america type culture collection tib 202 crl 2019  (ATCC)

Journal: bioRxiv

Article Title: A Prefusion Form of Herpes Simplex Virus 1 gB has a Distinct Antigenic Signature

doi: 10.64898/2026.04.12.718011

Figure Lengend Snippet: (A) CHO-K1 effector cells transiently expressing wild type gB or gB H516P and wild type gD, gH, and gL, and T7 polymerase were co-cultured for 18 h with CHO-HVEM cells transiently expressing the luciferase gene. Luciferase-induced luminescence was quantitated as a measure of fusion. (B) Detection of gB on the cell surface by CELISA. CHO-K1 cells were transfected with plasmids encoding HSV-1 gB or empty vector for 24 h. Cells were fixed with paraformaldehyde and then incubated with anti-gB monoclonal antibodies H126, H1359, and H1817. HRP-conjugated Protein A was added, followed by ABTS substrate. Absorbance was read at 405 nm. Data was normalized to wild type gB set to 100% for both experiments. Results are the means of three independent experiments. *, p < 0.05; ns, not significant, Student’s t- test.

Article Snippet: Chinese hamster ovary (CHO-K1) cells (American Type Culture Collection (ATCC), Manassas, VA, USA) were propagated in Ham’s F12 nutrient mixture (Gibco/Life Technologies, Grand Island, NY, USA) supplemented with 10% fetal bovine serum (FBS) (Atlanta Biologicals, Atlanta, GA, USA).

Techniques: Expressing, Cell Culture, Luciferase, Transfection, Plasmid Preparation, Incubation, Bioprocessing

Journal: bioRxiv

Article Title: A Prefusion Form of Herpes Simplex Virus 1 gB has a Distinct Antigenic Signature

doi: 10.64898/2026.04.12.718011

Figure Lengend Snippet: (A) Space-filling rendering (PyMOL) of prefusion gB (PDB 6Z9M) and postfusion gB (PDB 2GUM) ectodomain trimer . Epitopes of monoclonal antibodies used in this study are highlighted in color. The MAb H126 epitope includes residue 303 in gB domain I (blue) ( , ). The MAb H1838 epitope maps to residues 391 – 410 in domain II (green) . The H1359 epitope maps to residue 487-505 in domain III (yellow) . The SS10 epitope maps to residues 640 – 670 in domain IV (orange) . The SS106 epitope maps to residue 697 – 725 in domain V (red). The MAb DL16 epitope is trimer-specific and resides in domain V . The MAb H1817 epitope maps to residues 31 – 43 in gB domain VI (white) , which is unresolved in the structure. (B-H) gB H516P is antigenically distinct from wild type gB at domains II, IV, and V. Lysates of CHO-K1 cells expressing wild type gB (red) or H516P gB (purple) were bound to nitrocellulose membrane then probed with antibodies to gB. The appropriate fluorescent-conjugated secondary antibody was added for 20 min. Images were generated with Azure Biosystems imager. Rabbit polyclonal antibody R68 was used to standard loading. Reactivity of the antibodies was quantitated relative to the wild type by densitometry using Image Studio. The values shown represent antibody reactivity relative to wild type gB set to 100%. Results are the means and standard deviations of three independent experiments. ** p < 0.005; *** p < 0.001, Student’s t- test. (I) Summary of the antigenic analysis of gB H516P. The slopes of the lines of best fit for H516P gB relative to wild type (panels B-H) were calculated and used to determine fold changes, which were normalized to 1.0.

Article Snippet: Chinese hamster ovary (CHO-K1) cells (American Type Culture Collection (ATCC), Manassas, VA, USA) were propagated in Ham’s F12 nutrient mixture (Gibco/Life Technologies, Grand Island, NY, USA) supplemented with 10% fetal bovine serum (FBS) (Atlanta Biologicals, Atlanta, GA, USA).

Techniques: Bioprocessing, Residue, Expressing, Membrane, Generated

Journal: bioRxiv

Article Title: A Prefusion Form of Herpes Simplex Virus 1 gB has a Distinct Antigenic Signature

doi: 10.64898/2026.04.12.718011

Figure Lengend Snippet: Lysate from CHO-K1 cells expressing wild type gB or H516P gB was treated at a range of pH values (7.4 to 5.0) for 10 min at 37°C. Samples were blotted to nitrocellulose membranes and probed at neutral pH with gB MAbs targeting epitopes undergoing conformational change in (A) Domain 1, H126; (B) Domain V, SS106; and epitope not undergoing conformational change in (C) Domain IV, SS10 at neutral pH. Fluorescent-conjugated secondary antibody was added for 20 min. Images were generated with Azure Biosystems imager. Reactivity of three independent experiments was quantified by densitometry. The pH 7.4-treated samples were set to 100%.

Article Snippet: Chinese hamster ovary (CHO-K1) cells (American Type Culture Collection (ATCC), Manassas, VA, USA) were propagated in Ham’s F12 nutrient mixture (Gibco/Life Technologies, Grand Island, NY, USA) supplemented with 10% fetal bovine serum (FBS) (Atlanta Biologicals, Atlanta, GA, USA).

