Journal: ERJ Open Research

Article Title: Donor-derived airway air–liquid interface model for high-throughput screening of antiviral combinations with concurrent analysis of antiviral efficacy and epithelial toxicity using ciliR

doi: 10.1183/23120541.01283-2025

Figure Lengend Snippet: Viral inhibition and ciliary beat frequency (CBF) analysis with high-dose monotherapies of ALS-8112 and CPD23. a) Each well of 96-well high-throughput screening (HTS) air–liquid interface (ALI) culture was scanned for respiratory syncytial virus (RSV)-green fluorescent protein (GFP) fluorescence at days 3 and 7 post infection. b, c) Box plots and histograms showing the mean CBF and the distribution of CBF for ciliated cells with and those without RSV infection (paired t-test, ns : nonsignificant) per well (n=9). d) Dose–response curves showing inhibition of RSV-GFP by ALS-8112 and CPD23 (three plates). e) Representative fluorescence images of whole wells showing GFP expression, indicating RSV infection, at various concentrations of ALS-8112 (500–4000 nM) and CPD23 (5–80 nM). Wells treated with increasing concentrations of ALS-8112 and CPD23 exhibit reduced GFP expression, indicating inhibition of RSV-GFP. Scale bar represents 1 mm. f) Box plots showing the mean CBF and the distribution of CBF for ciliated cells treated with varying concentrations of ALS-8112 (500–4000 nM)) or CPD23 (5–80 nM) for 3 days (left panels) or 7 days (right panels) postinfection. No significant differences were detected (Wilcoxon test). ns : nonsignificant.

Article Snippet: Following infection, cells were fed basolaterally with media containing different concentrations of drugs: an inhibitor of RSV Fusion (F) protein (CPD23, compound 23 from C ockerill et al . [ ]) that blocks viral entry to the cell, and a polymerase inhibitor (ALS-8112, MedChemExpress) that causes lethal virus mutagenesis or disturbance of viral RNA synthesis to inhibit viral replication [ ].

Techniques: Inhibition, High Throughput Screening Assay, Virus, Fluorescence, Infection, Expressing

Journal: ERJ Open Research

Article Title: Donor-derived airway air–liquid interface model for high-throughput screening of antiviral combinations with concurrent analysis of antiviral efficacy and epithelial toxicity using ciliR

doi: 10.1183/23120541.01283-2025

Figure Lengend Snippet: Dose–response and toxicity analysis of combination therapies in primary human airway epithelial cells. a) Synergy analysis at day 3 and d) day 7. The 3D synergy maps illustrate the interaction between ALS-8112 and CPD23. The synergy scores were calculated using the Loewe model, where red regions indicate synergistic interactions, and green regions indicate antagonistic interactions. Overall values between −10 and 10 are considered additive. b, e) Toxicity matrices of the ciliary beat frequency (CBF) across the 96-well high-throughput screening (HTS) treatment plate at b) day 3 and e) day 7, shown as a 3D surface plot. The mean CBF remained between 8 and 9 Hz, indicating minimal impact on ciliary activity. c, f) The active area percentage, indicating overall cellular activity, remained consistent at c) day 3 and f) day 7 with untreated plates across different concentrations of ALS-8112 and CPD23, demonstrating no significant toxicity. g) Representative fluorescence images of whole wells showing green fluorescent protein (GFP) expression, indicating respiratory syncytial virus (RSV) infection, with ALS-8112 (4000 nM) and CPD23 (40 nM) and in combination. Wells treated with ALS-8112 and CPD23 exhibit reduced GFP expression, indicating inhibition of RSV. Scale bar represents 1 mm.

Article Snippet: Following infection, cells were fed basolaterally with media containing different concentrations of drugs: an inhibitor of RSV Fusion (F) protein (CPD23, compound 23 from C ockerill et al . [ ]) that blocks viral entry to the cell, and a polymerase inhibitor (ALS-8112, MedChemExpress) that causes lethal virus mutagenesis or disturbance of viral RNA synthesis to inhibit viral replication [ ].

