Journal: Oncotarget

Article Title: KRICT-9 inhibits neuroinflammation, amyloidogenesis and memory loss in Alzheimer’s disease models

doi: 10.18632/oncotarget.19818

Figure Lengend Snippet: Microglial BV-2 cells were incubated with anti-IBA-1 (green) and anti- Aβ 1−42 (red) primary antibodies and the cultured astrocytes were incubated with anti-GFAP (green) and anti- Aβ 1−42 (red) primary antibodies. (A-B) The effect of KRICT-9 on fluorescence was detected using Alexa 488-conjugated anti-mouse/goat and Alexa 568-conjugated anti-rabbit secondary antibodies. Expressions of APP, BACE1, C99, lba-1, and GFAP as detected by western blot using specific antibodies in (C) microglial BV-2 cells, and (D) astrocytes. Quantitative analysis of the western band was performed using ImageJ. Each blot is representative of three experiments. β-actin was used as an internal control. *p < 0.05 indicates significantly different from the LPS-treated group. Co-treatments with KRICT-9 and LPS for 24 h were used. (E-F) β-secretase activity. Cells were collected to determine Aβ 42 by ELISA from (G) microglial BV-2 cells, and (H) astrocytes. Values represent means ±SD for three independent experiments performed. *p <0.05 compared to control, #p < 0.05 compared LPS.

Article Snippet: Fixed cells were exposed to the following primary antibodies: GFAP (Cell Signaling Technology), Aβ (Novus Biologicals, Littleton, CO; NBP2-13075) (1:100 dilutions in blocking serum, Abcam), and Iba1 (1:100 dilution in blocking serum, Wako) at room temperature for 1 h. The cells were then washed twice with ice PBS and incubated with an anti-rabbit or mouse secondary antibody conjugated to Alexa Fluor 488 or 568 (Invitrogen-Molecular Probes, Carlsbad, CA) at room temperature for 1 h. Immunofluorescence images were acquired with an inverted fluorescent microscope Zeiss Axiovert 200 M (Carl Zeiss, Thornwood, NY).

Techniques: Incubation, Cell Culture, Fluorescence, Western Blot, Control, Activity Assay, Enzyme-linked Immunosorbent Assay