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Active targeting of tumors is believed to be the key to efficient cancer therapy and accurate, early-stage diagnostics. Active targeting implies minimized off-targeting and associated cytotoxicity towards healthy tissue. One way to acquire active targeting is to employ conjugates of therapeutic agents with ligands known to bind receptors overexpressed onto cancer cells. The integrin receptor family has been studied as a target for cancer treatment for almost fifty years. However, systematic knowledge on their effects on cancer cells, is yet lacking, especially when utilized as an active targeting ligand for particulate formulations. Decoration with various integrin-targeting peptides has been reported to increase nanoparticle accumulation in tumors ≥ 3-fold when compared to passively targeted delivery. In recent years, many newly discovered or rationally designed integrin-binding peptides with excellent specificity towards a single integrin receptor have emerged. Here, we show a comprehensive analysis of previously unreviewed integrin-binding peptides, provide diverse modification routes for nanoparticle conjugation, and showcase the most notable examples of their use for tumor and metastases visualization and eradication to date, as well as possibilities for combined cancer therapies for a synergetic effect. This review aims to highlight the latest advancements in integrin-binding peptide development and is directed to aid transition to the development of novel nanoparticle-based theranostic agents for cancer therapy. [ABSTRACT FROM AUTHOR]

The rapidly developing field of nanomedicine presents new challenges for researchers. Existing, clinically used preparations based on metal nanoparticles still have their limitations. Consequently, scientific attention is shifting from well-studied monometallic to bimetallic nanoparticles, which combine a synergistic combination of different metals in their composition. This review examines promising gold-containing bimetallic nanoparticles for use in biomedicine. Gold (Au) is the most popular initial choice in bimetallic nanoparticle (BNPs) composition due to its biocompatibility. As two metals combine in one particle, it becomes possible to reduce systemic toxicity and significantly increase the therapeutic effect. We provide a comprehensive assessment of the advantages and limitations of bimetallic nanoparticles and discuss potential solutions to the problems that have hindered their development. [ABSTRACT FROM AUTHOR]

A method for quantitative investigation of affinity constants of receptors immobilized on magnetic nanoparticles (MP) is developed based on spectral correlation interferometry (SCI). The SCI records with a picometer resolution the thickness changes of a layer of molecules or nanoparticles due to a biochemical reaction on a cover slip, averaged over the sensing area. The method is compatible with other types of sensing surfaces employed in biosensing. The measured values of kinetic association constants of magnetic nanoparticles are 4 orders of magnitude higher than those of molecular antibody association with antigen. The developed method also suggests highly sensitive detection of antigens in a wide dynamic range. The limit of detection of 92 pg/ml has been demonstrated for prostate-specific antigen (PSA) with 50-nm MP employed as labels, which produce 3-order amplification of the SCI signals. The calibration curve features high sensitivity (slope) of 3-fold signal raise per 10-fold increase of PSA concentration within 4-order dynamic range, which is an attractive compromise for precise quantitative and highly sensitive immunoassay. The proposed biosensing technique offers inexpensive disposable sensor chips of cover slips and represents an economically sound alternative to traditional immunoassays for disease diagnostics, detection of pathogens in food and environmental monitoring. [ABSTRACT FROM AUTHOR]

Nanoparticles with biocomputing capabilities could potentially be used to create sophisticated robotic devices with a variety of biomedical applications, including intelligent sensors and theranostic agents. DNA/RNA-based computing techniques have already been developed that can offer a complete set of Boolean logic functions and have been used, for example, to analyse cells and deliver molecular payloads. However, the computing potential of particle-based systems remains relatively unexplored. Here, we show that almost any type of nanoparticle or microparticle can be transformed into autonomous biocomputing structures that are capable of implementing a functionally complete set of Boolean logic gates (YES, NOT, AND and OR) and binding to a target as result of a computation. The logic-gating functionality is incorporated into self-assembled particle/biomolecule interfaces (demonstrated here with proteins) and the logic gating is achieved through input-induced disassembly of the structures. To illustrate the capabilities of the approach, we show that the structures can be used for logic-gated cell targeting and advanced immunoassays. [ABSTRACT FROM AUTHOR]

A bioengineering method for self-assembly of multifunctional superstructures with in-advance programmable properties has been proposed. The method employs two unique proteins, barnase and barstar, to rapidly join the structural components together directly in water solutions. The properties of the superstructures can be designed on demand by linking different agents of various sizes and chemical nature, designated for specific goals. As a proof of concept, colloidally stable trifunctional structures have been assembled by binding together magnetic particles, quantum dots, and antibodies using barnase and barstar. The assembly has demonstrated that the bonds between these proteins are strong enough to hold macroscopic (5 nm-3 μm) particles together. Specific interaction of such superstructures with cancer cells resulted in fluorescent labeling of the cells and their responsiveness to magnetic field. The method can be used to join inorganic moieties, organic particles, and single biomolecules for synergistic use in different applications such as biosensors, photonics, and nanomedicine. [ABSTRACT FROM AUTHOR]

Abstract: Methods of non-invasive in vivo quantification of magnetic nanoparticles (MP) have been proposed and realized. The methods are based on non-linear MP magnetization at two frequencies and measuring the response at combinatorial frequencies. The first method is developed for real-time study of MP dynamics and their clearance from the blood system of animals. High sensitivity of 3ng of Fe3O4 in 0.1ml was achieved for MP detection in mice tail veins. The second technique is proposed for MP detection inside animal tissues by an external probe. The proposed methods could essentially widen capabilities of biomedical research which involves magnetic nanoparticles. [Copyright &y& Elsevier]

A magnetic enzymatic composite was synthesized from non-toxic magnetite nanoparticles modified by biomimetic polymerization of modified dopamine and carbonic anhydrase enzyme, which has a significant role in neurodegenerative processes. The mass fraction of the enzyme in the composite was 10% wt, which corresponded to about 120 enzyme molecules per nanoparticle. The composite showed K m about 10 folds higher compared to free enzyme (0.22 and 0.019 mmol/L respectively) and V m a x four times lower (0.17 and 0.69 mmol/min). The enzymatic composite had 17% higher thermal stability compared to free enzyme and retained 95% of its initial activity after seven consecutive reuse cycles. In a colloidal state, the composite showed no response to the RF field due to the ballistic mechanism of heat dissipation but in concentrated form, the material showed up to 155% enhanced catalytic activity under the influence of heating radio frequency magnetic fields. The observed results are discussed and explained. [Display omitted] • Magnetic enzymatic composites were prepared using biomimetic polydopamine coatings. • The material showed a ballistic heat dissipation mechanism. • The material showed better thermal stability compared to free enzyme. • The material showed RF field promoted enzymatic activity. [ABSTRACT FROM AUTHOR]

