Cathodoluminescent and Characteristic X-ray-emissive Rare-Earth-doped Core/Shell Immunolabels for Spectromicroscopic Analysis of Cell Surface Receptors

Abstract

Understanding the localization and the interactions of biomolecules at the nanoscale and in the cellular context remains challenging. Electron microscopy (EM) as a non-Abbe limited technique gives access to the cellular ultrastructure yet results in grey-scale images and averts unambiguous (co-)localization of biomolecules. Multimodal nanoparticle-based immunolabels for correlative cathodoluminescence electron microscopy (CCLEM) and energy-dispersive X-ray spectromicroscopy (EDX-SM) are presented. The single-particle STEM-cathodoluminescence (CL) and characteristic X-ray emissivity of sub-20 nm lanthanide-doped nanoparticles were exploited as unique spectral fingerprints for precise localization and label identification. To maximize the nanoparticle brightness, lanthanides were incorporated in a low-phonon host lattice and separated from the environment using a passivating shell. The core/shell nanoparticles were then functionalized with either folic (terbium-doped) or caffeic acid (europium-doped). Their potential for immunolabeling was successfully demonstrated using HeLa cells expressing different surface receptors that bind to folic or caffeic acid, respectively. Both particle populations showed single-particle CL emission along with a distinctive energy-dispersive X-ray signal, with the latter enabling colour-based localization of receptors within swift imaging times well below 2 mins per µm2while offering high resolution with a pixel size of 2.78 nm. Taken together, these results open a route to color immunolabelling based on electron spectromicroscopy.