison of bacterial burden in the thigh tissue between vehicle ± human anti-αGal IgG and CTX-09 ± human anti-αGal IgG. Distribution of data from a single experiment are displayed as individual values with each point representative of an individual thigh. Outliers were excluded following ROUT outlier test (Q = 1%). ns, not significant; * P < 0.05; ** P < 0.01; *** P < 0.001; **** P < 0.0001 using Kruskal-Wallis test with Dunn's multiple comparisons ( n = 4–8).

Article Snippet: E. coli ATCC 700973, E. coli ATCC 25922–GFP, and P. aeruginosa ATCC 27853 were purchased from LGC standards; the remaining strains for in vitro and in vivo testing were purchased from UK Health Security Agency culture collections (see for more details on the above strains).

Techniques: Infection, Drug discovery, Clinical Proteomics, Comparison

Journal: Antibiotics

Article Title: Using GFP-Tagged Escherichia coli to Investigate the Persistence of Fecal Bacteria in Vegetated Wetlands: An Experimental Approach

doi: 10.3390/antibiotics9060335

Figure Lengend Snippet: Experimental design. Planted and unplanted rhizotrons inoculated with GFP- Escherichia coli in water or sediment.

Article Snippet: We used a fluorescent and nonpathogenic strain E. coli GFP ATCC ® 25922GFPTM (American Type Culture Collection, Manassas, VA, USA).

Techniques: Control

Journal: Antibiotics

Article Title: Using GFP-Tagged Escherichia coli to Investigate the Persistence of Fecal Bacteria in Vegetated Wetlands: An Experimental Approach

doi: 10.3390/antibiotics9060335

Figure Lengend Snippet: Effect of the type of inoculation—in water ( n = 6) or in sediment ( n = 6)—on the fluorescence of GFP- Escherichia coli in the rhizotrons. Boxplots of fluorescences measured on days 1, 2, 3, 6 and 8 (fluorescences > limit od detection (LOD)) in the water of rhizotrons inoculated with GFP- E. coli . Using an ANOVA with a multiple comparisons Tukey’s test on the l-mm, testing the effects of the type of inoculation on the fluorescence (i.e., bacterial concentration) of the water column. a and b are significantly different ( p -value (ANOVA) < 0.01). CTCF: corrected total cell fluorescence.

Article Snippet: We used a fluorescent and nonpathogenic strain E. coli GFP ATCC ® 25922GFPTM (American Type Culture Collection, Manassas, VA, USA).

Techniques: Fluorescence, Concentration Assay

Journal: Antibiotics

Article Title: Using GFP-Tagged Escherichia coli to Investigate the Persistence of Fecal Bacteria in Vegetated Wetlands: An Experimental Approach

doi: 10.3390/antibiotics9060335

Figure Lengend Snippet: Effects of Baldellia ranunculoides ( n = 3), Elodea canadensis ( n = 3), Mentha aquatica ( n = 3), Sparganium emersum ( n = 3), the mixture of these four species together ( n = 3) and the absence of plants ( n = 3) on the fluorescence of GFP- E. coli in the rhizotrons. Boxplots of fluorescences measured on days 1, 2, 3, 6 and 8 (fluorescences > LOD) in the water of rhizotrons inoculated with GFP- E. coli . Using an ANOVA with a multiple comparisons Tukey’s test on the l-mm, testing the effects of plant species on the fluorescence (i.e., bacterial concentration) of the water column. a and b are significantly different ( p -value (ANOVA) < 0.001). The dots are outliers.

Article Snippet: We used a fluorescent and nonpathogenic strain E. coli GFP ATCC ® 25922GFPTM (American Type Culture Collection, Manassas, VA, USA).

Techniques: Fluorescence, Concentration Assay

Journal: Antibiotics

Article Title: Using GFP-Tagged Escherichia coli to Investigate the Persistence of Fecal Bacteria in Vegetated Wetlands: An Experimental Approach

doi: 10.3390/antibiotics9060335

Figure Lengend Snippet: Example of planted rhizotrons after the inoculation of fluorescent E. coli (3rd day after inoculation). ( a ) Rhizotron under natural light. ( b ) Rhizotron under fluorescence conditions (enlightened only with a 480–530-nm light and observed with an amber filter). Photo obtained with an exposition of 13 s f/18 and ISO 800 with a Canon EOS 7D.

Article Snippet: We used a fluorescent and nonpathogenic strain E. coli GFP ATCC ® 25922GFPTM (American Type Culture Collection, Manassas, VA, USA).

Techniques: Fluorescence

Journal: Antibiotics

Article Title: Using GFP-Tagged Escherichia coli to Investigate the Persistence of Fecal Bacteria in Vegetated Wetlands: An Experimental Approach

doi: 10.3390/antibiotics9060335

Figure Lengend Snippet: Fitting curve between the fluorescence measurements (CTCF) and the bacterial concentration (McF). Twelve vials with known concentrations of GFP- E. coli (0.2, 0.4, 0.5, 0.7, 0.8, 0.9, 1, 1.5, 2, 3, 4 and 6 McF) were photographed three times under the same conditions as the rhizotrons. The values presented are the means of these three measurements for each vial.

Article Snippet: We used a fluorescent and nonpathogenic strain E. coli GFP ATCC ® 25922GFPTM (American Type Culture Collection, Manassas, VA, USA).

Techniques: Fluorescence, Concentration Assay