Fabrication of polyelectrolyte microcapsules and their use as carriers of drugs, fluorescent labels, and metal nanoparticles is a promising approach to designing theranostic agents. Semiconductor quantum dots (QDs) are characterized by extremely high brightness and photostability that make them attractive fluorescent labels for visualization of intracellular penetration and delivery of such microcapsules. Here, we describe an approach to design, fabricate, and characterize physico-chemical and functional properties of polyelectrolyte microcapsules encoded with water-solubilized and stabilized with three-functional polyethylene glycol derivatives core/shell QDs. Developed microcapsules were characterized by dynamic light scattering, electrophoretic mobility, scanning electronic microscopy, and fluorescence and confocal microscopy approaches, providing exact data on their size distribution, surface charge, morphological, and optical characteristics. The fluorescence lifetimes of the QD-encoded microcapsules were also measured, and their dependence on time after preparation of the microcapsules was evaluated. The optimal content of QDs used for encoding procedure providing the optimal fluorescence properties of the encoded microcapsules was determined. Finally, the intracellular microcapsule uptake by murine macrophages was demonstrated, thus confirming the possibility of efficient use of developed system for live cell imaging and visualization of microcapsule transportation and delivery within the living cells.Electronic supplementary materialThe online version of this article (10.1186/s11671-018-2447-z) contains supplementary material, which is available to authorized users.

Imaging agents and drug carriers are commonly targeted toward cancer cell through functionalization with specific recognition molecules. Quantum dots (QDs) are fluorescent semiconductor nanocrystals whose extraordinary brightness and photostability make them attractive for direct fluorescent labeling of biomolecules or optical encoding of the membranes and cells. Here, we analyse the cytotoxicity of QD-encoded microcapsules, validate an approach to the activation of the microcapsule's surface for further functionalization with monoclonal antibody Trastuzumab, a humanized monoclonal antibody targeting the extracellular domain of the human epidermal growth factor receptor 2 (HER2) and already in clinical use for the treatment of HER2 positive breast cancer. In addition, we characterize the cell-specific targeting activity of the resultant bio-conjugate by immunofluorescence assay (IFA) and real-time analysis of interaction of the conjugates with live HER2 overexpressing human breast cancer cells. We demonstrate, that encapsulation of QDs into the polymer shell using the layer-by-layer deposition method yields highly fluorescent polyelectrolyte microcapsules with a homogeneous size distribution and biocompatibility upon in vitro treatment of cancer cells. Carbodiimide surface activation ensures optimal disperse and optical characteristics of the QD-encoded microcapsules before antibody conjugation. The prepared conjugates of the microcapsules with cancer-specific monoclonal antibody targeting HER2 provide sufficiently sensitive and specific antibody-mediated binding of the microcapsules with live cancer cells, which demonstrated their potential as prospective cancer cell–targeting agents.

Early detection of malignant tumours and, especially, micrometastases and disseminated tumour cells is still a challenge. In order to implement highly sensitive diagnostic tools we demonstrate the use of nanoprobes engineered from nanobodies (single-domain antibodies, sdAbs) and fluorescent quantum dots (QDs) for single- and two-photon detection and imaging of human micrometastases and disseminated tumour cells in ex vivo biological samples of breast and pancreatic metastatic tumour mouse models expressing human epidermal growth factor receptor 2 (HER2) or carcinoembryonic antigen (CEA). By staining thin (5–10 µm) paraffin and thick (50 µm) agarose tissue sections, we detected HER2- and CEA-positive human tumour cells infiltrating the surrounding tissues or metastasizing to different organs, including the brain, testis, lung, liver, and lymph nodes. Compared to conventional fluorescently labelled antibodies the sdAb-HER2-QD and sdAb-CEA-QD nanoprobes are superior in detecting micrometastases in tissue sections by lower photobleaching and higher brightness of fluorescence signals ensuring much better discrimination of positive signals versus background. Very high two-photon absorption cross-sections of QDs and small size of the nanoprobes ensure efficient imaging of thick tissue sections unattainable with conventional fluorescent probes. The nanobody–QD probes will help to improve early cancer diagnosis and prognosis of progression by assessing metastasis.

utilizing near-infrared NPs. [3] While optical imaging can provide functional information, its main disadvantage is the low penetration depth and high background from tissue autofluorescence. [4] More recently however, second and third harmonic generation (SHG and THG) nanomaterials have been generated that can be efficiently employed in the second near infrared window (NIR-II) [5] using a multiphoton microscope. They have several inherent advantages, i) providing a significantly higher penetration depth; ii) no bleaching or blinking; and iii) emitting both, SHG and THG signals that allow them to be detected regardless of the endogenous background signal emitted by collagen and lipids for example. [6,7] Bismuth ferrite (BiFeO 3 ) harmonic nanoparticles (BFO-HNPs) have excellent SHG and THG capabilities [8] and were recently shown to be highly biocompatible in human cell lines, [9] and can be used for monitoring of human stem cells in depth of skeletal muscle tissue at more than one millimeter from the surface. [6] BFO is the material of choice among all metal-oxides since its nonlinear coefficients is one of the highest, along with its large wavelength excitation range, reaching further into the infrared window. [8] In this work, we demonstrate the possibility of using BFO-HNPs for monitoring pulmonary macrophages. The lung Recently, second harmonic generation (SHG) nanomaterials have been generated that are efficiently employed in the classical (NIR) and extended (NIR-II) near infrared windows using a multiphoton microscope. The aim was to test bismuth ferrite harmonic nanoparticles (BFO-HNPs) for their ability to monitor pulmonary macrophages in mice. BFO-loaded MH-S macro phages are given intratracheally to healthy mice or BFO-HNPs are intranasally instilled in mice with allergic airway inflammation and lung sections of up to 100 μM are prepared. Using a two-photon-laser scanning microscope, it is shown that bright BFO-HNPs signals are detected from superficially localized cells as well as from deep within the lung tissue. BFO-HNPs are identified with an excellent signal-to-noise ratio and virtually no background signal. The SHG from the nanocrystals can be distinguished from the endogenous collagen-derived SHG around the blood vessels and bronchial structures. BFO-HNPs are primarily taken up by M2 alveolar macro phages in vivo. This SHG imaging approach provides novel information about the inter action of macrophages with cells and the extracellular matrix in lung disease as it is capable of visualizing and tracking NP-loaded cells at high resolution in thick tissues with minimal background fluorescence. Second Harmonic NanoparticlesThe ORCID identification number(s) for the author(s) of this article can be found under https://doi.

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Bismuth ferrite harmonic nanoparticles (BFO‐HNPs) are a non‐linear material capable of second‐harmonic generation (SHG). In article number https://doi.org/10.1002/smll.201803776, Fernanda Ramos‐Gomes, Marietta Andrea Markus, and co‐workers use BFO‐HNPs to visualize macrophages in the lungs of mice with allergic airway inflammation. Using multiphoton microscopy, the intense SHG signals from BFO‐HNPs can track macrophages in the context of neighboring tissue structures in thick lung tissues at high resolution and with minimal background fluorescence.