Figure 2. Multiplexed diffraction-limited confocal imaging with DEI.
(a) Comparison of conventional staining using dye-conjugated antibodies and DEI using DNA-conjugated antibodies. Fixed neurons were stained with primary antibodies targeting SynapsinI, followed by both Alexa647-conjugated and DNA-conjugated antibodies, as shown in the schematic. DNA-conjugated antibody signals were visualized using Cy3b-imager strands. The correlation coefficient of the two images was 0.96. (b) Co-localization of SynapsinI and Synaptophysin in neurons visualized using two rounds of DEI. (c) Multiplexed eight-target imaging in neurons. Fixed DIV (Days in vitro) 14 mouse hippocampal neurons were stained with DNA-conjugated antibodies against SynapsinI, vGAT, MAP2, pNFH, GFAP, AlphaTubulin and AcetylTubulin. A 3D image stack of 14 μm thickness in z-axis was taken for each target and displayed as 2D color-coded maximum intensity projection (bottom to top: green to red). Scale bars: 10 μm. DNA docking strand sequences are listed in Table S4.