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Stability issues in membrane-free coacervates have been addressed with coating strategies, but these approaches often compromise the permeability of the coacervate. Here we report a facile approach to maintain both stability and permeability using tannic acid and then demonstrate the value of this approach in enzyme-triggered drug release. First, we develop size-tunable coacervates via self-assembly of heparin glycosaminoglycan with tyrosine and arginine-based peptides. A thrombin-recognition site within the peptide building block results in heparin release upon thrombin proteolysis. Notably, polyphenols are integrated within the nano-coacervates to improve stability in biofluids. Phenolic crosslinking at the liquid-liquid interface enables nano-coacervates to maintain exceptional structural integrity across various environments. We discover a pivotal polyphenol threshold for preserving enzymatic activity alongside enhanced stability. The disassembly rate of the nano-coacervates increases as a function of thrombin activity, thus preventing a coagulation cascade. This polyphenol-based approach not only improves stability but also opens the way for applications in biomedicine, protease sensing, and bio-responsive drug delivery.

Polymeric spherulites are typically formed by melt crystallization: spherulitic growth in solution is rare and requires complex polymers and dilute solutions. Here, we report the mild and unique formation of luminescent spherulites at room temperature via the simple molecule benzene-1,4-dithiol (BDT). Specifically, BDT polymerized into oligomers (PBDT) via disulfide bonds and assembled into uniform supramolecular nanoparticles in aqueous buffer; these nanoparticles were then dissolved back into PBDT in a good solvent (i.e., dimethylformamide) and underwent chain elongation to form spherulites (rPBDT) in 10 min. The spherulite geometry was modulated by changing the PBDT concentration and reaction time. Due to the step-growth polymerization and reorganization of PBDT, these spherulites not only exhibited robust structure but also showed broad clusterization-triggered emission. The biocompatibility and efficient cellular uptake of the spherulites further underscore their value as traceable drug carriers. This system provides a new pathway for designing versatile superstructures with value for hierarchical assembly of small molecules into a complicated biological system.

Colorimetric biosensors based on gold nanoparticle (AuNP) aggregation are often challenged by matrix interference in biofluids, poor specificity, and limited utility with clinical samples. Here, we propose a peptide-driven nanoscale disassembly approach, where AuNP aggregates induced by electrostatic attractions are dissociated in response to proteolytic cleavage. Initially, citrate-coated AuNPs were assembled via a short cationic peptide (RRK) and characterized by experiments and simulations. The dissociation peptides were then used to reversibly dissociate the AuNP aggregates as a function of target protease detection, i.e., main protease (Mpro), a biomarker for severe acute respiratory syndrome coronavirus 2. The dissociation propensity depends on peptide length, hydrophilicity, charge, and ligand architecture. Finally, our dissociation strategy provides a rapid and distinct optical signal through Mpro cleavage with a detection limit of 12.3 nM in saliva. Our dissociation peptide effectively dissociates plasmonic assemblies in diverse matrices including 100% human saliva, urine, plasma, and seawater, as well as other types of plasmonic nanoparticles such as silver. Our peptide-enabled dissociation platform provides a simple, matrix-insensitive, and versatile method for protease sensing.

Noroviruses are highly contagious and are one of the leading causes of acute gastroenteritis worldwide. Due to a lack of effective antiviral therapies, there is a need to diagnose and surveil norovirus infections to implement quarantine protocols and prevent large outbreaks. Currently, the gold standard of diagnosis uses reverse transcription polymerase chain reaction (RT-PCR), but PCR can have limited availability. Here, we propose a combination of a tunable peptide substrate and gold nanoparticles (AuNPs) to colorimetrically detect the Southampton norovirus 3C-like protease (SV3CP), a key protease in viral replication. Careful design of the substrate employs a zwitterionic peptide with opposite charged moieties on the C- and N- termini to induce a rapid color change visible to the naked eye; thus, this color change is indicative of SV3CP activity. This work expands on existing zwitterionic peptide strategies for protease detection by systematically evaluating the effects of lysine and arginine on nanoparticle charge screening. We also determine a limit of detection for SV3CP of 28.0 nM with comparable results in external breath condensate, urine, and fecal matter for 100 nM of SV3CP. The key advantage of this system is its simplicity and accessibility, thus making it an attractive tool for qualitative point-of-care diagnostics.

