Ashutosh Chilkoti

Point‑of‑care testing (POCT) demands diagnostic platforms that pair high analytical sensitivity with compact, low‑cost readout hardware, yet fluorescence‑based immunoassays face a fundamental tradeoff between detector cost and detection performance. Plasmonic nanocavities—such as metal–dielectric–metal nanopatch antennas—offer a powerful signal‑amplification strategy by dramatically enhancing fluorophore emission, enabling sensitive fluorescence detection with inexpensive optics. Here, we introduce a plasmonically enhanced fluorescence microarray immunoassay that integrates nanopatch antennas for high‑sensitivity, low‑cost quantification of key heart‑failure biomarkers, NT‑proBNP, NGAL, and galectin‑3, using a custom smartphone‑based detector. The nanopatch antennas yield up to 485‑fold fluorescence enhancement enabling quantitative and multiplexed detection with limits of detection of 0.055, 3.932, and 0.864 ng/mL, respectively. Analysis of NT‑proBNP–positive human samples shows strong correlation with clinically validated assays, demonstrating the potential of this plasmonically enhanced platform to deliver high‑performance heart‑failure biomarker monitoring at low cost in resource‑limited and point‑of‑care settings.

Peptide self-assembly and liquid–liquid phase separation (LLPS), often mediated by intrinsically disordered regions (IDRs), are natural mechanisms that translate protein molecular features into complex nano- and mesoscale architectures. Although the thermodynamics and kinetics of these processes are well understood, synthetic materials integrating both functionalities remain rare. Inspired by the conserved IDR–assembly domain (AD) architecture of amyloidogenic proteins, we hypothesized that modular recombinant constructs combining LLPS-capable IDRs with β-sheet-forming ADs could generate materials with tunable structural properties. To test this, we engineered a library of elastin-like polypeptides (ELPs) fused to amphiphilic anionic or cationic amyloidogenic peptides, enabling systematic investigation of how sequence parameters─including ELP length, AD charge, and hydrophilicity─and environmental conditions, including temperature, pH, and salt concentration, influence material behavior. Our results reveal links between molecular design and emergent multiscale structures, including micelles and vesicles embedded within coacervates. This work provides a framework for designing hybrid proteins coupling LLPS and self-assembly.

Tacrolimus is a cornerstone immunosuppressant used to prevent graft rejection in solid organ transplant recipients. While effective, tacrolimus therapy must be carefully monitored and managed due to the drug’s narrow therapeutic window and variable patient responses to dosing. Overdosing of tacrolimus can lead to opportunistic infections, nephrotoxicity, neurotoxicity, and cancer while under-dosing risks organ rejection. As a result, therapeutic drug monitoring (TDM) of tacrolimus is routinely performed. Current monitoring approaches face significant limitations, including slow (4–8 h or days) turnaround times, complex sample preparation requirements, and reliance on centralized infrastructure that delays critical dosing decisions. Furthermore, emerging evidence shows that the pharmacokinetic exposure parameter best associated with clinical outcomes is the area under the concentration–time curve (AUC). AUC-based monitoring requires sampling patients at multiple time points, which increases the logistical burdens to both patients and providers. Hence, there is an urgent need for a point-of-care test for tacrolimus that can be run at the patient bedside, in outpatient clinics, and eventually in patient homes. Here, we describe a novel point-of-care test, the TAC-D4 POCT, capable of measuring tacrolimus levels directly from whole blood samples without sample preparation steps or expensive analyzers with automated results available in 1 h. Our test helps overcome the current logistical barriers in the clinical management of patients receiving tacrolimus therapy ultimately leading to improved patient adherence, outcomes, and the ability to further personalize dosing regimens. Moreover, the Tac-D4 is highly modular and can be used to monitor clinically significantly biomarkers, such as interleukin-6 and matrix metalloproteinase 3, that complement tacrolimus TDM.

We previously reported the de novo design of three small (E. coli strain BL21(DE3) and up to three nonconsecutive disulfide bonds in the E. coli SHuffle T7 Express strain, without the need for refolding. The resulting SynIDP-DRP fusion proteins retain biological activity, confirming correct folding and disulfide-bond formation with minimal interference from the SynIDP tag, which obviates the need for tag removal. Collectively, these findings highlight the versatility and utility of SynIDPs as molecular tools to advance the production and development of protein therapeutics.