Recent developments in the field of nanomedicine have introduced a wide variety of nanomaterials that are capable of recognizing and killing tumor cells with increased specificity. A major limitation preventing the widespread introduction of nanomaterials into the clinical setting is their fast clearance from the bloodstream via the mononuclear phagocyte system (MPS). One of the most promising methods used to overcome this limitation is the MPS-cytoblockade, which forces the MPS to intensify the clearance of erythrocytes by injecting allogeneic anti-erythrocyte antibodies and, thus, significantly prolongs the circulation of nanoagents in the blood. However, on the way to the clinical application of this approach, the question arises whether the induced suppression of macrophage phagocytosis via the MPS-cytoblockade could pose health risks. Here, we show that highly cytotoxic doxorubicin- or clodronate-loaded liposomes, which are widely used for cancer therapy and biomedical research, induce a similar increase in the nanoparticle blood circulation half-life in mice as the MPS-cytoblockade, which only gently and temporarily saturates the macrophages with the organism's own erythrocytes. This result suggests that from the point of view of in vivo macrophage suppression, the MPS-cytoblockade should be less detrimental than the liposomal anti-cancer drugs that are already approved for clinical application while allowing for the substantial improvement in the nanoagent effectiveness. [ABSTRACT FROM AUTHOR]

Therapy for aggressive metastatic breast cancer remains a great challenge for modern biomedicine. Biocompatible polymer nanoparticles have been successfully used in clinic and are seen as a potential solution. Specifically, researchers are exploring the development of chemotherapeutic nanoagents targeting the membrane-associated receptors of cancer cells, such as HER2. However, there are no targeting nanomedications that have been approved for human cancer therapy. Novel strategies are being developed to alter the architecture of agents and optimize their systemic administration. Here, we describe a combination of these approaches, namely, the design of a targeted polymer nanocarrier and a method for its systemic delivery to the tumor site. Namely, PLGA nanocapsules loaded with a diagnostic dye, Nile Blue, and a chemotherapeutic compound, doxorubicin, are used for two-step targeted delivery using the concept of tumor pre-targeting through the barnase/barstar protein "bacterial superglue". The first pre-targeting component consists of an anti-HER2 scaffold protein, DARPin9_29 fused with barstar, Bs-DARPin9_29, and the second component comprises chemotherapeutic PLGA nanocapsules conjugated to barnase, PLGA-Bn. The efficacy of this system was evaluated in vivo. To this aim, we developed an immunocompetent BALB/c mouse tumor model with a stable expression of human HER2 oncomarkers to test the potential of two-step delivery of oncotheranostic nano-PLGA. In vitro and ex vivo studies confirmed HER2 receptor stable expression in the tumor, making it a feasible tool for HER2-targeted drug evaluation. We demonstrated that two-step delivery was more effective than one-step delivery for both imaging and tumor therapy: two-step delivery had higher imaging capabilities than one-step and a tumor growth inhibition of 94.9% in comparison to 68.4% for the one-step strategy. The barnase*barstar protein pair has been proven to possess excellent biocompatibility, as evidenced by the successful completion of biosafety tests assessing immunogenicity and hemotoxicity. This renders the protein pair a highly versatile tool for pre-targeting tumors with various molecular profiles, thereby enabling the development of personalized medicine. [ABSTRACT FROM AUTHOR]

Nanoparticle-based chemotherapy is considered to be an effective approach to cancer diagnostics and therapy in modern biomedicine. However, efficient tumor targeting remains a great challenge due to the lack of specificity, selectivity, and high dosage of chemotherapeutic drugs required. A two-step targeted drug delivery strategy (DDS), involving cancer cell pre-targeting, first with a first nontoxic module and subsequent targeting with a second complementary toxic module, is a solution for decreasing doses for administration and lowering systemic toxicity. To prove two-step DDS efficiency, we performed a direct comparison of one-step and two-step DDS based on chemotherapy loaded PLGA nanoparticles and barnase*barstar interface. Namely, we developed and thoroughly characterized the two-step targeting strategy of HER2-overexpressing cancer cells. The first targeting block consists of anti-HER2 scaffold polypeptide DARPin9_29 fused with barstar. Barstar exhibits an extremely effective binding to ribonuclease barnase with Kaff = 1014 M−1, thus making the barnase*barstar protein pair one of the strongest known protein*protein complexes. A therapeutic PLGA-based nanocarrier coupled to barnase was used as a second targeting block. The PLGA nanoparticles were loaded with diagnostic dye, Nile Blue, and a chemotherapeutic drug, doxorubicin. We showed that the two-step DDS increases the performance of chemotherapy-loaded nanocarriers: IC50 of doxorubicin delivered via two-step DDS was more than 100 times lower than that for one-step DDS: IC50 = 43 ± 3 nM for two-step DDS vs. IC50 = 4972 ± 1965 nM for one-step DDS. The obtained results demonstrate the significant efficiency of two-step DDS over the classical one-step one. We believe that the obtained data will significantly change the direction of research in developing targeted anti-cancer drugs and promote the creation of new generation cancer treatment strategies. [ABSTRACT FROM AUTHOR]

Nanostructured materials and systems find various applications in biomedical fields. Hybrid organo–inorganic nanomaterials are intensively studied in a wide range of areas, from visualization to drug delivery or tissue engineering. One of the recent trends in material science is biomimetic approaches toward the synthesis or modification of functional nanosystems. Here, we describe an approach toward multifunctional nanomaterials through the biomimetic polymerization of dopamine derivatives. Magnetite nanoparticles were modified with a combination of dopamine conjugates to give multifunctional magneto-fluorescent nanocomposites in one synthetic step. The obtained material showed excellent biocompatibility at concentrations up to 200 μ g/mL and an in vivo biodistribution profile typical for nanosized formulations. The synthesized systems were conjugated with antibodies against HER2 to improve their selectivity toward HER2-positive cancer cells. The produced material can be used for dual magneto-optical in vivo studies or targeted drug delivery. The applied synthetic strategy can be used for the creation of various multifunctional hybrid nanomaterials in mild conditions. [ABSTRACT FROM AUTHOR]