Electrostatic interactions are a key driving force that mediates colloidal assembly. The Schulze-Hardy rule states that nanoparticles have a higher tendency to coagulate in the presence of counterions with high charge valence. However, it is unclear how the Schulze-Hardy rule works when the simple electrolytes are replaced with more sophisticated charge carriers. Here, we designed cationic peptides of varying valencies and demonstrate that their charge screening behaviors on anionic gold nanoparticles (AuNPs) follow the six-power relationship in the Schulze-Hardy rule. This finding further inspires a simple yet effective strategy for measuring SARS-CoV-2 main protease (Mpro) via naked eyes. This work provides a unique avenue for fundamental NP disassembly based on the Schulze-Hardy rule and can help design versatile substrates for colorimetric sensing of other proteases.

Transmembrane protease serine 2 (TMPRSS2) is a plasma membrane protease that activates both spike protein of coronaviruses for cell entry and oncogenic signaling pathways for tumor progression. TMPRSS2 inhibition can reduce cancer invasion and metastasis and partially prevent the entry of SARS-CoV-2 into host cells. Thus, there is an urgent need for both TMPRSS2-selective imaging and precise screening of TMPRSS2 inhibitors. Here, we report a TMPRSS2-responsive surface-potential-tunable peptide-conjugated probe (EGTP) with aggregation-induced emission (AIE) features for TMPRSS2 selective imaging and accurate inhibitor screening. The amphiphilic EGTP was constructed with tunable surface potential and responsive efficiency with TMPRSS2 and its inhibitor. The rational construction of AIE luminogens (AIEgens) with modular peptides indicated that the cleavage of EGTP led to a gradual aggregation with bright fluorescence in high TMPRSS2-expressing cells. This strategy may have value for selective detection of cancer cells, SARS-CoV-2-target cells, and screening of protease inhibitors.

Aromatic interactions are commonly involved in the assembly of naturally occurring building blocks, and these interactions can be replicated in an artificial setting to produce functional materials. Here we describe a colorimetric biosensor using co-assembly experiments with plasmonic gold and surfactant-like peptides (SLPs) spanning a wide range of aromatic residues, polar stretches, and interfacial affinities. The SLPs programmed in DDD-(ZZ)x -FFPC self-assemble into higher-order structures in response to a protease and subsequently modulate the colloidal dispersity of gold leading to a colorimetric readout. Results show the strong aggregation propensity of the FFPC tail without polar DDD head. The SLPs were specific to the target protease, i.e., Mpro , a biomarker for SARS-CoV-2. This system is a simple and visual tool that senses Mpro in phosphate buffer, exhaled breath condensate, and saliva with detection limits of 15.7, 20.8, and 26.1 nM, respectively. These results may have value in designing other protease testing methods.

3D-bioprinted skin-mimicking phantoms with skin colors ranging across the Fitzpatrick scale are reported. These tools can help understand the impact of skin phototypes on biomedical optics. Synthetic melanin nanoparticles of different sizes (70-500 nm) and clusters are fabricated to mimic the optical behavior of melanosome. The absorption coefficient and reduced scattering coefficient of the phantoms are comparable to real human skin. Further the melanin content and distribution in the phantoms versus real human skins are validated via photoacoustic (PA) imaging. The PA signal of the phantom can be improved by: 1) increasing melanin size (3-450-fold), 2) increasing clustering (2-10.5-fold), and 3) increasing concentration (1.3-8-fold). Then, multiple biomedical optics tools (e.g., PA, fluorescence imaging, and photothermal therapy) are used to understand the impact of skin tone on these modalities. These well-defined 3D-bioprinted phantoms may have value in translating biomedical optics and reducing racial bias.

