Journal: Virology Journal

Article Title: Cell culture medium influences RSV G protein glycosylation and recognition by monoclonal antibodies

doi: 10.1186/s12985-026-03318-8

Figure Lengend Snippet: Medium-dependent effects on RSV G protein size, relative protein signals in virus stocks, and viral spread. ( A ) Western blot analysis of soluble G (sG) protein derived from HEK293T cells (top), HEK293T GALE/GALK1 knockout cells (middle), and whole-cell lysates of RSV A-0594-EGFP-infected A549 cells (bottom, 24 hpi), all produced or infected in cell culture medium A, B or C. ( B ) Culture medium-dependent viral spread of RSV A-0594-EGFP in A549 cells infected at an MOI of 1 and cultured in media A, B, or C. GFP integrated fluorescence intensity (GCU × µm²/image; mean ± SD, n = 9 from three independent experiments) was quantified over time using an IncuCyte system (Sartorius). Statistical significance was assessed by two-way ANOVA. ( C ) Virus spread following infection with RSV virus stocks A-C in A549 cells infected at an MOI of 1, with medium replaced by Opti-MEM (medium B) after adsorption. GFP integrated fluorescence intensity (GCU × µm²/image; mean ± SD, n = 9 from three independent experiments) was quantified over time using an IncuCyte system. ( D ) Relative G protein signal normalised to N protein signal in lysed RSV A-0594-EGFP virus stocks A-C, determined by Western blot densitometry (mean ± SD, n = 3). Statistical significance was assessed by one-way ANOVA. ( E ) Relative F protein signal normalised to N protein signal in lysed RSV A-0594-EGFP virus stocks A-C, determined by Western blot densitometry (mean ± SD, n = 3). Statistical significance was assessed by one-way ANOVA. * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001, **** P ≤ 0.0001. ns, not significant

Article Snippet: Integrated GFP fluorescence (GCU × μm2/image) was quantified over 5 days using an IncuCyte S3 system (Sartorius).

Techniques: Virus, Western Blot, Analysis, Derivative Assay, Knock-Out, Infection, Produced, Cell Culture, Fluorescence, Adsorption

Journal: Virology Journal

Article Title: Cell culture medium influences RSV G protein glycosylation and recognition by monoclonal antibodies

doi: 10.1186/s12985-026-03318-8

Figure Lengend Snippet: Antibody binding and neutralization of recombinant RSV produced in medium A, B or C. ( A ) Quantification of red (therapeutic F-specific antibodies: palivizumab, which binds both the pre-F and post-F conformations, and the pre-F-specific antibodies nirsevimab and clesrovimab) and blue (post-F-specific control antibody MAB8599) fluorescence intensity, normalized to GFP signal, for infected cells cultured in medium A, B or C (mean ± SD, n = 3). Statistical significance was assessed by one-way ANOVA. ( B ) Representative fluorescence images of infected cells stained with palivizumab, nirsevimab and clesrovimab (red) and the post-F control antibody (blue). ( C ) IC50 values of palivizumab, nirsevimab and clesrovimab, against virus produced with medium B or C, determined by 4-PL regression (mean ± SD, n = 3 independent experiments). Statistical significance was assessed using fitted IC50 values. ( D ) Scatter plot showing RT-qPCR Cq values versus TCID50 values for virus stocks A-C. * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001, **** P ≤ 0.0001. ns, not significant

Article Snippet: Integrated GFP fluorescence (GCU × μm2/image) was quantified over 5 days using an IncuCyte S3 system (Sartorius).

Techniques: Binding Assay, Neutralization, Recombinant, Produced, Control, Fluorescence, Infection, Cell Culture, Staining, Virus, Quantitative RT-PCR