Phospho sapk jnk 81e11

Protein samples were prepared from (n = ~50) adult eyes from Canton-S (wild-type), GMR> Aβ42, GMR> Aβ42+hpo following standardized protocols (Gogia et al., 2017 ). The samples were loaded in the following sequence: Lane 1-Molecular weight marker (BIORAD Precision Plus Protein Kaleidoscope Prestained Catalog Number #1610375), Lane 2-Wild-type (Canton-S), Lane 3-GMR> Aβ42, Lane 4-GMR> Aβ42+hpo (gain-of-function), Lane 5-GMR> Aβ42+hpo^RNAi (loss-of-function). We incubated the blots with the Phospho SAPK/JNK (81E11) (1:3,000, Cell Signaling) and after appropriate washes incubated with horseradish peroxidase conjugated goat anti–rabbit IgG (1:5,000) secondary antibody. Signal was detected using super signal chemiluminescence substrate (Pierce). We captured the images using the BioSpectrum® 500 Imaging System and analyzed the blot images and band intensity. We used Microsoft Excel 2017 software for statistical analysis. We calculated the p-values using the Student two-tailed t-test. The error bars represent standard deviation from means.

Total liver lysates were analyzed by immunoblotting using antibodies against HBsAg (20-HR20, Fitzgerald), GADD153 (F-168, Santa Cruz), phospho-PERK (16F8, Cell Signaling), PERK (H-300, Santa Cruz), phospho-eIF2α (119A11, Cell Signaling), eIF2α (L57A5, Cell Signaling), β-actin (13E5, Cell Signaling), JNK2 (N-18, Santa Cruz), c-Jun (60A8, Cell Signaling), phospho-c-Jun (D47G9, Cell Signaling), phospho-SAPK/JNK (81E11, Cell Signaling), STAT3 (79D7, Cell Signaling), phospho-STAT3 (D3A7, Cell Signaling).

Sample preparation, sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blotting was performed as described before [48 (link)]. Total liver lysates were analyzed by using antibodies against HBsAg (20-HR20, Fitzgerald, Acton, Ma, USA), phospho-PERK (16F8, Cell Signaling, Beverly, MA, USA), phospho-eIF2α (119A11, Cell Signaling), β-actin (13E5, Cell Signaling), c-Jun (60A8, Cell Signaling), phospho-c-Jun (D47G9, Cell Signaling), phospho-SAPK/JNK (81E11, Cell Signaling), STAT3 (79D7, Cell Signaling), phospho-STAT3 (D3A7, Cell Signaling).