Existing tools to detect and visualize severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) suffer from low selectivity, poor cell permeability, and high cytotoxicity. Here we report a novel self-immolative fluorescent probe (MP590) for the highly selective and sensitive detection of the SARS-CoV-2 main protease (Mpro). This fluorescent probe was prepared by connecting a Mpro-cleavable peptide (N-acetyl-Abu-Tle-Leu-Gln) with a fluorophore (i.e., resorufin) via a self-immolative aromatic linker. Fluorescent titration results show that MP590 can detect Mpro with a limit of detection (LoD) of 35 nM and is selective over interferents such as hemoglobin, bovine serum albumin (BSA), thrombin, amylase, SARS-CoV-2 papain-like protease (PLpro), and trypsin. The cell imaging data indicate that this probe can report Mpro in HEK 293T cells transfected with a Mpro expression plasmid as well as in TMPRSS2-VeroE6 cells infected with SARS-CoV-2. Our results suggest that MP590 can both measure and monitor Mpro activity and quantitatively evaluate Mpro inhibition in infected cells, making it an important tool for diagnostic and therapeutic research on SARS-CoV-2.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a serious threat to human health and lacks an effective treatment. There is an urgent need for both real-time tracking and precise treatment of the SARS-CoV-2-infected cells to mitigate and ultimately prevent viral transmission. However, selective triggering and tracking of the therapeutic process in the infected cells remains challenging. Here, we report a main protease (Mpro)-responsive, mitochondrial-targeting, and modular-peptide-conjugated probe (PSGMR) for selective imaging and inhibition of SARS-CoV-2-infected cells via enzyme-instructed self-assembly and aggregation-induced emission (AIE) effect. The amphiphilic PSGMR was constructed with tunable structure and responsive efficiency and validated with recombinant proteins, cells transfected with Mpro plasmid or infected by SARS-CoV-2, and a Mpro inhibitor. By rational construction of AIE luminogen (AIEgen) with modular peptides and Mpro, we verified that the cleavage of PSGMR yielded gradual aggregation with bright fluorescence and enhanced cytotoxicity to induce mitochondrial interference of the infected cells. This strategy may have value for selective detection and treatment of SARS-CoV-2-infected cells.

Synchronization is important for photoacoustic (PA) tomography, but some fixed delays between the data acquisition (DAQ) and the light pulse are a common problem degrading imaging quality. Here, we present a simple yet versatile method named Scissors to help synchronize ultrasound open platforms (UOPs) for PA imaging. Scissors is a programed function that can cut or add a fixed delay to radio frequency (RF) data and, thus, synchronize it before reconstruction. Scissors applies the programmable metric of UOPs and has several advantages. It is compatible with many setups regardless of the synchronization methods, light sources, transducers, and delays. The synchronization is adjustable in steps reciprocal to the UOPs sampling rate (20-ns step with a 50-MHz sampling rate). Scissors works in real-time PA imaging, and no extra hardware is needed. We programed Scissors in Vantage UOP (Verasonics, Inc., Kirkland, WA, USA) and then imaged two 30- [Formula: see text] nichrome wires with a 20.2-MHz central frequency transducer. The PA image was severely distorted by an 828-ns delay; over 90% delay was caused by our Q -switch laser. The axial and lateral resolutions are 112 and [Formula: see text], respectively, after using Scissors. We imaged a human finger in vivo, and the imaging quality is tremendously improved after solving the 828-ns delay by using Scissors.

