Avidin-biotin Interaction Research Articles

Electrochemical detection of poly(ADP-ribose) polymerase-1 with silver nanoparticles as signal labels by integrating the advantages of homogeneous reaction with surface-tethered detection

Poly(ADP-ribose)polymerase-1 (PARP1) could be activated by binding to nucleic acids with specific sequences, thus catalyzing the poly-ADP-ribosylation (PARylation) of target proteins including PARP1 itself. Most of the previously reported electrochemical methods for the determination of PARP1 were relied on the electrostatic interactions, which required the pre-immobilization of DNA on an electrode for the capture of PARP1. Herein, we reported an “immobilization−free” electrochemical strategy for the assays of PARP1 on the basic of avidin-biotin interaction. Once PARP1 was activated by binding with the specific double-stranded DNA (dsDNA) in a homogeneous solution, the biotinylated nicotinamide adenine dinucleotide (biotin-NAD+) was transferred onto PARP1, resulting in the formation of biotinylated PAR polymers. The resulting biotinylated PAR polymers were then captured by a neutravidin (NA)-modified electrode through avidin-biotin interactions. The rich biotin moieties in the PAR polymers allowed for the capture of NA-modified silver nanoparticles (NA-AgNPs) through the avidin-biotin interactions. The surface-tethered AgNPs produced a well-defined electrochemical signal due to the characteristic solid-state Ag/AgCl process. The “immobilization−free”, electrostatic interaction-independent electrochemical biosensor exhibited low background current, high sensitivity, and good stability. It has achieved the determination of PARP1 with a detection limit down to 0.7 mU. The biosensor was further applied to determine the inhibition efficiency of potential inhibitors with a satisfactory result. This method shows promising potential applications in PARP1-related clinical diagnosis and drug discovery.

Probing the interaction between biomolecules under sub-zero temperature conditions by electrophoresis in ice grain boundaries

BackgroundPsychrophiles can survive under cryogenic conditions because of various biomolecules. These molecules interact with cells, ice crystals, and lipid bilayers to enhance their functionality. Previous studies typically measured these interactions by thawing frozen samples and conducting biological assays at room temperature; however, studying these interactions under cryogenic conditions is crucial. This is because these biomolecules can function at lower temperatures. Therefore, a platform for measuring chemical interactions under sub-zero temperature conditions must be established. ResultsThe chemical interactions between biomolecules under sub-zero temperature conditions were evaluated within ice grain boundaries with a channel-like structure, which circumvents the need for thawing. An aqueous solution of sucrose was frozen within a microfluidic channel, facilitating the formation of freeze-concentrated solutions (FCSs) that functioned as size-tunable electrophoretic fields. Avidin proteins or single-stranded DNA (ssDNA) were introduced into the FCS in advance. Probe micro/nanospheres whose surfaces were modified with molecules complementary to the target analytes were introduced into the FCS. If the targets have functionalities under sub-zero temperature conditions, they interact with complementary molecules. The chemical interactions between the target molecules and nanospheres led to the aggregation of the particles. The size tunability of the diameter of the FCS channels enabled the recognition of aggregation levels, which is indicative of interaction reactivity. The avidin–biotin interaction and ssDNA hybridization served as models for chemical interactions, demonstrating interactivity under sub-zero temperature conditions. The results presented herein suggest the potential for in situ measurement of biochemical assays in the frozen state, elucidating the functionality of bio-related macromolecules at or slightly below 0 °C. SignificanceThis is the first methodology to evaluate chemical interactions under sub-zero temperature conditions without employing the freeze-and-thaw process. This method has the advantage of revealing the chemical interactions only at low temperatures. Therefore, it can be used to screen and evaluate the functionality of cryo-related biomolecules, including cold-shock and antifreeze proteins.

Smart Biointerfaces via Click Chemistry-Enabled Nanopatterning of Multiple Bioligands and DNA Force Sensors.