A dry-reagent immunomagnetic (DRIM) biosensing platform is developed for rapid high-precision quantitative analyses for in vitro diagnostics. The platform combines the advantages of immunochromatography with highly sensitive quantification of 200-nm magnetic nanoparticles (MP) from the entire volume of lateral flow membranes. The MP are registered by their non-linear magnetization at combinatorial frequencies by a portable reader that offers the detection limit of 60 zeptomoles or 0.4 ng of magnetic nanolabels and extremely wide linear dynamic range of 7 orders. The efficient combination of small MP with large reaction surface of the 3D membranes has permitted the detection in human serum of as low as 25 pg/ml total prostate specific antigen (PSA) during 30-min assay. The featured 3-fold signal increase per every order of concentration within 3.5 orders of magnitude allows precise analysis of antigen concentrations in a wide range. The system also provides the first tool for quantitative MP mapping along the lateral flow strip for easy development and optimization of assays. The DRIM detection can be used for simple, rapid and sensitive quantification of protein biomarkers for in vitro diagnostics both in laboratory and near-patient conditions, for food analysis, environmental monitoring, security, and safety applications. [ABSTRACT FROM AUTHOR]

Injuries of the respiratory system caused by viral infections (e.g., by influenza virus, respiratory syncytial virus, metapneumovirus, or coronavirus) can lead to long-term complications or even life-threatening conditions. The challenges of treatment of such diseases have become particularly pronounced during the recent pandemic caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). One promising drug is the anti-fibrinolytic and anti-inflammatory protease inhibitor aprotinin, which has demonstrated considerable inhibition of the replication of some viruses. Encapsulation of aprotinin in liposomes can significantly improve the effectiveness of the drug, however, the use of nanoparticles as carriers of aprotinin can radically change its biodistribution in the body. Here we show that the liposomal form of aprotinin accumulates more efficiently in the lungs, heart, and kidneys than the molecular form by side-by-side comparison of the ex vivo biodistribution of these two fluorescently labeled formulations in mice using bioimaging. In particular, we synthesized liposomes of different compositions and studied their accumulation in various organs and tissues. Direct comparison of the biodistributions of liposomal and free aprotinin showed that liposomes accumulated in the lungs 1.82 times more effectively, and in the heart and kidneys – 3.56 and 2.00 times, respectively. This suggests that the liposomal formulation exhibits a longer residence time in the target organ and, thus, has the potential for a longer therapeutic effect. The results reveal the great potential of the aprotinin-loaded liposomes for the treatment of respiratory system injuries and heart- and kidney-related complications of viral infections. • The biodistribution study of molecular and liposomal form of aprotinin was performed. • The liposomal form accumulates more efficiently in the lungs, heart, and kidneys. • The composition of liposomes affects the biodistribution of aprotinin. [ABSTRACT FROM AUTHOR]

N -Heterocyclic carbenes (NHC) are well-recognized ligands of choice for preparing robust transition metal species. However, their use for fabrication of biomedically relevant nanoparticles has been limited to the synthesis of non-targeted particles showing increased tolerance to different aqueous coagulants. In this work, the first example of carbene-coated metal nanoparticles suitable for in vivo applications is presented. Directed design of a novel biscarbene NHC ligand allowed to prepare the first magnetite/gold (Fe 3 O 4 @AuNP@NHC) nanostructures and carbene gold (AuNP@NHC) nanoparticles with significant stability in aqueous solutions and enhanced ability to form bioconjugates. Furthermore, these nanoparticles exhibit an extraordinary property for inorganic nanoparticles: they can endure several additive-free air drying/redispersion cycles without deterioration of their colloidal behavior. Bioconjugated AuNP@NHC and multimodal Fe 3 O 4 @AuNP@NHC demonstrated a successful performance in three distinct applications: lateral flow tests, specific cancer cell targeting, and bioimaging. Thus, the results show the notable advantages of the N -heterocyclic carbene coating of inorganic nanoparticles and their utility for complex biomedical applications. [Display omitted] • The first example of carbene-coated nanoparticles successfully used for in vivo research. • The first magnetic gold nanoparticles coated with carbene ligand. • Nanoparticles showed remarkable stability in aqueous solutions and ability to be conjugated with bioreceptors. • Developed carbene-coated nanoparticle conjugates were applied for lateral flow assays, cell targeting, and bioimaging. [ABSTRACT FROM AUTHOR]

Magnetic nanoparticles are widely used in biomedicine for MRI imaging and anemia treatment. The aging of these nanomaterials in vivo may lead to gradual diminishing of their contrast properties and inducing toxicity. Here, we describe observation of the full lifecycle of 40-nm magnetic particles from their injection to the complete degradation in vivo and associated impact on the organism. We found that in 2 h the nanoparticles were eliminated from the bloodstream, but their initial biodistribution changed over time. In 1 week, a major part of the nanoparticles was transferred to the liver and spleen, where they degraded with a half-life of 21 days. MRI and a magnetic spectral approach revealed preservation of contrast in these organs for more than 1 month. The particle degradation led to the increased number of red blood cells and blood hemoglobin level due to released iron without causing any toxicity in tissues. We also observed an increase in gene expression level of Fe-associated proteins such as transferrin, DMT1, and ferroportin in the liver in response to the iron particle degradation. A deeper understanding of the organism response to the particle degradation can bring new directions to the field of MRI contrast agent design. [ABSTRACT FROM AUTHOR]

[Display omitted] Nanoparticles based on poly(lactic-co-glycolic acid) (PLGA) with various surface chemistry are widely used in biomedicine for theranostic applications. The nature of the external coating of nanoparticles has a significant influence on their efficiency as drug carriers or visualization agents. However, information about the mechanisms of nanoparticle accumulation in tumors and the influence of their surface properties on biodistribution is scarce due to the lack of systematic evaluation. Here we investigate the effect of different polymer coatings of the surface on in vitro and in vivo properties of PLGA nanoparticles. Namely, cell binding efficiency, cytotoxicity, efficiency of fluorescent bioimaging, and tumor accumulation were tested. The highest binding efficiency in vitro and cytotoxicity were observed for positively charged polymers. Interestingly, in vivo fluorescent visualization of tumor-bearing mice and quantitative measurements of biodistribution of magnetite-loaded nanoparticles indicated different dependences of accumulation in tumors on the coating of PLGA nanoparticles. This means that nanoparticle surface properties can simultaneously enhance imaging efficiency and decrease quantitative accumulation in tumors. The obtained data demonstrate the complexity of the dependence of nanoparticles' effectiveness for theranostic applications on surface features. We believe that this study will contribute to the rational design of nanoparticles for effective cancer diagnostics and therapy. [ABSTRACT FROM AUTHOR]

