Correlated light and electron microscopic imaging of multiple endogenous proteins using Quantum dots

The importance of Locating proteins in their context within cells has been heightened recently by the accomplishments in molecular structure and systems biology. Although tight microscopy (LM) has been extensively used for mapping protein Localization, many studies require the additional resolution of the electron microscope. Here we report the application of small nanocrystals (Quantum dots; QDs) to specifically and efficiently label multiple distinct endogenous proteins. QDs are both fluorescent and electron dense, facilitating their use for correlated microscopic analysis. Furthermore, QDs can be discriminated optically by their emission wavelength and physically by size, making them invaluable for multilabeling analysis. We developed pre-embedding labeling criteria using QDs that allows optimization at the light Level, before continuing with electron microscopy (EM). We provide examples of double and triple immunolabeling using tight, electron and correlated microscopy in rat cells and mouse tissue. We conclude that QDs aid precise high-throughput determination of protein distribution.