Nanoscale biomolecular placement is crucial for advancing cellular signaling, sensor technology, and molecular interaction studies. Despite this, current methods fall short in enabling large-area nanopatterning of multiple biomolecules while minimizing nonspecific interactions. Using bioorthogonal tags at a submicron scale, we introduce a novel hole-mask colloidal lithography method for arranging up to three distinct proteins, DNA, or peptides on large, fully passivated surfaces. The surfaces are compatible with single-molecule fluorescence microscopy and microplate formats, facilitating versatile applications in cellular and single-molecule assays. We utilize fully passivated and transparent substrates devoid of metals and nanotopographical features to ensure accurate patterning and minimize nonspecific interactions. Surface patterning is achieved using bioorthogonal TCO-tetrazine (inverse electron-demand Diels-Alder, IEDDA) ligation, DBCO-azide (strain-promoted azide-alkyne cycloaddition, SPAAC) click chemistry, and biotin-avidin interactions. These are arranged on surfaces passivated with dense poly(ethylene glycol) PEG brushes crafted through the selective and stepwise removal of sacrificial metallic and polymeric layers, enabling the directed attachment of biospecific tags with nanometric precision. In a proof-of-concept experiment, DNA tension gauge tether (TGT) force sensors, conjugated to cRGD (arginylglycylaspartic acid) in nanoclusters, measured fibroblast integrin tension. This novel application enables the quantification of forces in the piconewton range, which is restricted within the nanopatterned clusters. A second demonstration of the platform to study integrin and epidermal growth factor (EGF) proximal signaling reveals clear mechanotransduction and changes in the cellular morphology. The findings illustrate the platform's potential as a powerful tool for probing complex biochemical pathways involving several molecules arranged with nanometer precision and cellular interactions at the nanoscale.

All-stage targeted red blood cell membrane-coated docetaxel nanocrystals for glioma treatment

Challenges for glioma treatment with nanomedicines include physio-anatomical barriers (the blood-brain barrier and blood-brain tumor barrier), low drug loading capacity, and limited circulation time. Here, a red blood cell membrane-coated docetaxel drug nanocrystal (pV-RBCm-NC(DTX)), modified with pHA-VAP (pV) for all-stage targeting of glioma, was designed. The NC(DTX) core exhibited a high drug loading capacity but low in vivo stability, and the RBCm coating significantly enhanced the stability and prolonged in vivo circulation. Moreover, the Y-shaped targeting ligand pV was modified by a mild avidin-biotin interaction, which endowed RBCm-NC(DTX) with superior barrier-crossing ability and therapeutic efficacy. The integration of nanocrystal technology, cell membrane coating, and the avidin-biotin insertion method into this active targeting biomimetic formulation represents a promising drug delivery strategy for glioma.

Absorption-based colorimetric detection of nickel(II) ion by phase separation of thermoresponsive magnetic nanoparticles under microflow.

Therma-Max™ LSA Streptavidin is a thermoresponsive magnetic nanoparticle (TMNP). It can be introduced conveniently to molecular recognition groups by avidin-biotin interaction. In this study, we demonstrated the detection of nickel(II) ions by the magnetic separation of TMNP induced by their phase transition under microflow. The NTA-tagged TMNP solution mixed with a Ni2+ sample was introduced into a microchannel with a well structure. Moreover, the sample was heated to induce the thermally induced aggregation of TMNP. The Ni-capturing TMNP were trapped in the well by magnetic fields. The supernatant was removed from the outlet, and a dimethylglyoxime (DMG) solution was introduced into the device for colorimetric detection in the well. Because DMG has a higher stability constant with Ni2+, sensitive colorimetric detection of Ni2+ can be achieved in devices where the sample volume, e.g., optical pathlength, is short. To demonstrate the feasibility of the proposed method, a recovery test was conducted using a commercially available cosmetic sample. Therein, complete collection was achieved.

Biotin-functionalized nanoparticles: an overview of recent trends in cancer detection.

Electrochemical bio-sensing is a potent and efficient method for converting various biological recognition events into voltage, current, and impedance electrical signals. Biochemical sensors are now a common part of medical applications, such as detecting blood glucose levels, detecting food pathogens, and detecting specific cancers. As an exciting feature, bio-affinity couples, such as proteins with aptamers, ligands, paired nucleotides, and antibodies with antigens, are commonly used as bio-sensitive elements in electrochemical biosensors. Biotin-avidin interactions have been utilized for various purposes in recent years, such as targeting drugs, diagnosing clinically, labeling immunologically, biotechnology, biomedical engineering, and separating or purifying biomolecular compounds. The interaction between biotin and avidin is widely regarded as one of the most robust and reliable noncovalent interactions due to its high bi-affinity and ability to remain selective and accurate under various reaction conditions and bio-molecular attachments. More recently, there have been numerous attempts to develop electrochemical sensors to sense circulating cancer cells and the measurement of intracellular levels of protein thiols, formaldehyde, vitamin-targeted polymers, huwentoxin-I, anti-human antibodies, and a variety of tumor markers (including alpha-fetoprotein, epidermal growth factor receptor, prostate-specific Ag, carcinoembryonic Ag, cancer antigen 125, cancer antigen 15-3, etc.). Still, the non-specific binding of biotin to endogenous biotin-binding proteins present in biological samples can result in false-positive signals and hinder the accurate detection of cancer biomarkers. This review summarizes various categories of biotin-functional nanoparticles designed to detect such biomarkers and highlights some challenges in using them as diagnostic tools.