Nanoparticles exhibiting the localized surface plasmon resonance (LSPR) phenomenon are promising tools for diagnostics and cancer treatment. Among widely used metal nanoparticles, silver nanoparticles (Ag NPs) possess the strongest light scattering and surface plasmon strength. However, the therapeutic potential of Ag NPs has until now been underestimated. Here we show targeted photothermal therapy of solid tumors with 35 nm HER2-targeted Ag NPs, which were produced by the green synthesis using an aqueous extract of Lavandula angustifolia Mill. Light irradiation tests demonstrated effective hyperthermic properties of these NPs, namely heating by 10 °C in 10 min. To mediate targeted cancer therapy, Ag NPs were conjugated to the scaffold polypeptide, affibody ZHER2:342, which recognizes a clinically relevant oncomarker HER2. The conjugation was mediated by the PEG linker to obtain Ag-PEG-HER2 nanoparticles. Flow cytometry tests showed that Ag-PEG-HER2 particles successfully bind to HER2-overexpressing cells with a specificity comparable to that of full-size anti-HER2 IgGs. A confocal microscopy study showed efficient internalization of Ag-PEG-HER2 into cells in less than 2 h of incubation. Cytotoxicity assays demonstrated effective cell death upon exposure to Ag-PEG-HER2 and irradiation, caused by the production of reactive oxygen species. Xenograft tumor therapy with Ag-PEG-HER2 particles in vivo resulted in full primary tumor regression and the prevention of metastatic spread. Thus, for the first time, we have shown that HER2-directed plasmonic Ag nanoparticles are effective sensitizers for targeted photothermal oncotherapy. [ABSTRACT FROM AUTHOR]

Method of highly sensitive registration of magnetic nanopartides by their nonlinear magnetization is used in a novel sandwich-type immunoassay for detection of staphylococcal toxins in complex media of virtually any volume, with increasing sensitivity at higher sample volume. The signal is read out from the entire volume of a nontransparent 3D fiber structure employed as a solid phase, which provides large reaction surface, quick reagent mixingi as well as antigen imnsunofiltration directly in the course of the assay. The method has demonstrated near-linear dose-response curves withm a wide range of ~3 decades, while detection of staphylococcal enterotoxin A (SEA) and toxic shock syndrome toxin (TSST) in neat milk without sample preparation. The limits of detection (LOD) as low as 4 and 10 pg/mL for TSST and SEA, respectively, were obtained in 2-h format using 30-mL samples. The second, 25-mm format, showed the LOD of 0.1 and 0.3 nglmL for the same toxins in a 150 j~L sample. The developed immunoassay can be applied in food safety control, in vitro diagnostics, and veterinary for a variety of research from express tests in the field to highly sensitive laboratory tests. [ABSTRACT FROM AUTHOR]

Nanoparticle-based technologies are rapidly expanding into many areas of biomedicine and molecular science. The unique ability of magnetic nanoparticles to respond to the magnetic field makes them especially attractive for a number of in vivo applications including magnetofection. The magnetofection principle consists of the accumulation and retention of magnetic nanoparticles carrying nucleic acids in the area of magnetic field application. The method is highly promising as a clinically efficient tool for gene delivery in vivo. However, the data on in vivo magnetofection are often only descriptive or poorly studied, insufficiently systematized, and sometimes even contradictory. Therefore, the aim of the review was to systematize and analyze the data that influence the in vivo magnetofection processes after the systemic injection of magnetic nanostructures. The main emphasis is placed on the structure and coating of the nanomagnetic vectors. The present problems and future trends of the method development are also considered. [ABSTRACT FROM AUTHOR]

Magnetic nanoparticles have been widely used in nanobiomedicine for diagnostics and the treatment of diseases, and as carriers for various drugs. The unique magnetic properties of "magnetic" drugs allow their delivery in a targeted tumor or tissue upon application of a magnetic field. The approach of combining magnetic drug targeting and gene delivery is called magnetofection, and it is very promising. This method is simple and efficient for the delivery of genetic material to cells using magnetic nanoparticles controlled by an external magnetic field. However, magnetofection in vivo has been studied insufficiently both for local and systemic routes of magnetic vector injection, and the relevant data available in the literature are often merely descriptive and contradictory. In this review, we collected and systematized the data on the efficiency of the local injections of magnetic nanoparticles that carry genetic information upon application of external magnetic fields. We also investigated the efficiency of magnetofection in vivo, depending on the structure and coverage of magnetic vectors. The perspectives of the development of the method were also considered. [ABSTRACT FROM AUTHOR]

Smart nanomaterials, contrast nanoparticles and drug nanocarriers of advanced targeting architecture were designed for various biomedical applications. Most of such agents demonstrate poor pharmacokinetics in vivo due to rapid elimination from the bloodstream by cells of the mononuclear phagocyte system (MPS). One of the promising methods to prolong blood circulation of the nanoparticles without their modification is MPS blockade. The method temporarily decreases macrophage endocytosis in response to uptake of a low-toxic non-functional material. The effect of different factors on the efficiency of macrophage blockade in vivo induced by solid nanomaterials has been studied here. Those include: blocker nanoparticle size, ζ-potential, surface coating, dose, mice strain, presence of tumor or inflammation. We found that the blocker particle coating type had the strongest effect on MPS blockade efficiency, which allowed to prolong functional particle blood circulation half-life 18 times. The mechanisms capable of regulation of the MPS blockade have been demonstrated, which can promote application of this phenomenon in medicine for improving delivery of diagnostic and therapeutic nanomaterials. Image 1 [ABSTRACT FROM AUTHOR]