Techniques: Expressing, Generated

Journal: bioRxiv

Article Title: A Prefusion Form of Herpes Simplex Virus 1 gB has a Distinct Antigenic Signature

doi: 10.64898/2026.04.12.718011

Figure Lengend Snippet: (A) CHO-K1 cells were transfected with plasmids encoding wild type gB or H516P gB. Cell lysates were either prepared in 2% SDS sample buffer with reducing agent and heated (denatured conditions), and then resolved by SDS-PAGE (left panel) or prepared with 0.2% SDS, no reducing agent, no heating (“native” conditions) and resolved by SDS-PAGE (right panel). Western blots were probed with anti-gB polyclonal antibody R68. Molecular weight standards (kDa) are indicated to the right. β-actin was included as a loading control. (B) Lysates of CHO-K1 cells expressing wild type gB (red) or H516P gB (purple) were bound to nitrocellulose membrane then probed with MAb DL16. Anti-mouse fluorescent-conjugated secondary antibody was added for 20 min. Images were generated with Azure Biosystems imager. Reactivity of DL16 was quantitated relative to the wild type by densitometry using Image Studio. gB wild type, 1.00; gB H516P, 0.31+/-0.01****. The values shown represent antibody reactivity relative to wild type gB set to 100%. Results are the means and standard deviations of three independent experiments. *** p < 0.0001, Student’s t- test.

Article Snippet: Chinese hamster ovary (CHO-K1) cells (American Type Culture Collection (ATCC), Manassas, VA, USA) were propagated in Ham’s F12 nutrient mixture (Gibco/Life Technologies, Grand Island, NY, USA) supplemented with 10% fetal bovine serum (FBS) (Atlanta Biologicals, Atlanta, GA, USA).

Techniques: Transfection, SDS Page, Western Blot, Molecular Weight, Control, Expressing, Membrane, Generated

Journal: Frontiers in Medicine

Article Title: Protective effects of the RAGE inhibitor azeliragon as a potential anti- Streptococcus pneumoniae therapeutic in sepsis models

doi: 10.3389/fmed.2026.1793580

Figure Lengend Snippet: Minimum inhibitory concentrations (MICs) of azeliragon, penicillin, tetracycline, and vancomycin against clinical S. pneumoniae isolates were determined using the broth microdilution method. MIC 50 [the lowest concentration inhibiting 50% of isolates; (A) ] and MIC 90 [the lowest concentration inhibiting 90% of isolates; (B) ] were calculated. Values shown above the bars indicate the corresponding MIC 50 or MIC 90 (μg/mL). Data were presented as mean ± SD. The experiments were performed with three independent biological replicates ( n = 3).

Article Snippet: Streptococcus pneumoniae American Type Culture Collection (ATCC) 49,619 and the clinical isolates 1057 and 1044 were used for in vitro antimicrobial assays, mechanistic studies, and in vivo infection models.

Techniques: Concentration Assay

Journal: Frontiers in Medicine

Article Title: Protective effects of the RAGE inhibitor azeliragon as a potential anti- Streptococcus pneumoniae therapeutic in sepsis models

doi: 10.3389/fmed.2026.1793580

Figure Lengend Snippet: Effects of azeliragon on the in vitro growth of S. pneumoniae . Dose–response relationships of S. pneumoniae ATCC 49619 (A) , S. pneumoniae 1057 (B) , and S. pneumoniae 1044 (C) following treatment with increasing concentrations of azeliragon. Bacterial growth was measured after 18 h of incubation and expressed as OD 600 . Effects of azeliragon on the growth kinetics of S. pneumoniae ATCC 49619 (D) , S. pneumoniae 1057 (E) , and S. pneumoniae 1044 (F) . Bacteria were cultured under control conditions or with different concentrations of azeliragon, and OD 600 was recorded at indicated time points to generate growth curves. Data were presented as mean ± SD. The experiments were performed with three independent biological replicates ( n = 3). “ns” represented no significant difference, ** p < 0.01.

Article Snippet: Streptococcus pneumoniae American Type Culture Collection (ATCC) 49,619 and the clinical isolates 1057 and 1044 were used for in vitro antimicrobial assays, mechanistic studies, and in vivo infection models.

Techniques: In Vitro, Incubation, Bacteria, Cell Culture, Control

Journal: Frontiers in Medicine

Article Title: Protective effects of the RAGE inhibitor azeliragon as a potential anti- Streptococcus pneumoniae therapeutic in sepsis models

doi: 10.3389/fmed.2026.1793580

Figure Lengend Snippet: Time-kill activity and biofilm disruption effects of azeliragon against S. pneumoniae . Time-kill curves of S. pneumoniae ATCC 49619 (A) , S. pneumoniae 1057 (B) , and S. pneumoniae 1044 (C) treated with azeliragon (4× MIC). Bacteria were cultured under control conditions or treated with azeliragon (4× MIC) or vancomycin (4× MIC), and viable counts (CFU/mL) were determined at indicated time points. Effects of azeliragon (4× MIC) on biofilm biomass of S. pneumoniae ATCC 49619 (D) , S. pneumoniae 1057 (E) , and S. pneumoniae 1044 (F) . Mature biofilms were treated with the indicated drugs for 24 h and quantified by crystal violet staining. Results are expressed as OD₅₇₀. Data were presented as mean ± SD. Data were presented as mean ± SD. The experiments were performed with six independent biological replicates ( n = 6). Group comparisons for biofilm assays were analyzed by one-way ANOVA followed by Tukey’s multiple-comparison test. “ns” represented no significant difference, *** p < 0.001.

