Journal: Vaccine

Article Title: Robust Antigen-Specific Humoral Immune Responses to Sublingually Delivered Adenoviral Vectors Encoding HIV-1 Env: Association with Mucoadhesion and Efficient Penetration of the Sublingual Barrier

doi: 10.1016/j.vaccine.2011.07.008

Figure Lengend Snippet: An Ad5 vector encoding HIV-1 gp120 (1 × 108 VP), or a HSV amplicon vector (1 × 106 VP) encoding the same antigen was delivered IM or SL to female BALB/c mice; control animals were inoculated with PBS. A homologous boost was performed on day 14 and sera samples were collected on day 28. Sera from immunized mice were diluted 1:10, 1:100, and 1:1000 in PBS and incubated with ELISA plates coated with recombinant HIV-1 gp160 (MN strain, Protein Sciences). Peroxidase conjugated anti-mouse IgG (panel A) or anti-mouse IgA (panel B) were then added, as appropriate, and results were quantitated by measuring absorbance at 450 nm. The gray dotted line represents the background absorbance in this assay (this was set at 2 times the mean background absorbance in samples from PBS-treated control animals). Results represent mean values from 4 animals/group; bars denote the standard error of the mean. Data shown are representative of 2 experiments, performed with similar results. *: Denotes a statistically significant difference in the magnitude of the Env-specific serum IgG or IgA response in mice immunized with Ad:gp120 via the SL route versus the serum IgG or IgA response in mice immunized with HSV-1:Luc via the same route (p=0.0286; Mann-Whitney test).

Article Snippet: ELISA plates (Dynex Technologies) were coated overnight with recombinant soluble HIV-1 MN gp160 (Protein Sciences) or purified oligomeric HIV-1 YU2 gp140 [ 25 ] at 500 ng/well in 0.1 M NaHCO 3 (pH 9.6).

Techniques: Plasmid Preparation, Amplification, Control, Incubation, Enzyme-linked Immunosorbent Assay, Recombinant, MANN-WHITNEY

Journal: Vaccine

Article Title: Robust Antigen-Specific Humoral Immune Responses to Sublingually Delivered Adenoviral Vectors Encoding HIV-1 Env: Association with Mucoadhesion and Efficient Penetration of the Sublingual Barrier

doi: 10.1016/j.vaccine.2011.07.008

Figure Lengend Snippet: An Ad5 vector encoding oligomeric HIV-1 Env (gp140-YU2) was administered to female BALB/c mice via the SL, IM or intragastric (IG) route at a dose of 1×109 VP. At day 28, animals received a booster immunization with the same vaccines/routes, and at day 38, animals were sacrificed and samples collected for analysis. (A, B) Quantitation of endpoint Env-specific serum IgG (A) and IgA (B) antibody titers in sera from groups of immunized mice. Sera were serially diluted, two-fold, in PBS and incubated with ELISA plates coated with recombinant gp140 (YU2 Strain). Peroxidase conjugated anti-Mouse IgG (Sigma) or anti-mouse IgA (Sigma) were then added, as appropriate, and results were quantitated by measuring absorbance at 450 nm. Data represent mean endpoint titers ± SEM, calculated from 4 mice per group. Results shown are representative of 3 experiments, performed with similar results. *: Denotes a statistically significant difference in the magnitude of the Env-specific serum IgA response in mice immunized with Ad:gp140 via the SL route versus the IM route (p=0.0286, Mann-Whitney test). (C, D) Quantitation of Env-specific mucosal IgG (C) and IgA (D) antibody titers in vaginal washes from groups of immunized mice. In this case, antibodies were measured only in undiluted wash samples, and results are presented directly as mean absorbance (A450) values plus or minus SEM calculated from 4 mice per group. Results shown are representative of 2 experiments, performed with similar results.

Article Snippet: ELISA plates (Dynex Technologies) were coated overnight with recombinant soluble HIV-1 MN gp160 (Protein Sciences) or purified oligomeric HIV-1 YU2 gp140 [ 25 ] at 500 ng/well in 0.1 M NaHCO 3 (pH 9.6).