Blood circulation is the key parameter that determines the in vivo efficiency of nanoagents. Despite clinical success of the stealth liposomal agents with their inert and shielded surfaces, a great number of non-stealth nanomaterials is being developed due to their potential of enhanced functionality. By harnessing surface phenomena, such agents can offer advanced control over drug release through intricately designed nanopores, catalysis-propelled motion, computer-like analysis of several disease markers for precise target identification, etc. However, investigation of pharmacokinetic behavior of these agents becomes a great challenge due to ultra-short circulation (usually around several minutes) and impossibility to use the invasive blood-sampling techniques. Accordingly, the data on circulation of such agents has been scarce and irregular. Here, we demonstrate high-throughput capabilities of the developed magnetic particle quantification technique for nanoparticle circulation measurements and present a comprehensive investigation of factors that affect blood circulation of the non-stealth nanoparticles. Namely, we studied the following 9 factors: particle size, zeta-potential, coating, injection dose, repetitive administration, induction of anesthesia, mice strain, absence/presence of tumors, tumor size. Our fundamental findings demonstrate potential ways to extend the half-life of the agents in blood thereby giving them a better chance of achieving their goal in the organism. The study will be valuable for design of the next generation nanomaterials with advanced biomedical functionality. With the real-time magnetic particle quantification technique, we performed investigation of factors that influence blood circulation of non-stealth nanoparticles: from nanoagent properties to aspects of tumor growth. Unlabelled Image [ABSTRACT FROM AUTHOR]

Graphical abstract Abstract Exploration of novel types of iron oxide nanoparticles as well as novel versatile ways to prepare them in a controlled manner keeping in mind necessity of narrow size distributions and high colloidal and chemical stability is an important task for modern nanochemistry. Most of the procedures for preparation of nanocrystalline iron oxides require drastic conditions and complex mixtures of reagents, therefore there is a high demand for methods of synthesis of such nanoparticles (NPs) in mild conditions. In this study, we discovered a new way to prepare crystalline goethite-like hydrous ferric oxide (HFO) NPs by fast and simple precipitation procedure in aqueous media and probed modification strategies aimed at the development of modified HFO nanoparticles for biomedical applications, including express-diagnostics and specific cell targeting. [ABSTRACT FROM AUTHOR]

Graphical abstract Abstract Three types of polymer-coated glass slides were prepared by acylation of their γ–aminopropyl derivatives with succinic anhydride or poly(p -nitrophenyl acylate) (PNPA) at various reaction times. The variations in duration of chemical adsorption of PNPA allowed control over nano-corrugation of the reactive surfaces employed for biosensing. The visually smooth and optically clear polymer-coated slides were used for immobilization of a conjugate of bovine serum albumin with chloramphenicol (CAP) and label-free measurement of concentration of this antibiotic. The sensor chips prepared in this way provided strong biosensing response due to fast biomolecular interactions and efficient competition of the target analyte during indirect immunoassay. The kinetics of formation and dissociation of immune complexes "conjugate-antibody" on the sensor chip surface was registered by the label-free methods of spectral-correlation and spectral-phase interferometry. The average thickness of immune complex adsorption layers was in the range of 5–12 nm. The highest value of kinetic association constant (k on = 5 × 104 M−1s−1) and the lowest equilibrium dissociation constant (K diss = 2 × 10-8 M−1) were achieved with the chips originated in the PNPA-coated slides, which were produced under short (2 min) polymer chemisorption and exhibited nano-corrugation almost unchanged with respect to the parent glass, according to the atomic-force microscopy. The detection limit for CAP detection was 80 pg/mL, and the dynamic range was 5 orders. The proposed methods for fabrication of sensor chips with nano-corrugated surface, the results of their characterization, as well as the developed assay can be employed for high-throughput recording of multiple molecular interactions by spectral interferometry and in a wide range of biosensing platforms for registration of biologically active compounds in medicine, veterinary, food control and ecological monitoring. [ABSTRACT FROM AUTHOR]

Highlights • Method of magnetite particles stabilization and functionalization was developed. • Barnase ∗ Barstar interface makes the method universal in terms of self-assembly. • Fusion protein of C-terminal part of Mms6 and Barstar stabilized particles in PBS. • Targeted delivery of modified particles to HER2/neu+ cancer cells was realized. Abstract Surface modification of nanoparticles with various biologically active molecules makes it possible to realize the huge biomedical potential of various nanoobjects. Functionally active particles are usually obtained through stabilization by polymers and subsequent chemical conjugation with biomolecules, such as, antibodies or aptamers. This entails a number of problems, such as non-oriented conjugation, low coupling yield and the inability to easily vary the attached components on demand. To solve this problem, we developed a novel method of magnetite nanoparticles stabilization with simultaneous modification by functionally active protein – Barstar, for subsequent self-assembly of the necessary components with these particles through the interaction of high-affinity protein pair Barnase ∗ Barstar (K aff = 1014 M−1). Namely, we developed a biocompatible Bs-C-Mms6 fusion protein containing the C-terminal part of the Mms6 (magnetite-binding protein of magnetotactic bacteria) and Barstar (an inhibitor of bacterial ribonuclease Barnase). We obtained stable in PBS magnetite nanoparticles modified with Bs-C-Mms6. These particles can be used for self-assembly with any type of Barnase-containing molecules. To demonstrate the effectiveness of this approach for the development of targeted nanoparticles, we performed a self-assembly of these particles with a fusion protein of Barnase and DARPin9.29, namely DARPin9.29-Bn. DARPin9.29 recognizes the extracellular domain of clinically important HER2/neu oncomarker. We have shown that such particles selectively bind this oncomarker and can be used to detect HER2/neu-positive cancer cells for diagnostic purposes. [ABSTRACT FROM AUTHOR]