A Biotinylated Glycosylphosphatidylinositol (GPI) as the Universal Platform To Access GPI-Anchored Protein Analogues.

A glycosylphosphatidylinositol (GPI) derivative with biotin linked to its mannose III 6-O-position was prepared by a convergent strategy. This biotinylated GPI was demonstrated to bind avidinated proteins readily through biotin-avidin interaction and, therefore, can serve as a universal platform to access various biologically significant GPI-anchored protein analogues.

Mitochondrion-targeted carboxymethyl chitosan hybrid nanoparticles loaded with Coenzyme Q10 protect cardiac grafts against cold ischaemia‒reperfusion injury in heart transplantation

BackgroundHeart transplantation (HT) has been approved as an optimal therapeutic regimen for patients with terminal-stage cardiac failure. However, cold ischaemia‒reperfusion (I/R) injury remains an unavoidable and outstanding challenge, which is a major factor in early graft dysfunction and an obstacle to long-term survival in HT. Cold I/R injury induces cardiac graft injury by promoting mitochondrial dysfunction and augmenting free radical production and inflammatory responses. We therefore designed a mitochondrion-targeted nanocarrier loaded with Coenzyme Q10 (CoQ10) (CoQ10@TNPs) for treatment of cold I/R injury after cardiac graft in a murine heterotopic cardiac transplantation model.MethodsHybrid nanoparticles composed of CaCO3/CaP/biotinylated-carboxymethylchitosan (CaCO3/CaP/BCMC) were synthesized using the coprecipitation method, and the mitochondria-targeting tetrapeptide SS31 was incorporated onto the surface of the hybrid nanoparticles through biotin-avidin interactions. Transmission electron microscopy (TEM) and dynamic light scattering (DLS) analysis were used for characterisation. In vitro, the hypoxia-reoxygenation model of H9c2 cells was employed to replicate in vivo cold I/R injury and treated with CoQ10@TNPs. The impact of CoQ10@TNPs on H9c2 cell injury was assessed by analysis of oxidative damage and apoptosis. In vivo, donor hearts (DHs) were perfused with preservation solution containing CoQ10@TNPs and stored in vitro at 4 °C for 12 h. The DHs were heterotopically transplanted and analysed for graft function, oxidative damage, apoptosis, and inflammatory markers 1 day post-transplantation.ResultsCoQ10@TNPs were successfully synthesized and delivered CoQ10 to the mitochondria of the cold ischaemic myocardium. In vitro experiments demonstrated that CoQ10@TNPs was taken up by H9c2 cells at 4 °C and localized within the mitochondria, thus ameliorating oxidative stress damage and mitochondrial injury in cold I/R injury. In vivo experiments showed that CoQ10@TNPs accumulated in DH tissue at 4 °C, localized within the mitochondria during cold storage and improved cardiac graft function by attenuating mitochondrial oxidative injury and inflammation.ConclusionsCoQ10@TNPs can precisely deliver CoQ10 to the mitochondria of cold I/R-injured cardiomyocytes to effectively eliminate mitochondrial reactive oxygen species (mtROS), thus reducing oxidative injury and inflammatory reactions in cold I/R-injured graft tissues and finally improving heart graft function. Thus, CoQ10@TNPs offer an effective approach for safeguarding cardiac grafts against extended periods of cold ischaemia, emphasizing the therapeutic potential in mitigating cold I/R injury during HT. These findings present an opportunity to enhance existing results following HT and broaden the range of viable grafts for transplantation.