Heparin is a common anticoagulant, but heparin overdose is a common intensive care unit (ICU) medication error due to the narrow therapeutic window of heparin. Conventional methods to monitoring heparin suffer from long turnaround time, the need for skilled personnel, and low frequency of sampling. To overcome these issues, we describe here a fiber optic photoacoustic (PA) sensor for real-time heparin monitoring. The proposed sensor was validated with in vitro testing and in a simulated in vivo model using the following samples: (1) phosphate-buffered saline (PBS), (2) spiked human plasma, (3) spiked whole human blood, and (4) clinical samples from patients treated with heparin. Samples were validated by comparing the PA signal to the activated partial thromboplastin time (aPTT) as well as the activated clotting time (ACT). Importantly, the proposed sensor has a short turnaround time (3 min) and a limit of detection of 0.18 U/ml in whole human blood. The PA signal is linear with heparin dose and correlates with the aPTT value (Pearson's r = 0.99). The PA signal from 32 clinical samples collected from eight patients linearly correlated with ACT values (Pearson's r = 0.89, in vitro; Pearson's r = 0.93, simulated in vivo). The PA signal was also validated against the cumulative heparin dose (Pearson's r = 0.94, in vitro; Pearson's r = 0.96, simulated in vivo). This approach could have applications in both in vitro and real-time in vivo heparin monitoring.

Phenolics are ubiquitous in nature and have gained immense research attention because of their unique physiochemical properties and widespread industrial use. In recent decades, their accessibility, versatile reactivity, and relative biocompatibility have catalysed research in phenolic-enabled nanotechnology (PEN) particularly for biomedical applications which have been a major benefactor of this emergence, as largely demonstrated by polydopamine and polyphenols. Therefore, it is imperative to overveiw the fundamental mechanisms and synthetic strategies of PEN for state-of-the-art biomedical applications and provide a timely and comprehensive summary. In this review, we will focus on the principles and strategies involved in PEN and summarize the use of the PEN synthetic toolkit for particle engineering and the bottom-up synthesis of nanohybrid materials. Specifically, we will discuss the attractive forces between phenolics and complementary structural motifs in confined particle systems to synthesize high-quality products with controllable size, shape, composition, as well as surface chemistry and function. Additionally, phenolic's numerous applications in biosensing, bioimaging, and disease treatment will be highlighted. This review aims to provide guidelines for new scientists in the field and serve as an up-to-date compilation of what has been achieved in this area, while offering expert perspectives on PEN's use in translational research.

N95 decontamination protocols and KN95 respirators have been described as solutions to a lack of personal protective equipment. However, there are a few material science studies that characterize the charge distribution and physical changes accompanying disinfection treatments, particularly heating. Here, we report the filtration efficiency, dipole charge density, and fiber integrity of N95 and KN95 respirators before and after various decontamination methods. We found that the filter layers in N95 and KN95 respirators maintained their fiber integrity without any deformations during disinfection. The filter layers of N95 respirators were 8-fold thicker and had 2-fold higher dipole charge density than that of KN95 respirators. Emergency Use Authorization (EUA)-approved KN95 respirators showed filtration efficiencies as high as N95 respirators. Interestingly, although there was a significant drop in the dipole charge in both respirators during decontamination, there was no remarkable decrease in the filtration efficiencies due to mechanical filtration. Cotton and polyester face masks had a lower filtration efficiency and lower dipole charge. In conclusion, a loss of electrostatic charge does not directly correlate to the decreased performance of either respirator.

Personal protective equipment (PPE) including N95 respirators are critical for persons exposed to SARS-CoV-2. KN95 respirators and N95 decontamination protocols have been described as solutions to a lack of such PPE. However, there are a few materials science studies that characterize the charge distribution and physical changes accompanying disinfection treatments particularly heating. Here, we report the filtration efficiency, dipole charge density, and fiber integrity of pristine N95 and KN95 respirators before and after various decontamination methods. We found that the filter layer of N95 is 8-fold thicker than that of KN95, which explains its 10% higher filtration efficiency (97.03 %) versus KN95 (87.76 %) under pristines condition. After 60 minutes of 70 °C treatment, the filtration efficiency and dipole charge density of N95 became 97.16% and 12.48 µC/m2, while those of KN95 were 83.64% and 1.48 µC/m2 ; moreover, fit factor of N95 was 55 and that of KN95 was 2.7. In conclusion, the KN95 respirator is an inferior alternative of N95 respirator. In both systems, a loss of electrostatic charge does not directly correlate to a decrease in performance.