Abstract An ultrasensitive lateral-flow assay is developed for rapid quantitative detection of small molecules on-site. The conceptual novelty, which transfers lateral-flow assays to the category of highly sensitive quantitative systems, is due to employment of a bifunctional ligand combined with volumetric registration of magnetic nanolabels. The ligand provides extremely high affinity for trapping the nanolabels and, simultaneously, efficiently competes with the analyzed molecules for the limited quantity of antigen-binding sites on the nanolabels. The developed assay has been demonstrated as the first express method for measuring in human serum of free thyroxine (fT4). The limit of detection is 20 fМ or 16 fg/ml at the assay time <30 min with the dynamic range of 3 orders. Besides, we present the results of first characterization of kinetic parameters of interaction between free thyroxine and monoclonal antibody, as well as of competitive relationship between fT4 and fT4-biotin. The proposed universal platform can be used for ultrasensitive detection of small molecules in human in vitro diagnostics, veterinary, biosafety and counter-terrorism, food quality control, environmental monitoring, etc., as well as for search of new, previously undetectable, diagnostic markers in medicine. Graphical abstract Image 1 Highlights • The first express method for highly sensitive quantitative detection of free thyroxine. • On-site lateral-flow assay, which is 100 times more sensitive than the traditional ELISA. • The first immunochromatographic assay that uses a bifunctional ligand. • The first characterization of competitive relationship between fT4 and bifunctional ligand. • The convenient and affordable method for search of new disease markers. [ABSTRACT FROM AUTHOR]

Smart materials that can autonomously implement sensing, analyze information and execute a pre-programmed action are extremely attractive for advanced biosensing and theranostics. Recently, novel smart “biocomputing” nanoagents have emerged, which exhibit remarkable ability to adapt their behavior to changes in microenvironment. These agents use Boolean logic for simultaneous processing of complex combinations of several biochemical stimuli. One of the major challenges in design of such nanoagents is development of biocomputing sensing interfaces and their tuning for proper logic-gating characteristics. Here, we present a versatile method for precise, rapid and controllable design of smart logic-gating layers by means of a label-free optical instrument. We demonstrate real-time monitoring and control of every stage of the logic gating at a molecular level with detailed study of kinetic parameters of interactions between all participating components. Such label-free investigation is exceptionally valuable for fine tuning of the layers’ components for logic-gating biosensing. We show that direct translation onto gold nanoparticles of the optimized smart biolayers, which implement basic YES and NOT logic gates, yields intelligent labels for lateral flow immunoassay. The proposed method is promising for development of advanced biosensors and theranostic agents based on smart biolayers, which can “make decisions” as a result of complex analysis of various biochemical cues. [ABSTRACT FROM AUTHOR]

Enhancement of sensitivity and reproducibility of measuring the concentration of small molecules (haptens) is crucially important for a wide variety of applications. The immunochemical detection of haptens by different biosensors is commonly realized by competitive assays that strongly depend on sorption capacity of the sensor chip and accessibility of antigen determinants in a conjugate immobilized on its surface. Here, we propose a method for significant improvement of limits of label-free detection of haptens due to enhancement of reproducibility of the competitive assay by taking into account the specific sorption capacity within the sensing area. The method is validated by mycotoxin detection in complex food matrices such as white wine. The limits of detection of 0.25 ng/mL, 0.48 ng/mL and 1 ng/mL are achieved for ochratoxin A, zearalenone, and aflatoxin B1, respectively, using compact label-free interferometric biosensors that employ microscope cover slips as affordable single-used sensor chips. The proposed method of drastic improvement of reproducibility of competitive assays can be used by any other label-free biosensors. This feature combined with the well-known advantages of label-free biosensors could make them attractive for precise, rapid, sensitive and multiplex detection of small molecules in food analysis, medicine, biosafety and environmental monitoring, etc. [ABSTRACT FROM AUTHOR]

Background Theranostics, a fusion of two key parts of modern medicine - diagnostics and therapy of the organism's disorders, promises to bring the efficacy of medical treatment to a fundamentally new level and to become the basis of personalized medicine. Extrapolating today's progress in the field of smart materials to the long-run prospect, we can imagine future intelligent agents capable of performing complex analysis of different physiological factors inside the living organism and implementing a built-in program thereby triggering a series of therapeutic actions. These agents, by analogy with their macroscopic counterparts, can be called nanorobots. It is quite obscure what these devices are going to look like but they will be more or less based on today's achievements in nanobiotechnology. Scope of Review The present Review is an attempt to systematize highly diverse nanomaterials, which may potentially serve as modules for theranostic nanorobotics, e.g., nanomotors, sensing units, and payload carriers. Major Conclusions Biocomputing-based sensing, externally actuated or chemically “fueled” autonomous movement, swarm inter-agent communication behavior are just a few inspiring examples that nanobiotechnology can offer today for construction of truly intelligent drug delivery systems. General Significance The progress of smart nanomaterials toward fully autonomous drug delivery nanorobots is an exciting prospect for disease treatment. Synergistic combination of the available approaches and their further development may produce intelligent drugs of unmatched functionality. [ABSTRACT FROM AUTHOR]

Biomolecule-driven assembly of nanoparticles is a powerful and convenient approach for development of advanced nanosensors and theranostic agents with diverse “on-demand” composition and functionality. While a lot of research is being devoted to fabrication of such agents, the development of non-invasive analytical tools to monitor self-assembly/disassembly processes in real-time substantially lags behind. Here, we demonstrate the capabilities of localized surface plasmon resonance (SPR) phenomenon to study non-covalent interactions not just between plasmonic particles, but between gold nanoparticles (AuNP) and non-plasmonic ones. We show its potential to investigate assembly and performance of a novel type of advanced smart materials, namely, biocomputing agents. These agents, self-assembled from nanoparticles via biomolecular interfaces such as proteins, DNA, etc., can analyze presence of biomolecular inputs according to Boolean logic and undergo the input-induced disassembly in order to implement the proper output action. Using UV–Vis spectroscopy to monitor the assembly/disassembly processes of the basic YES-gate structure that consists of a polymer core particle with a multitude of associated gold nanoparticles, we found that the structure transformations are well-characterized by pronounced difference in SPR spectral band position (shifting up to 50 nm). This SPR shift correlates remarkably well with biochemical estimation of the assembly/disassembly extent, and can provide valuable real-time kinetic analysis. We believe that the obtained data can be easily extended to other non-plasmonic nanoparticle systems having similar chemical and colloidal properties. SPR method can become a valuable addition to analytical toolbox for characterization of self-assembled smart nanosystems used in biosensing, imaging, controlled release and other applications. [ABSTRACT FROM AUTHOR]