Biotinylated Quinone as a Chemiluminescence Sensor for Biotin-Avidin Interaction and Biotin Detection Application

Biotin, or vitamin B7, is essential for metabolic reactions. It must be obtained from external sources such as food and biotin/vitamin supplements because it is not biosynthesized by mammals. Therefore, there is a need to monitor its levels in supplements. However, biotin detection methods, which include chromatographic, immune, enzymatic, and microbial assays, are tedious, time-consuming, and expensive. Thus, we synthesized a product called biotin-naphthoquinone, which produces chemiluminescence upon its redox cycle reaction with dithiothreitol and luminol; then it was used as a chemiluminescence sensor for biotin–avidin interaction. When a quinone biotinylated compound binds avidin, the chemiluminescence decreases noticeably due to the proximity between quinone and avidin, and when free biotin is added in a competitive assay, the chemiluminescence returns. The chemiluminescence is regained as the free biotin displaces biotinylated quinone in its complex with avidin, freeing biotin-naphthoquinone. Many experiments, including the use of a biotin-free quinone, proved the competitive nature of the assay. The competitive assay method used in this study was linear in the range of 1.0–100 µM with a detection limit of 0.58 µM. The competitive chemiluminescence assay could detect biotin in vitamin B7 tablets with good recovery of 91.3 to 110% and respectable precision (RSD < 8.7%).

A novel visual biosensor for mercury detection based on a cleavable phosphorothioate-RNA probe and microfluidic bead trap

Mercury ion (Hg2+) detection plays a crucial role in safeguarding the environment and protecting human health. In this study, we developed a microfluidic biosensing platform that utilizes a phosphorothioate-RNA (PS-RNA) probe and gold-coated magnetic nanoparticles to quantify Hg2+ visually. The PS-RNA probe was linked to gold-coated magnetic nanoparticles via Au–S interaction and biotin–avidin interaction and then combined with polystyrene microspheres to form a self-assembled structure called a magnetic bead-AuNP-probe-polystyrene (MAPP) bead. We designed and fabricated a microfluidic chip that contained only a magnetic separator and a particle trap. The presence of Hg2+ cut the PS-RNA probe, leading to the separation of magnetic beads and polystyrene microspheres. The MAPP was removed by the magnetic separator, while the cut polystyrene microspheres flowed along the channel and accumulated in the particle trap, forming a long visible strip. The length of the polystyrene microsphere strip was proportional to the Hg2+ concentration. The microfluidic biosensing platform exhibited high sensitivity and specificity for Hg2+ detection, with a detection limit of 21.9 nM. By integrating the microfluidic magnetic manipulation system with a sensitive nucleic acid probe, we developed an automated and visual biosensor that is a valuable reference for the on-site visual detection of heavy metal ions.

Antibody-conjugated nanoparticles for target-specific drug delivery of chemotherapeutics.

Nanotechnology provides effective methods for precisely delivering chemotherapeutics to cancer cells, thereby improving efficacy and reducing off-target side effects. The targeted delivery of nanoscale chemotherapeutics is accomplished by two different approaches, namely the exploitation of leaky tumor vasculature (EPR effect) and the surface modification of nanoparticles (NPs) with various tumor-homing peptides, aptamers, oligonucleotides, and monoclonal antibodies (mAbs). Because of higher binding affinity and specificity, mAbs have received a lot of attention for the detection of selective cancer biomarkers and also for the treatment of various types of cancer. Antibody-conjugated nanoparticles (ACNPs) are an effective targeted therapy for the efficient delivery of chemotherapeutics specifically to the targeted cancer cells. ACNPs combine the benefits of NPs and mAbs to provide high drug loads at the tumor site with better selectivity and delivery efficiency. The mAbs on the NP surfaces recognize their specific receptors expressed on the target cells and release the chemotherapeutic agent in a controlled manner. Appropriately designed and synthesized ACNPs are essential to fully realize their therapeutic benefits. In blood stream, ACNPs instantly interact with biological molecules, and a protein corona is formed. Protein corona formation triggers an immune response and affects the targeting ability of the nanoformulation. In this review, we provide recent findings to highlight several antibody conjugation methods such as adsorption, covalent conjugation, and biotin-avidin interaction. This review also provides an overview of the many effects of the protein corona and the theranostic applications of ACNPs for the treatment of cancer.

