rabbit anti hsp70 af1663  (R&D Systems)

Journal: Nature Communications

Article Title: Augmentation of vaccine-induced humoral and cellular immunity by a physical radiofrequency adjuvant

doi: 10.1038/s41467-018-06151-y

Figure Lengend Snippet: RFA increases HSP70 levels and activates MyD88. a C57BL/6 mice were exposed to RF and skin HSP70, HSc70, and HSP90 levels were then analyzed by western blotting at 6 and 24 h (hr) using GAPDH as internal control. b Representative IHC images of HSP70 expression in RF-treated and non-treated skin at 24 h. Scale: 250 µm. c WT and MyD88 KO mice were exposed to RF or sham treatment followed by ID injection of 10 µg OVA into RF or sham-treated skin. Serum anti-OVA antibody titer was measured 2 weeks later. n = 4–5. d WT mice were intradermally injected with 100 µg Pepinh-Control or Pepinh-MyD, or the same volume of PBS 3 and 1 h before RF treatment and ID OVA immunization at 10 µg dose. OVA immunization alone served as control (No adjuvant). Serum anti-OVA antibody titer was measured 2 weeks later. n = 4–5. One-way ANOVA with Tukey’s multiple comparison test was used to compare differences between groups in c and d . * p < 0.05; ** p < 0.01. NS not significant. Representative of two independent experiments in c and d

Article Snippet: PVDF membrane was then incubated with rabbit anti-HSP70 antibodies (1:2000, AF1663, R&D Systems) (no cross-reactivity with HSc70) at room temperature for 90 min. After washing in Tris-buffered saline (TBS) containing 0.05% Tween 20 (TBST), PVDF membrane was incubated with HRP-conjugated anti-rabbit secondary antibodies (1:2000, 7074P2, Cell Signaling Technology) at room temperature for 1 h. After washing in TBST, PVDF membrane was incubated with Pierce ECL Western Blotting Substrate (32109, Thermo Fisher Scientific).

Techniques: Western Blot, Control, Expressing, Injection, Adjuvant, Comparison

Journal: Nature Communications

Article Title: Augmentation of vaccine-induced humoral and cellular immunity by a physical radiofrequency adjuvant

doi: 10.1038/s41467-018-06151-y

Figure Lengend Snippet: RFA increases HSP70 levels and activates MyD88. a C57BL/6 mice were exposed to RF and skin HSP70, HSc70, and HSP90 levels were then analyzed by western blotting at 6 and 24 h (hr) using GAPDH as internal control. b Representative IHC images of HSP70 expression in RF-treated and non-treated skin at 24 h. Scale: 250 µm. c WT and MyD88 KO mice were exposed to RF or sham treatment followed by ID injection of 10 µg OVA into RF or sham-treated skin. Serum anti-OVA antibody titer was measured 2 weeks later. n = 4–5. d WT mice were intradermally injected with 100 µg Pepinh-Control or Pepinh-MyD, or the same volume of PBS 3 and 1 h before RF treatment and ID OVA immunization at 10 µg dose. OVA immunization alone served as control (No adjuvant). Serum anti-OVA antibody titer was measured 2 weeks later. n = 4–5. One-way ANOVA with Tukey’s multiple comparison test was used to compare differences between groups in c and d . * p < 0.05; ** p < 0.01. NS not significant. Representative of two independent experiments in c and d

Article Snippet: Tissue sections were then incubated with SuperBlock (Thermo Fisher Scientific) and then rabbit anti-mouse polyclonal HSP70 antibodies (1:100, AF1663, R&D Systems) at 4 ̊C overnight.

Techniques: Western Blot, Control, Expressing, Injection, Adjuvant, Comparison