We present a multiplex quantitative lateral flow (LF) assay for simultaneous on-site detection of botulinum neurotoxin (BoNT) types A, B, and E in complex matrixes, which is innovative by virtually no sacrifice in performance while transition from the single-plex assays and by characteristics on the level of laboratory quantitative methods. The novel approach to easy multiplexing is realized via joining an on-demand set of single-plex LF strips, which employ magnetic nanolabels, into a miniature cylinder cartridge that mimics LF strip during all assay stages. The cartridge is read out by an original portable multichannel reader based on the magnetic particle quantification technique. The developed reader offers the unmatched 60 zmol detection limit and 7-order linear dynamic range for volumetric registration of magnetic labels inside a cartridge of several millimeters in diameter regardless of its optical transparency. Each of the test strips, developed here as building blocks for the multiplex assay, can be used "as is" for autonomous quantitative single-plex detection with the same measuring setup, exhibiting the limits of detection (LOD) of 0.22, 0.11, and 0.32 ng/mL for BoNT-A, -B, and -E, respectively. The proposed multiplex assay has demonstrated the remarkably similar LOD values of 0.20, 0.12, 0.35 ng/mL under the same conditions. The multiplex assay performance was successfully validated by BoNT detection in milk and apple and orange juices. The developed methods can be extended to other proteins and used for rapid multianalyte tests for point-of-care in intro diagnostics, food analysis, biosafety and environmental monitoring, forensics, and security, etc. [ABSTRACT FROM AUTHOR]

A 3-channel biosensor based on spectral correlation interferometry (SCI) has been adapted for direct optical detection of antigens by measuring changes in thickness of a biolayer on functionalized glass slips employed as affordable single-use sensor chips. The instrument is insensitive to the bulk refractive index of a solution under test and provides signals in metrological units (pm or nm). Using real-time monitoring with the SCI, protocols for fabrication of sensor chips with different functional (epoxylated, carboxylated, and biotinylated) surfaces for antibody immobilization have been developed and optimized to minimize chip-to-chip variations and achieve better limit of detection (LOD), shorter assay time, and longer shelf life. The optimized coupling surfaces have been compared for detection of human serum albumin (HSA) used as a model agent of medical significance. The dynamic ranges for measuring the HSA concentration were 0.07–20, 0.12–30, and 0.25–10 μg/ml, and the assay durations were less than 20, 15, and 30 min for the epoxylated, carboxylated, and biotinylated chips, respectively. The advantages of each type of sensor chip have been shown, namely, the carboxylated chips feature the shortest assay time, the epoxylated ones demonstrate the best LOD, and the biotinylated chips exhibit the longest shelf life in an unprotected environment. The developed protocols of antibody immobilization can be used in different biosensors and assay techniques including those based on fluorescent, magnetic or plasmonic labels, etc. The SCI is well compatible with various partially transparent layers used in biosensing and with microarrays for multi-analyte detection. [ABSTRACT FROM AUTHOR]

A device for the generation of nanoaerosols from biological substances was designed and the resulting nanoaerosols were characterized. The function of the generator is based on the electrospray-neutralization process in which nanoclusters, obtained by electrohydrodynamic atomization, are neutralized by counter-ions generated by the electrospraying of ethanol at the opposite potential. The device is capable of generating nanoaerosol from suspensions of nanoparticles and solutions of non-volatile organic substances (e.g., antibiotics), proteins, and other biomolecules. This device produces nanoaerosols (10–300nm in size) with a concentration up to 2×107 cm−3. A quartz crystal microbalance based device was designed to monitor the performance of the generator. Using magnetic Fe3O4 nanoparticles as reporters, it has been demonstrated that approximately 0.4µg of nanoaerosol was deposited into a mouse lung in 1h in a nose-only exposure system. Thus, we present an approach to both generate nanoaerosol from biological and therapeutic substances and to estimate the doses delivered to mice exposed to these nanoaersolized substances. [ABSTRACT FROM AUTHOR]

[Display omitted] The rapid elimination of systemically administered drug nanocarriers by the mononuclear phagocyte system (MPS) compromises nanomedicine delivery efficacy. To mitigate this problem, an approach to block the MPS has been introduced and implemented by intravenous pre-administering blocker nanoparticles. The required large doses of blocker nanoparticles appeared to burden the MPS, raising toxicity concerns. To alleviate the toxicity issues in MPS blockade, we propose an intrinsically biocompatible blocker, ferrihydrite – a metabolite ubiquitous in a biological organism. Ferrihydrite particles were synthesized to mimic endogenous ferritin-bound iron. Ferrihydrite surface coating with carboxymethyl-dextran was found to improve MPS blockade dramatically with a 9-fold prolongation of magnetic nanoparticle circulation in the bloodstream and a 24-fold increase in the tumor targeted delivery. The administration of high doses of ferrihydrite caused low toxicity with a rapid recovery of toxicological parameters after 3 days. We believe that ferrihydrite particles coated with carboxymethyl-dextran represent superior blocking biomaterial with enviable biocompatibility. [ABSTRACT FROM AUTHOR]

Superparamagnetic particles of Fe3O4 in ferrofluid were injected transcranially in the ventricle of the rat brain. At three months after the injection the rat was sacrificed and the brain was investigated by Mössbauer spectroscopy and histological Perls Prussian blue method. Histological examination demonstrated increased concentration of blue areas in parenchyma and on the dura mater of the brain of experimental rat in comparison with the brain of control rat, indicating appearance in the brain of some undegradable compound of iron. Mössbauer spectra showed the presence in the ferrofluid of both Fe3O4 nanoparticles and an additional chemical compound which contains ferric iron in the high-spin state. Comparison of the Mössbauer spectra of the ferrofluid, of the brain before injection and of the brain three months after injection shows that the Fe3O4 nanoparticles were removed from the brain in three months. At the same time this additional chemical compound does not decompose and remains in the brain intact. [ABSTRACT FROM AUTHOR]