(Invited) Microenvironment Responsiveness of Defect Photoluminescence of Locally Functionalized Single-Walled Carbon Nanotubes and Its Sensing Applications

Photoluminescence (PL) of single-walled carbon nanotubes (SWCNTs) in the near infrared (NIR) region is applicable to advanced optical applications such as bio-imaging and sensing. Local chemical functionalization (slight amount of chemical modification) of SWCNTs has been developed to introduce sp3 carbon defects in the sp2 carbon network of the tube walls for the luminescent defect formation.[1-3] The resultant locally functionalized SWCNTs (lf-SWCNTs) show defect PL (E 11* PL) with red-shifted wavelengths and increased quantum yields compared to original E 11 PL of pristine SWCNTs. For lf-SWCNTs, molecularly designed modifier molecules have enabled the defect PL wavelength modulation[4] and the sensing function creation.[5,6]Recently, we have reported the unique microenvironment responsiveness of defect PL of lf-SWCNTs; namely, the greater wavelength shifts of E 11* PL than E 11 PL by dielectric effects[7] and the PL shifting behavior modulation based on the chemical structure difference of the functionalized sites and the exciton localization effects[8] were observed. The microenvironment responsiveness of the defect PL was applied to the biological sensing application development.[9] To investigate protein detection/recognition functions of lf-SWCNTs, we introduced an avidin-biotin interaction with a selective and strong binding fashion at the functionalized sites of lf-SWCNTs. The lf-SWCNTs tethering biotin groups (lf-SWCNTs-b) were synthesized through diazonium chemistry, followed by its subsequent-modification. When neutravidin was mixed with a lf-SWCNTs-b solution, E 11* PL peak was red-shifted, indicating the higher polarity environment formation by the neutravidin adsorption on lf-SWCNTs-b. When avidin or streptavidin was used, wavelength shifting behaviors of E 11* PL of lf-SWCNTs-b were clearly changed depending on the used avidin derivatives. The results indicate the different polarity environment formation deriving from structural differences of each avidin derivative. Moreover, the detection signal enhancement was observed in a film device using lf-SWCNTs-b. Therefore, lf-SWCNTs have a high potential for development of advanced protein detection/recognition systems using NIR PL.[1] T. Shiraki et al. Acc. Chem. Res., 53, 1846 (2020). [2] S. Tretiak et al., Acc. Chem. Res., 53, 1791(2020) [3] Y. Wang et al. Nat. Rev. Chem., 3, 375 (2019). [4] T. Shiraki et al. ACS Nano, DOI: 10.1021/acsnano.2c09897 (2022). [5] T. Shiraki et al. Chem. Commun., 52, 12972 (2016). [6] S. Kruss et al, Angew. Chem. Int. Ed., 59, 17732 (2020). [7] T. Shiraki et al. Chem. Commun., 55, 3662 (2019). [8] T. Shiraki et al. J. Phys. Chem. C, 125, 12758 (2021). [9] T. Shiraki et al. Nanoscale, 14, 13090 (2022). Figure 1

Biotin-Avidin System-Based Delivery Enhances the Therapeutic Performance of MSC-Derived Exosomes.

Exosomes (EXs) shed by mesenchymal stem cells (MSCs) are potent therapeutic agents that promote wound healing and regeneration, but when used alone in vivo, their therapeutic potency is diminished by rapid clearance and bioactivity loss. Inspired by the biotin-avidin interaction, we developed a simple yet versatile method for the immobilization of MSC-derived EXs (MSC-EXs) into hydrogels and achieved sustained release for regenerative purposes. First, biotin-modified gelatin methacryloyl (Bio-GelMA) was fabricated by grafting NHS-PEG12-biotin onto the amino groups of GelMA. Biotin-modified MSC-EXs (Bio-EXs) were then synthesized using an in situ self-assembling biotinylation strategy, which provided sufficient binding sites for MSC-EX delivery with little effect on their cargo composition. Thereafter, Bio-EXs were immobilized in Bio-GelMA via streptavidin to generate Bio-GelMA@Bio-EX hydrogels. An in vitro analysis demonstrated that Bio-EXs could be taken up by macrophages and exerted immunomodulatory effects similar to those of MSC-EXs, and Bio-GelMA@Bio-EX hydrogels provided sustained release of MSC-EXs for 7 days. After subcutaneous transplantation, a more constant retention of MSC-EXs in Bio-GelMA@Bio-EX hydrogels was observed for up to 28 days. When placed in an artificial periodontal multitissue defect, the functionalized hydrogels exhibited an optimized therapeutic performance to regrow complex periodontal tissues, including acellular cementum, periodontal ligaments (PDLs), and alveolar bone. In this context, Bio-GelMA@Bio-EX hydrogels exerted a robust immunomodulatory effect that promoted macrophage polarization toward an M2 phenotype. Our findings demonstrate that MSC-EXs delivered with the aid of the biotin-avidin system exhibit robust macrophage-modulating and repair-promoting functions and suggest a universal approach for the development of MSC-EX-functionalized biomaterials for advanced therapies.