Techniques: Plasmid Preparation, Vaccines, Quantitation Assay, Incubation, Enzyme-linked Immunosorbent Assay, Recombinant, MANN-WHITNEY

Journal: eLife

Article Title: Mannose receptor is an HIV restriction factor counteracted by Vpr in macrophages

doi: 10.7554/eLife.51035

Figure Lengend Snippet:

Article Snippet: Antibodies to the following proteins were used for immunoblot analysis: MR (cat# ab64693, Abcam, Cambridge, Massachusetts), GAPDH (clone 3C2, cat# H00002597-M01, Abnova, Taipei, Taiwan), Gag pr55 (HIV-Ig AIDS Reagent Program cat# 3957), Env gp160/120 (AIDS Reagent Program cat# 288 from Dr. Michael Phelan), 89.6 and YU-2 Env gp41 (clone z13e1, AIDS Reagent Program cat# 11557 from Dr. Michael Zwick), NL4-3 Env gp41 (clone CHESSIE-8, AIDS Reagent Program cat# 526 from Dr. George Lewis), Vpr (AIDS Reagent Program cat# 3951 from Dr. Jeffrey Kopp), GFP (cat# ab13970, Abcam), Nef (AIDS Reagent Program cat# 2949 from Dr. Ronald Swanstrom), FLAG (clone M2, cat# F3165, Sigma), STING (D2P2F, cat# 13647, Cell Signaling Technology), GBP5 (sc-160353, which was a generous gift from Dr. Frank Kirchhoff), and IFITM3 (cat# 11714–1-AP, Proteintech, Rosemont, IL).

Techniques: Recombinant, Mutagenesis, Luciferase, Selection, Isolation, Gene Expression, Sequencing, Purification, Enzyme-linked Immunosorbent Assay, Neutralization, Software, shRNA

Journal: eLife

Article Title: Mannose receptor is an HIV restriction factor counteracted by Vpr in macrophages

doi: 10.7554/eLife.51035

Figure Lengend Snippet:

Article Snippet: Antibodies to the following proteins were used for immunoblot analysis: MR (cat# ab64693, Abcam, Cambridge, Massachusetts), GAPDH (clone 3C2, cat# H00002597-M01, Abnova, Taipei, Taiwan), Gag pr55 (HIV-Ig AIDS Reagent Program cat# 3957), Env gp160/120 (AIDS Reagent Program cat# 288 from Dr. Michael Phelan), 89.6 and YU-2 Env gp41 (clone z13e1, AIDS Reagent Program cat# 11557 from Dr. Michael Zwick), NL4-3 Env gp41 (clone CHESSIE-8, AIDS Reagent Program cat# 526 from Dr. George Lewis), Vpr (AIDS Reagent Program cat# 3951 from Dr. Jeffrey Kopp), GFP (cat# ab13970, Abcam), Nef (AIDS Reagent Program cat# 2949 from Dr. Ronald Swanstrom), FLAG (clone M2, cat# F3165, Sigma), STING (D2P2F, cat# 13647, Cell Signaling Technology), GBP5 (sc-160353, which was a generous gift from Dr. Frank Kirchhoff), and IFITM3 (cat# 11714–1-AP, Proteintech, Rosemont, IL).

Techniques: Recombinant, Mutagenesis, Luciferase, Selection, Isolation, Gene Expression, Sequencing, Purification, Enzyme-linked Immunosorbent Assay, Neutralization, Software, shRNA

Journal: eLife

Article Title: Mannose receptor is an HIV restriction factor counteracted by Vpr in macrophages

doi: 10.7554/eLife.51035

Figure Lengend Snippet:

Article Snippet: Antibodies to the following proteins were used for immunoblot analysis: MR (cat# ab64693, Abcam, Cambridge, Massachusetts), GAPDH (clone 3C2, cat# H00002597-M01, Abnova, Taipei, Taiwan), Gag pr55 (HIV-Ig AIDS Reagent Program cat# 3957), Env gp160/120 (AIDS Reagent Program cat# 288 from Dr. Michael Phelan), 89.6 and YU-2 Env gp41 (clone z13e1, AIDS Reagent Program cat# 11557 from Dr. Michael Zwick), NL4-3 Env gp41 (clone CHESSIE-8, AIDS Reagent Program cat# 526 from Dr. George Lewis), Vpr (AIDS Reagent Program cat# 3951 from Dr. Jeffrey Kopp), GFP (cat# ab13970, Abcam), Nef (AIDS Reagent Program cat# 2949 from Dr. Ronald Swanstrom), FLAG (clone M2, cat# F3165, Sigma), STING (D2P2F, cat# 13647, Cell Signaling Technology), GBP5 (sc-160353, which was a generous gift from Dr. Frank Kirchhoff), and IFITM3 (cat# 11714–1-AP, Proteintech, Rosemont, IL).

Techniques: Recombinant, Mutagenesis, Luciferase, Selection, Isolation, Gene Expression, Sequencing, Purification, Enzyme-linked Immunosorbent Assay, Neutralization, Software, shRNA