Figure 3.

The binding of Toll-like receptor 4 (TLR4) and glutamate receptor N-methyl-D-aspartate subunit 1 (GluN1) was investigated with antibody-based fluorescence resonance energy transfer (FRET) in N9 and EOC 20 microglial cells. (A–F) N9 and (G–L) EOC 20 microglial cells were treated with 1 µg̸ml lipopolysaccharide (LPS) for 30 min and stained for TLR4 (red) and GluN1 (green). (A and G) Pre-bleach and (B and H) post-bleach images for TLR4. (C and I) Pre-bleach and (D and J) post-bleach images for GluN1. (F and L) FRET efficiency images. When one cell was randomly selected as the region of interest 1 (ROI1), four other cells and background were selected as controls, which were not bleached. (E and K) Five cells as ROI1 were analyzed from each coverslip and FRET efficiency was calculated. ^*P<0.05 vs. ROI1. Similar results were obtained from at least three independent experiments and two coverslips were examined for each condition in one test (n=6). Scale bar, 20 µM.