Sortase A transpeptidation produces seamless, unbranched biotinylated nanobodies for multivalent and multifunctional applications.

Exploitation of the biotin-streptavidin interaction for advanced protein engineering is used in many bio-nanotechnology applications. As such, researchers have used diverse techniques involving chemical and enzyme reactions to conjugate biotin to biomolecules of interest for subsequent docking onto streptavidin-associated molecules. Unfortunately, the biotin-streptavidin interaction is susceptible to steric hindrance and conformational malformation, leading to random orientations that ultimately impair the function of the displayed biomolecule. To minimize steric conflicts, we employ sortase A transpeptidation to produce quantitative, seamless, and unbranched nanobody-biotin conjugates for efficient display on streptavidin-associated nanoparticles. We further characterize the protein-nanoparticle complex and demonstrate its usefulness in optical microscopy and multivalent severe acute respiratory syndrome coronavirus (SARS-CoV-2) antigen interaction. The approach reported here provides a template for making novel multivalent and multifunctional protein complexes for avidity-inspired technologies.

"Clickable" Organic Electrochemical Transistors.

Interfacing the surface of an organic semiconductor with biological elements is a central quest when it comes to the development of efficient organic bioelectronic devices. Here, we present the first example of "clickable" organic electrochemical transistors (OECTs). The synthesis and characterization of an azide-derivatized EDOT monomer (azidomethyl-EDOT, EDOT-N3) are reported, as well as its deposition on Au-interdigitated electrodes through electropolymerization to yield PEDOT-N3-OECTs. The electropolymerization protocol allows for a straightforward and reliable tuning of the characteristics of the OECTs, yielding transistors with lower threshold voltages than PEDOT-based state-of-the-art devices and maximum transconductance voltage values close to 0 V, a key feature for the development of efficient organic bioelectronic devices. Subsequently, the azide moieties are employed to click alkyne-bearing molecules such as redox probes and biorecognition elements. The clicking of an alkyne-modified PEG4-biotin allows for the use of the avidin-biotin interactions to efficiently generate bioconstructs with proteins and enzymes. In addition, a dibenzocyclooctyne-modified thrombin-specific HD22 aptamer is clicked on the PEDOT-N3-OECTs, showing the application of the devices toward the development of organic transistors-based biosensors. Finally, the clicked OECTs preserve their electronic features after the different clicking procedures, demonstrating the stability and robustness of the fabricated transistors.

Detection of Salmonella Typhimurium with Gold Nanoparticles Using Quartz Crystal Microbalance Biosensor.

Demonstration of the Salmonella Typhimurium detection system was shown utilizing a quartz crystal microbalance (QCM) biosensor and signal enhancement by gold nanoparticles. In this study, a benchtop system of a QCM biosensor was utilized for the detection of Salmonella Typhimurium. It was designed with a peristaltic pump system to achieve immobilization of antibodies, detection of Salmonella, and the addition of gold nanoparticles to the sensor. As a series of biochemical solutions were introduced to the surface, the proposed system was able to track the changes in the resonant frequency which were proportional to the variations of mass on the sensor. For antibody immobilization, polyclonal antibodies were immobilized via self-assembled monolayers to detect Salmonella O-antigen. Subsequently, Salmonella Typhimurium was detected by antibodies and the average frequency before and after detecting Salmonella was compared. The highest frequency shifts were -26.91 Hz for 109 CFU/mL while the smallest frequency shift was -3.65 Hz corresponding to 103 CFU/mL. For the specificity tests, non-Salmonella samples such as E. coli, Listeria, and Staphylococcus resulted in low cross-reactivity. For signal amplification, biotinylated antibodies reacted to Salmonella followed by streptavidin-100 nm AuNPs through biotin-avidin interaction. The frequency shifts of 103 CFU/mL showed -28.04 Hz, and consequently improved the limit of detection.

Twelve Colors of Streptavidin-Fluorescent Proteins (SA-FPs): A Versatile Tool to Visualize Genetic Information in Single-Molecule DNA.