Techniques: High Throughput Screening Assay, Activity Assay, Fluorescence, Expressing, Virus, Infection, Inhibition

Journal: medRxiv

Article Title: Mucosal IgA to pre-fusion F protein predicts protection from RSV infection in a high burden setting

doi: 10.64898/2026.03.16.26348479

Figure Lengend Snippet: (A) Longitudinal trajectories of serum IgG (top row) and mucosal IgA (bottom row) antibody responses against four RSV-B proteins (PreF, PostF, G, NP) over time. Individual participant trajectories are shown as thin lines with low opacity, colored by infection status: not infected (black), sero-detected infections (orange), and PCR-confirmed infections (green). (B) Mean fold-change in antibody titres between the first bleed (pre-epidemic baseline) and second bleed (post-epidemic) for serum IgG and mucosal IgA responses to RSV-B proteins. Bars represent mean fold-change (log10 scale) stratified by infection status, with error bars indicating standard error.

Article Snippet: RSV antigens (RSV-A Pre-Fusion (ProteoGenix, product code: PX-P6126), RSV-B Pre-Fusion (SinoBiological, product code: 40832-V08B), RSV-A Post-Fusion (SinoBiological, product code: 11049-V08B), RSV-B Post-Fusion (SinoBiological, product code: 40999-V08H), RSV-A G protein (SinoBiological, product code: 11070-V08H2), RSV-B G protein (SinoBiological, product code: 13029-V08H, RSV-A Nucleoprotein (SinoBiological, product code: 40821-V08E and RSV-B Nucleoprotein (SinoBiological, product code: 40822-V08F)), were coupled to distinct bead regions.

Techniques: Infection

Journal: medRxiv

Article Title: Mucosal IgA to pre-fusion F protein predicts protection from RSV infection in a high burden setting

doi: 10.64898/2026.03.16.26348479

Figure Lengend Snippet: (A) Post-infection longitudinal antibody titers for four viral RSV-B proteins (PreF, PostF, G, and NP) across different antibody types; serum IgG and mucosal IgA. Lines show the median posterior predictive fit from the fitted Bayesian model, and the points show the observational titre data, with the size correlating with the sample size for that bin. (B) Peak antibody (x axis) and persistence measured as duration above a 2-fold (left panel) and 4-fold titre rise (right panel) in days. Data points show median posterior values for measurements of four viral RSV-B proteins (PreF, PostF, G, and NP) across different antibody types: serum IgG and mucosal IgA.

Article Snippet: RSV antigens (RSV-A Pre-Fusion (ProteoGenix, product code: PX-P6126), RSV-B Pre-Fusion (SinoBiological, product code: 40832-V08B), RSV-A Post-Fusion (SinoBiological, product code: 11049-V08B), RSV-B Post-Fusion (SinoBiological, product code: 40999-V08H), RSV-A G protein (SinoBiological, product code: 11070-V08H2), RSV-B G protein (SinoBiological, product code: 13029-V08H, RSV-A Nucleoprotein (SinoBiological, product code: 40821-V08E and RSV-B Nucleoprotein (SinoBiological, product code: 40822-V08F)), were coupled to distinct bead regions.

Techniques: Infection

Journal: medRxiv

Article Title: Mucosal IgA to pre-fusion F protein predicts protection from RSV infection in a high burden setting

doi: 10.64898/2026.03.16.26348479

Figure Lengend Snippet: Serum IgG, top row; mucosal IgA, bottom row and columns are viral antigen target (PreF, PostF, G, and NP for both RSV-A and RSV-B strains). The solid green line represents the mean estimated probability of protection given exposure to infection as a function of antibody titre, with shaded ribbons indicating 95% credible intervals. Background histograms show the distribution of antibody titres at infection for infected individuals (orange) versus non-infected individuals (gray).

Article Snippet: RSV antigens (RSV-A Pre-Fusion (ProteoGenix, product code: PX-P6126), RSV-B Pre-Fusion (SinoBiological, product code: 40832-V08B), RSV-A Post-Fusion (SinoBiological, product code: 11049-V08B), RSV-B Post-Fusion (SinoBiological, product code: 40999-V08H), RSV-A G protein (SinoBiological, product code: 11070-V08H2), RSV-B G protein (SinoBiological, product code: 13029-V08H, RSV-A Nucleoprotein (SinoBiological, product code: 40821-V08E and RSV-B Nucleoprotein (SinoBiological, product code: 40822-V08F)), were coupled to distinct bead regions.