The development of non-invasive photothermal therapy (PTT) methods utilizing nanoparticles as sensitizers is one of the most promising directions in modern oncology. Nanoparticles loaded with photothermal dyes are capable of delivering a sufficient amount of a therapeutic substance and releasing it with the desired kinetics in vivo. However, the effectiveness of oncotherapy methods, including PTT, is often limited due to poor penetration of sensitizers into the tumor, especially into solid tumors of epithelial origin characterized by tight cellular junctions. In this work, we synthesized 200 nm nanoparticles from the biocompatible copolymer of lactic and glycolic acid, PLGA, loaded with magnesium phthalocyanine, PLGA/Pht-Mg. The PLGA/Pht-Mg particles under the irradiation with NIR light (808 nm), heat the surrounding solution by 40 °C. The effectiveness of using such particles for cancer cells elimination was demonstrated in 2D culture in vitro and in our original 3D model with multicellular spheroids possessing tight cell contacts. It was shown that the mean inhibitory concentration of such nanoparticles upon light irradiation for 15 min worsens by more than an order of magnitude: IC50 increases from 3 µg/mL for 2D culture vs. 117 µg/mL for 3D culture. However, when using the JO-4 intercellular junction opener protein, which causes a short epithelial–mesenchymal transition and transiently opens intercellular junctions in epithelial cells, the efficiency of nanoparticles in 3D culture was comparable or even outperforming that for 2D (IC50 = 1.9 µg/mL with JO-4). Synergy in the co-administration of PTT nanosensitizers and JO-4 protein was found to retain in vivo using orthotopic tumors of BALB/c mice: we demonstrated that the efficiency in the delivery of such nanoparticles to the tumor is 2.5 times increased when PLGA/Pht-Mg nanoparticles are administered together with JO-4. Thus the targeting the tumor cell junctions can significantly increase the performance of PTT nanosensitizers. [ABSTRACT FROM AUTHOR]

This review is dedicated to a comprehensive description of the latest achievements in the chemical functionalization routes and applications of carbon nanomaterials (CNMs), such as carbon nanotubes, graphene, and graphene nanoribbons. The review starts from the description of noncovalent and covalent exohedral modification approaches, as well as an endohedral functionalization method. After that, the methods to improve the functionalities of CNMs are highlighted. These methods include the functionalization for improving the hydrophilicity, biocompatibility, blood circulation time and tumor accumulation, and the cellular uptake and selectivity. The main part of this review includes the description of the applications of functionalized CNMs in bioimaging, drug delivery, and biosensors. Then, the toxicity studies of CNMs are highlighted. Finally, the further directions of the development of the field are presented. [ABSTRACT FROM AUTHOR]

Thyroid stimulating hormone (TSH) is the first-line marker for initial evaluation of the thyroid gland function. We present a lateral flow immunoassay based on superparamagnetic nanolabels for rapid (<25 min) quantitative determination of TSH at a point of care. The demonstrated limit of detection (LOD) of 0.017 μIU/mL in human serum is on the level of third-generation TSH laboratory tests. The wide linear dynamic range of more than 3 orders covers the whole range of clinically relevant TSH concentrations for confident quantitative diagnostics of the gland function from hyper- to hypothyroidism, and different states in-between. The attractive values of LOD and linear dynamic range are due to counting of the superparamagnetic nanolabels over the whole reaction volume by their non-linear magnetization at two frequencies of an alternating magnetic field and detecting the response at combinatorial frequencies. The developed cost-efficient and user-friendly immunoassay can be used for express in vitro diagnostics and long-term quantitative monitoring of thyroid dysfunctions, especially in distant regions, developing countries, and sparsely populated areas. Image 1 • Highly sensitive and rapid (25-min) assay for quantitative measurements of TSH in human serum. • Limit of detection – 0.017 μIU/mL suitable for diagnostics of hyperthyroidism. • Steep slope of calibration plot within 3-order linear dynamic range. • Accurate discrimination of virtually all TSH concentrations, pivotal for treatment strategy. • Easy-to-use, can be implemented on site and cost-efficient even for a single test. [ABSTRACT FROM AUTHOR]

The development of synthetic ways to fabricate nanosized materials with a well-defined shape, narrow-sized distribution, and high stability is of great importance to a rapidly developing area of nanotechnology. Here, we report an unusual reaction between amorphous two-line ferrihydrite and concentrated sulfuric or other mineral and organic acids. Instead of the expected dissolution, we observed the formation of new narrow-distributed brick-red nanoparticles (NPs) of hematite. Different acids produce similar nanoparticles according to scanning (SEM) and transmission electron microscopy (TEM), selected area electron diffraction (SAED), X-ray diffraction (XRD), infrared spectroscopy (FTIR), and energy-dispersive X-ray spectroscopy (EDX). The reaction demonstrates new possibilities for the synthesis of acid-resistant iron oxide nanoparticles and shows a novel pathway for the reaction of iron hydroxide with concentrated acids. The biomedical potential of the fabricated nanoparticles is demonstrated by the functionalization of the particles with polymers, fluorescent labels, and antibodies. Three different applications are demonstrated: i) specific targeting of the red blood cells, e.g., for red blood cell (RBC)-hitchhiking; ii) cancer cell targeting in vitro; iii) infrared ex vivo bioimaging. This novel synthesis route may be useful for the development of iron oxide materials for such specificity-demanding applications such as nanosensors, imaging, and therapy. [ABSTRACT FROM AUTHOR]

A rapid lateral flow immunoassay is presented that uses carboxyl-modified superparamagnetic nanoparticles as labels that can be quantified by highly sensitive multi-channel electronic readers. The approach is generic in that it is likely to be applicable to numerous small molecules. The method permits both single- and multiplex assays at a point-of-need without sample pretreatment. It is user-friendly and offers attractive characteristics demonstrated here for detection of morphine, fentanyl and methamphetamine in urine. The competitive immunoassay uses commercially available reagents that do not require special permissions. After migration of sample, the lateral flow test strips are subjected to an alternating magnetic field at two frequencies. The response from the nanolabels is readout at a combinatorial frequency from the entire volume of a porous immunochromatographic membrane by the magnetic particle quantification technique. Even trace concentrations can be quantified within ≤20 min with the limits of detection (LOD) of 0.20 ng·mL−1, 0.36 ng·mL−1 and 1.30 ng·mL−1 for morphine, fentanyl and methamphetamine, respectively. The second variant presented here features highly sensitive quantification of haptens (LOD for fentanyl - 0.05 ng·mL−1). This is due to high-affinity trapping of magnetic nanolabels in a universal streptavidin-based test strip, which can be also used for detection of virtually any other small molecule. The third variant is of the multiplexed type and intended for rapid and simultaneous detection of the drugs of abuse in human urine with LODs equal to 0.60 ng·mL−1 and 3.0 ng·mL−1 for morphine and methamphetamine, respectively. In addition to the low LODs, the RSDs did not exceed 7%, 9%, and 11% for methamphetamine, morphine and fentanyl, respectively. [ABSTRACT FROM AUTHOR]