Streptavidin-fluorescent proteins (SA-FPs) are a versatile tool to visualize a broad range of biochemical applications on a fluorescence microscope. Although the avidin-biotin interaction is widely used, the use of SA-FPs has not been applied to single-molecule DNA visualization. Here, we constructed 12 bright SA-FPs for DNA staining or labeling reagents. To date, 810 FPs are available, many of which are brighter than organic dyes. In this study, 12 bright FPs were selected to construct SA-FP plasmids covering green to red colors. Their brightness ranges from 40 to 165 mM-1 cm-1. Moreover, SA-FP is brighter than FP itself because streptavidin forms a tetramer complex; thus, four FPs are in a single complex. In addition, FPs often form a dimer or a tetramer, resulting in multiple FPs in a single spot on a microscopic image. This feature is advantageous because multiple fluorescent β-barrels on a single biotin tag provide enough brightness to be easily visualized by epifluorescence microscopy. Using SA-FPs, we visualized DNA backbones, nickase-based optical mapping, and AT-frequency profiling. Finally, we demonstrated the combination of nickase-based optical mapping using SA-FP and AT-frequency profiling.

PH-Sensitive Twin Liposomes Containing Quercetin and Laccase for Tumor Therapy.

In this study, functional twin liposomes (TLs) were designed by linking avidin-anchored single liposomes and biotin-anchored single liposomes via avidin-biotin interactions. Here, we first punched a hole on the liposome surface using the liposome magnetoporation method to prepare functional single liposomes, which were used for safely encapsulating quercetin (QER, as a model prodrug) or laccase (LAC, as a bioactive enzyme) inside the liposomes without the use of organic solvents; the pores were then plugged by pH-sensitive glycol chitosan grafted with 3-diethylaminopropylamine (GDEAP) and avidin (or biotin). As a result, single liposomes with QER and biotin-GDEAP were efficiently coupled with other liposomes with LAC and avidin-GDEAP. We demonstrated that the TLs could accelerate QER and LAC release at acidic pH (6.8), improving the LAC-mediated oxidization of QER and significantly elevating tumor cell death, suggesting that this strategy can be used as an efficient method for the programmed action of prodrugs.

Radionuclide Delivery Strategies in Tumor Treatment: A Systematic Review.

The aim of this review was to assess recent progress in targeted radionuclide tumor therapy, focusing on the best delivery strategies. A literature search was conducted in PubMed, Web of Science, and Scopus using the terms “radionuclides”, “liposomes”, “avidin–biotin interaction”, “theranostic”, and “molecular docking”. The 10 year filter was applied, except for the avidin–biotin interaction. Data were retrieved from both preclinical and clinical settings. Three targeting strategies were considered: pretargeting, liposomes, and ligands. Pretargeting can be achieved by exploiting the avidin–biotin interaction. This strategy seems very promising, although it has been investigated mainly in resectable tumors. Radiolabeled liposomes have attracted new interest as probes to identify the most suitable patients for treatment with liposomal formulations of common chemotherapeutics. The use of ligands for the delivery of radiotherapeutics to a specific target is still the most appealing strategy for treating tumors. The most appropriate ligand can be identified by virtually simulating its interaction with the receptor. All strategies showed great potential for use in targeted radionuclide therapy, but they also have numerous drawbacks. The most promising option is probably the one based on the use of new ligands.

Live Macrophage-Delivered Doxorubicin-Loaded Liposomes Effectively Treat Triple-Negative Breast Cancer.

Triple-negative breast cancer is often aggressive and resistant to various cancer therapies, especially corresponding targeted drugs. It is shown that targeted delivery of chemotherapeutic drugs to tumor sites could enhance treatment outcome against triple-negative breast cancer. In this study, we exploited the active tumor-targeting capability of macrophages by loading doxorubicin-carrying liposomes on their surfaces via biotin-avidin interactions. Compared with conventional liposomes, this macrophage-liposome (MA-Lip) system further increased doxorubicin accumulation in tumor sites, penetrated deeper into tumor tissue, and enhanced antitumor immune response. As a result, the MA-Lip system significantly lengthened the survival rate of 4T1 cell-bearing mice with low toxicity. Besides, the MA-Lip system used highly biocompatible and widely approved materials, which ensured its long-term safety. This study provides a system for triple-negative breast cancer treatment and offers another macrophage-based strategy for tumor delivery.