Journal: Nature Communications

Article Title: Contemporary HIV-1 consensus Env with AI-assisted redesigned hypervariable loops promote antibody binding

doi: 10.1038/s41467-024-48139-x

Figure Lengend Snippet: For each panel, comparisons of IC50 values for short and long HV loops are shown for representative antibodies: PGT145 ( A ) and VRC26.25 ( B ), two V2 apex-targeting antibodies whose epitopes are close to V2HV loops; 10-1074 ( C ) and the PGT135 ( D ), two glycan supersite antibodies whose epitopes are close to V1HV loops; VRC01 ( E ) and 3BNC117 ( F ), two CD4 binding site antibodies with epitopes close to the V5HV loops. The HV loop nearest to the epitope of each antibody is shown in bold font. The box plots depicted the median and 25th/75th percentiles of the distribution, with the minima and maxima as whiskers. The p -values from non-paired, non-parametric one-tailed Mann-Whitney U tests are provided. IC50 values equal to or greater than 100 μg/ml are grouped (upper limit of antibody neutralization measurements in the CATNAP database). Source data for all panels are provided in the Source Data files.

Article Snippet: 293 T cells were transfected with 1 ug of plasmid DNA encoding an HIV-1 env gene (PolyJet™ In Vitro DNA Transfection Reagent (SignaGen Laboratories, Cat. No. SL100688) in 6-well plates and harvested at 48 h. Cells were split into separate staining reactions for monoclonal antibodies 3BNC117 (Cat. No. ARP-12474), 10-1074 (Cat. No. ARP-12477), 10E8 (Cat. No. ARP-12294), 447-52D (Cat. No. ARP-4030, all from BEI resources), b12 (Polymun, Cat. No. AB011), PGDM1400 and PGT151 (both kindly provided by Dr. Devon Sok and International AIDS Vaccine Initiative).

Techniques: Binding Assay, One-tailed Test, MANN-WHITNEY, Neutralization

Journal: Nature Communications

Article Title: Contemporary HIV-1 consensus Env with AI-assisted redesigned hypervariable loops promote antibody binding

doi: 10.1038/s41467-024-48139-x

Figure Lengend Snippet: Consensus Env sequences were expressed as gp140 glycoproteins. Comparison of the binding of A plasma pools from 13 cohorts of PLWH and B monoclonal antibodies to consensus B Env with unmodified (in red) and redesigned (in blue) HV loops. The number of plasma samples in each pool is shown in parenthesis in panel A . Binding of plasma samples ( n = 13) C and monoclonal antibodies ( n = 7, with PGDM1400 and MPER antibodies excluded) ( D ) to the subtype B, C and CRF01_AE consensus Env with unmodified HV loops (red) and redesigned HV loops (blue) is compared using paired, non-parametric one-tailed Wilcoxon signed-rank tests. MPER antibodies and PGDM1400 were not included in the statistical tests as their epitopes are not presented on the gp140 glycoproteins. E Neutralization sensitivity of the consensus Env with or without redesigned HV loops for subtypes B, C, and CRF01_AE measured for ten bnAbs that targeted the CD4 binding site (VRC01 and 3BNC117), V2-apex (PG9 and PGT145), glycan supersite (10-1074 and PGT128), MPER (10E8 and 4E10) and mannose-dependent (2G12) epitopes as well as the sCD4. The neutralization sensitivity is reported as IC50 values (μg/ml) with the most sensitive viruses colored in red and the most resistant in gray. Neutralization sensitivity (IC50 values) are shown for the unmodified and redesigned HV loops and displayed separately for each Env clade ( n = 10) ( F ) or each bnAb ( G ). The increase in sensitivity with the redesigned HV loops is tested for each clade with one-tailed exact Wilcoxon-Pratt signed-rank tests. Only the 01_AE_2010s.hv_redesigned.1 is shown in panels F and G as it has a shorter V1HV than 01_AE_2010s.hv_redesigned.2. Source data for all panels are provided in the Source Data files.

Article Snippet: 293 T cells were transfected with 1 ug of plasmid DNA encoding an HIV-1 env gene (PolyJet™ In Vitro DNA Transfection Reagent (SignaGen Laboratories, Cat. No. SL100688) in 6-well plates and harvested at 48 h. Cells were split into separate staining reactions for monoclonal antibodies 3BNC117 (Cat. No. ARP-12474), 10-1074 (Cat. No. ARP-12477), 10E8 (Cat. No. ARP-12294), 447-52D (Cat. No. ARP-4030, all from BEI resources), b12 (Polymun, Cat. No. AB011), PGDM1400 and PGT151 (both kindly provided by Dr. Devon Sok and International AIDS Vaccine Initiative).

Techniques: Comparison, Binding Assay, One-tailed Test, Neutralization

Journal: iScience

Article Title: Chimpanzee adenovirus-mediated multiple gene therapy for age-related macular degeneration

doi: 10.1016/j.isci.2023.107939

Figure Lengend Snippet: In vivo expression of eGFP, PEDF, sFlt-1, and sCD59 following IVT administration of AdCs (A) Representative fundus images of the retina from individual mice following IVT injection of 1.5×10 9 vp and 7.5×10 9 vp AdC68-eGFP. The eGFP signal could be detected from 48 h to 35 days post-injection. The dotted circles represent the edge of mouse retina. (B–D) Assessment of PEDF, sFlt-1, and sCD59 mRNA expression in retina-choroid-sclera complexes isolated from 10 mice. In each mouse, one eye was injected with AdC68-PFC (five mice for 1.5×10 9 vp and five mice for 7.5×10 9 vp) whereas the contralateral, un-injected eye served as control (only five eyes were used for analysis). At 4 days post-injection, RNA was purified from the retina-choroid complexes and real-time qPCR was conducted. Absolute number of mRNA copies were calculated using the standard curve method. (E–H) Images of western blot and quantification of the PEDF, sFlt-1, and sCD59 protein amount expressed in retina-choroid complexes of five mice. In each mouse, one eye was injected with AdC68-PFC (7.5×10 9 vp) whereas the contralateral, un-injected eye served as control. Total protein was obtained from retina-choroid-sclera complexes isolated from AdC68-PFC-treated (7.5×10 9 vp) and un-injected eyes 7 days post-injection. Antibodies against GAPDH were used for the internal control. The relative expression of PEDF, sFlt-1, and sCD59 in the un-injected eyes was set to 1. Data are expressed as mean ± SEM, and analyzed using one-way ANOVA multiple comparisons with Tukey’s method among groups in (B) and Student’s t test (two-tailed) in (C) (∗p < 0.05, ∗∗p < 0.01). PEDF, pigment epithelium-derived factor; sFlt-1, soluble fms-like tyrosine kinase-1; sCD59, soluble forms of CD59; IVT, intravitreal.

Article Snippet: Rabbit polyclonal anti-CD59 , Sino Biological , Cat#12474-RP02.

Techniques: In Vivo, Expressing, Injection, Isolation, Control, Purification, Western Blot, Two Tailed Test, Derivative Assay

Journal: iScience

Article Title: Chimpanzee adenovirus-mediated multiple gene therapy for age-related macular degeneration

doi: 10.1016/j.isci.2023.107939

Figure Lengend Snippet:

Article Snippet: Rabbit polyclonal anti-CD59 , Sino Biological , Cat#12474-RP02.

Techniques: Virus, Recombinant, Transfection, Protease Inhibitor, Plasmid Preparation, Gel Extraction, Software