Antibody Conjugated, Raman Tagged Hollow Gold–Silver Nanospheres for Specific Targeting and Multimodal Dark-Field/SERS/Two Photon-FLIM Imaging of CD19(+) B Lymphoblasts

In this Research Article, we propose a new class of contrast agents for the detection and multimodal imaging of CD19(+) cancer lymphoblasts. The agents are based on NIR responsive hollow gold-silver nanospheres conjugated with antiCD19 monoclonal antibodies and marked with Nile Blue (NB) SERS active molecules (HNS-NB-PEG-antiCD19). Proof of concept experiments on specificity of the complex for the investigated cells was achieved by transmission electron microscopy (TEM). The microspectroscopic investigations via dark field (DF), surface-enhanced Raman spectroscopy (SERS), and two-photon excited fluorescence lifetime imaging microscopy (TPE-FLIM) corroborate with TEM and demonstrate successful and preferential internalization of the antibody-nanocomplex. The combination of the microspectroscopic techniques enables contrast and sensitivity that competes with more invasive and time demanding cell imaging modalities, while depth sectioning images provide real time localization of the nanoparticles in the whole cytoplasm at the entire depth of the cells. Our findings prove that HNS-NB-PEG-antiCD19 represent a promising type of new contrast agents with great possibility of being detected by multiple, non invasive, rapid and accessible microspectroscopic techniques and real applicability for specific targeting of CD19(+) cancer cells. Such versatile nanocomplexes combine in one single platform the detection and imaging of cancer lymphoblasts by DF, SERS, and TPE-FLIM microspectroscopy.