Article Snippet: Streptococcus pneumoniae American Type Culture Collection (ATCC) 49,619 and the clinical isolates 1057 and 1044 were used for in vitro antimicrobial assays, mechanistic studies, and in vivo infection models.

Techniques: Activity Assay, Disruption, Bacteria, Cell Culture, Control, Staining, Comparison

Journal: Frontiers in Medicine

Article Title: Protective effects of the RAGE inhibitor azeliragon as a potential anti- Streptococcus pneumoniae therapeutic in sepsis models

doi: 10.3389/fmed.2026.1793580

Figure Lengend Snippet: Effects of azeliragon on membrane integrity and oxidative stress in S. pneumoniae . (A) Propidium iodide (PI) staining was used to assess membrane permeability. Bacteria were treated under control conditions or with azeliragon (4× MIC) or vancomycin (4× MIC). Fluorescence intensity (A.U.) was measured to evaluate membrane integrity. (B) Intracellular oxidative stress was assessed using a reactive oxygen species (ROS) fluorescent probe. Bacteria were treated as indicated, and ROS fluorescence intensity (A.U.) was quantified. Data were presented as mean ± SD. Group comparisons were performed using one-way ANOVA followed by Tukey’s multiple-comparison test. Data were presented as mean ± SD. The experiments were performed with three independent biological replicates ( n = 3). “ns” represented no significant difference, * p < 0.05, ** p < 0.01.

Article Snippet: Streptococcus pneumoniae American Type Culture Collection (ATCC) 49,619 and the clinical isolates 1057 and 1044 were used for in vitro antimicrobial assays, mechanistic studies, and in vivo infection models.

Techniques: Membrane, Staining, Permeability, Bacteria, Control, Fluorescence, Comparison

Journal: Frontiers in Medicine

Article Title: Protective effects of the RAGE inhibitor azeliragon as a potential anti- Streptococcus pneumoniae therapeutic in sepsis models

doi: 10.3389/fmed.2026.1793580

Figure Lengend Snippet: In vivo protective effects of azeliragon in a mouse infection model. (A) Determination of the infectious dose of S. pneumoniae 1044. (B) Therapeutic efficacy of azeliragon in the pneumonia model. (C) Seven-day survival curves of mice with sepsis induced by S. pneumoniae ATCC 49619. Mice received vehicle control, bacterial control, azeliragon (5 mg/kg or 10 mg/kg), or vancomycin (10 mg/kg). Each group contained 10 mice. Survival was monitored continuously. (D) Seven-day survival curves of mice with sepsis induced by the clinical S. pneumoniae isolate 1044. Experimental groups and treatments were identical to those in (A) . Survival data were plotted using the Kaplan–Meier method, and statistical significance was determined using the log-rank (Mantel–Cox) test. “ns” represented no significant difference, * p < 0.05, ** p < 0.01, and *** p < 0.001.

Article Snippet: Streptococcus pneumoniae American Type Culture Collection (ATCC) 49,619 and the clinical isolates 1057 and 1044 were used for in vitro antimicrobial assays, mechanistic studies, and in vivo infection models.

Techniques: In Vivo, Infection, Drug discovery, Control

Journal: Frontiers in Medicine

Article Title: Protective effects of the RAGE inhibitor azeliragon as a potential anti- Streptococcus pneumoniae therapeutic in sepsis models

doi: 10.3389/fmed.2026.1793580

Figure Lengend Snippet: Effects of azeliragon on tissue bacterial burden and inflammatory cytokines in a non-lethal mouse model of S. pneumoniae infection. (A) Bacterial loads in blood (CFU/mL). (B) Bacterial loads in lung tissue (CFU/g). (C) Serum interleukin-6 (IL-6) levels (pg/mL). (D) Serum tumor necrosis factor-α (TNF-α) levels (pg/mL). Mice received vehicle control, bacterial control, azeliragon (10 mg/kg), or vancomycin (10 mg/kg). Blood and lung samples were collected for quantitative bacterial culture. Serum cytokine levels were measured by ELISA. Data are presented as mean ± SD. Group comparisons were performed using one-way ANOVA followed by Tukey’s multiple-comparison test. “ns” represented no significant difference, ** p < 0.01 and *** p < 0.001.

Article Snippet: Streptococcus pneumoniae American Type Culture Collection (ATCC) 49,619 and the clinical isolates 1057 and 1044 were used for in vitro antimicrobial assays, mechanistic studies, and in vivo infection models.

Techniques: Infection, Control, Enzyme-linked Immunosorbent Assay, Comparison