Techniques: Infection

Journal: medRxiv

Article Title: Mucosal IgA to pre-fusion F protein predicts protection from RSV infection in a high burden setting

doi: 10.64898/2026.03.16.26348479

Figure Lengend Snippet: Model performance comparison across single biomarker models and the dual biomarker model, defined by out-of-sample predictive accuracy (LOO-ELPD, x-axis) and discrimination ability (area under the ROC curve, AUC, y-axis). Circles indicate serum IgG models, squares represent mucosal IgA models, and the triangle denotes the dual biomarker model combining serum IgG and mucosal IgA to RSV-B PreF. The best-performing model within each biomarker class is highlighted with darker shading. Error bars show the standard error of LOO-ELPD (horizontal) and 95% confidence intervals for AUC (vertical). The dashed horizontal line indicates an AUC of 0.7.

Article Snippet: RSV antigens (RSV-A Pre-Fusion (ProteoGenix, product code: PX-P6126), RSV-B Pre-Fusion (SinoBiological, product code: 40832-V08B), RSV-A Post-Fusion (SinoBiological, product code: 11049-V08B), RSV-B Post-Fusion (SinoBiological, product code: 40999-V08H), RSV-A G protein (SinoBiological, product code: 11070-V08H2), RSV-B G protein (SinoBiological, product code: 13029-V08H, RSV-A Nucleoprotein (SinoBiological, product code: 40821-V08E and RSV-B Nucleoprotein (SinoBiological, product code: 40822-V08F)), were coupled to distinct bead regions.

Techniques: Comparison, Biomarker Discovery

Journal: PLoS ONE

Article Title: Discovery and Characterization of Phage Display-Derived Human Monoclonal Antibodies against RSV F Glycoprotein

doi: 10.1371/journal.pone.0156798

Figure Lengend Snippet: Heavy chain C-terminal 6xHis tagged antigen specific bivalent Fabs were purified with Ni-NTA column and then tested in ELISA binding to RSV prefusion and postfusion F proteins. (A) Antibodies preferentially binding to RSV postfusion F protein; (B) Antibodies binding to both RSV postfusion and prefusion F proteins; (C) antibody binding specifically to RSV prefusion F protein. Full-length human IgG1 D25 (prefusion F specific) and palivizumab (binding to both prefusion and postfusion F) were used as control antibodies in the above experiments.

Article Snippet: Three groups of RSV F protein specific antibodies were identified from the Morphosys HuCAL GOLD ® phage display library ( ): group 1 mAbs (3B1, 3B5, 3E2, 3H3, 3G4) which bind to the postfusion RSV F protein strongly with minimal binding to the prefusion F protein ( ); group 2 mAbs (3B2, 3D3, 3B7 and 3C2) which bind to both prefusion and postfusion F proteins ( ); and group 3 mAb (2E1) which binds specifically to prefusion F protein, although weakly as a bivalent Fab ( ).

Techniques: Purification, Enzyme-linked Immunosorbent Assay, Binding Assay

Journal: PLoS ONE

Article Title: Discovery and Characterization of Phage Display-Derived Human Monoclonal Antibodies against RSV F Glycoprotein