Metabolic dysfunction-associated steatohepatitis is a progressive liver disease characterized by inflammation and fibrosis, for which effective pharmacological treatments remain limited. Hormones that regulate metabolism, such as glucagon-like peptide-1 and fibroblast growth factor-21, have shown therapeutic promise through complementary metabolic and hepatoprotective actions. This study aims to develop and evaluate a single molecular therapy that engages both pathways to address multiple drivers of disease progression. We designed a unimolecular dual agonist by linking glucagon-like peptide-1 and fibroblast growth factor-21 using a thermally responsive elastin-like polypeptide linker to enable sustained drug exposure after subcutaneous administration. Receptor activity was assessed in cell-based assays. Therapeutic efficacy was evaluated in male C57Bl6/J mice with diet-induced advanced steatohepatitis and fibrosis using metabolic measurements, gene expression analyses, protein quantification, and histological assessment of liver tissue. Here we show that the dual agonist retains potent activity at both target receptors and forms a reversible subcutaneous depot that prolongs systemic exposure. Treatment improves body weight, liver mass, blood glucose, and cholesterol levels in mice with advanced liver disease. We further observe reduced liver inflammation and fibrosis, accompanied by decreased expression of inflammatory and fibrotic markers, reduced fat accumulation, and increased hepatocyte proliferation. These findings demonstrate that a single long-acting molecule engaging both metabolic hormone pathways improves metabolic and hepatic features of advanced steatohepatitis in a preclinical model. This work supports continued development of unimolecular multi-pathway therapies as a potential strategy for chronic liver disease.

Physically cross-linked hydrogels formed through noncovalent interactions offer reversible gelation, injectability, and shear-thinning behavior, making them ideal for biomedical applications. Temperature-responsive systems are particularly attractive, as they enable injection as a solution, and at physiological temperature for minimally invasive delivery. Poly(oligo(ethylene glycol) methacrylate) (POEGMA) is a promising nonimmunogenic polymer that combines PEG-like hydration and protein resistance with tunable thermal responsiveness that does not bind pre-existing anti-PEG antibodies that are ubiquitous and elicits a minimal antibody response against itself. Here, we show that diblock POEGMA copolymers─composed solely of short ethylene glycol side chains─spontaneously self-assemble into nanoparticles and form physically cross-linked hydrogels above a controllable transition temperature, without chemical modification. Using fluorescence recovery after photobleaching and video particle tracking nanorheology, we identify a thermally induced transition from dynamically arrested viscoelastic hydrogels to dynamically arrested liquid coacervates. These results establish a new class of single-component, nonimmunogenic POEGMA hydrogels with tunable phase behavior and mechanical properties, offering a versatile platform for localized drug delivery, sustained release, and regenerative medicine.

Biological organization has traditionally been viewed through the lens of distinct organelles and lock‐and‐key molecular interactions. The recent explosion of interest in phase separation has reshaped this view, revealing that cells also organize through membraneless organelles, also called biomolecular condensates, for spatial, temporal, and functional organization of its constituents. A key outcome of this shift has been the integration of polymer physics into cell biology, opening new avenues for understanding macromolecular behavior in living systems. This review traces the evolution of the field from foundational polymer physics, connects polymer properties to biological processes, and proposes a framework for interrogating cellular function through the lens of polymer science. While polymers are ubiquitous in daily life, their relevance to natural macromolecules and biological organization in cells is less apparent outside the discipline. Notably, current explorations at the biology–polymer interface echo much of the early pioneering work in polymer science. Accordingly, this review serves both as a primer on polymer concepts central to cellular processes and as a synthesis of recent advances and emerging tools for studying condensates in living cells.

Excessive alcohol consumption remains a major public health challenge with limited therapeutic options. Both glucagon-like peptide-1 (GLP-1) and fibroblast growth factor-21 (FGF21) independently regulate alcohol intake through complementary metabolic and reward pathways, but their combined potential has not been explored. Here, we report that a long-acting dual agonist, GLP1-ELP-FGF21 modulates behavioural, neurophysiological, and cognitive components of alcohol seeking in mice. A single GLP1-ELP-FGF21 dose reversibly reduces voluntary alcohol intake for at least 72 hours in male mice, has sustained effects in female mice, and markedly blunts nucleus accumbens dopamine transients aligned to the initiation and termination of lick bouts during alcohol consumption. To assess its effects on decision-making, we used a novel two-choice (alcohol versus food) decision task modelled with evidence-accumulation frameworks. Alcohol choice behaviour conformed to evidence accumulation decision models: Linear Ballistic Accumulator (LBM) and Racing diffusion models (RDM). Critically, GLP1-ELP-FGF21 selectively reduces choices for alcohol and slows the latent accumulation rate for alcohol options, without affecting food-directed choice or non-decision processes. Sensory-specific satiety devaluation confirms that reductions in reward value are explained by reductions in accumulation rates. Together, these results highlight GLP1-ELP-FGF21 as a therapeutic strategy for alcohol use disorder via modulation of central reward pathways and decision-making when confronted with alcohol reward