doi: 10.1371/journal.pone.0156798

Figure Lengend Snippet: (A-B) ELISA analysis of 2E1 IgG (A) and 3B1 IgG (B) binding to RSV pre- (red circle) and postfusion F (blue square) proteins. (C-H) Surface plasmon resonance (SPR) analysis of 2E1 and 3B1 Fabs binding to pre- and postfusion RSV F proteins. RU = Resonance Units. Monovalent Fab antibody fragments were captured on the surface of a Series S Sensor Chip CM5 previously functionalized with Human Fab Binder. Prefusion or postfusion F protein diluted 2-fold serially starting at 100 nM or 200 nM was then injected over captured 2E1 (C) or 3B1 (F), respectively. To determine steady-state affinity, response levels at equilibrium were plotted over concentration of pre- (D) or postfusion F (G) protein. 50 nM of post- (E) or prefusion F (H) was injected to demonstrate the specificity of 2E1 binding to prefusion F and 3B1 to postfusion F. (I-K) SPR based competition analysis of 2E1 and 3B1 against palivizumab (I), D25 (J) and MPE8 (K) in binding to RSV prefusion F protein. RU = Resonance Units. Palivizumab and D25 were amine coupled to the surface of separate flow channels of a CM5 chip. A third flow channel was subjected to amine coupling activation without an antibody and used for reference subtraction. Prefusion F (40 μg/mL) was then injected over all surfaces. After a brief stabilization period, 2E1, 3B1, or running buffer was injected to measure binding to sites not occupied by the capturing antibody (palivizumab or D25). To assess competition to MPE8, the MPE8 antibody was captured (6000 RU, not shown) to flow channel 2 of a Biacore Sensor Chip Protein A. Prefusion F (40 μg/mL) was passed over channels 1 and 2 followed by 2E1 Fab, 3B1 Fab, D25 Fab and buffer to measure binding to sites not occupied by MPE8. (L) Bio-Layer Interferometry (BLI) based competition experiment of 3B1 IgG against site I antibody 131-2a. Palivizumab (blue) is able to bind to postfusion F protein in an Octet sandwich competition assay using 131-2a as the capture antibody, but antibody 3B1 (red) does not bind.

Article Snippet: Three groups of RSV F protein specific antibodies were identified from the Morphosys HuCAL GOLD ® phage display library ( ): group 1 mAbs (3B1, 3B5, 3E2, 3H3, 3G4) which bind to the postfusion RSV F protein strongly with minimal binding to the prefusion F protein ( ); group 2 mAbs (3B2, 3D3, 3B7 and 3C2) which bind to both prefusion and postfusion F proteins ( ); and group 3 mAb (2E1) which binds specifically to prefusion F protein, although weakly as a bivalent Fab ( ).

Techniques: Enzyme-linked Immunosorbent Assay, Binding Assay, SPR Assay, Injection, Concentration Assay, Activation Assay, Competitive Binding Assay

Journal: PLoS ONE

Article Title: Discovery and Characterization of Phage Display-Derived Human Monoclonal Antibodies against RSV F Glycoprotein

doi: 10.1371/journal.pone.0156798

Figure Lengend Snippet: (A) Heat map plot showing the difference in deuteration levels of the RSV prefusion F protein alone compared to RSV prefusion F protein in the presence of the 2E1 monovalent Fab at five time points (15, 50, 150, 500, and 1500 sec). Slower deuterium exchange indicates regions containing the binding sites. White areas are ‘gaps’ for which there was no sequence coverage, and thus no HDX-MS information was obtained. Dashed greyed-out areas represent sequences of the signal and P27 peptides which are not present in the mature purified protein. (B) Uptake plots of several RSV F peptides spanning the conformational epitope region. Red curves show the deuteration levels of peptides of RSV F protein alone, while blue curves show the peptides of the RSV F / 2E1 complex. Peptides containing the residues of antibody epitope (417–434, 441–448, and 457–467) showed decreased deuteration level upon 2E1 binding. In contrast, peptides with no significant decrease in the deuteration level upon 2E1 exposure represent non-epitope sequences (435–440 and 449–457). The residues identified as critical for binding by shotgun mutagenesis are indicated in red font.

Article Snippet: Three groups of RSV F protein specific antibodies were identified from the Morphosys HuCAL GOLD ® phage display library ( ): group 1 mAbs (3B1, 3B5, 3E2, 3H3, 3G4) which bind to the postfusion RSV F protein strongly with minimal binding to the prefusion F protein ( ); group 2 mAbs (3B2, 3D3, 3B7 and 3C2) which bind to both prefusion and postfusion F proteins ( ); and group 3 mAb (2E1) which binds specifically to prefusion F protein, although weakly as a bivalent Fab ( ).

Techniques: Binding Assay, Sequencing, Purification, Mutagenesis