Raman spectroscopy (RS) is a widely employed technique for analyzing emerging environmental pollutants, microplastics and nanoplastics (MNPs), detection of biomolecules, identification of cells and pathogens. Detecting, identifying, and quantifying these particles in environmental samples and living organisms poses significant challenges due to their minute size, irregular shapes, diverse polymer compositions, surface coatings, and large surface areas that readily attract chemical and microbial contaminants. Raman Spectroscopy is a reliable, specific, fast, more sensitive method for the characterization of small sized particles. Moreover, the handheld Raman device is easily deployable in the field. This review addresses the key analytical strengths and the challenges that limit precise characterization of MNPs and provide recommendations to improve data reliability, that further include strategies to mitigate common quality control issues, particularly the challenge of distinguishing between plastic particles present in the sample and those introduced through contamination during sampling, processing, or analysis. Recently, the use of artificial intelligence (AI) and machine learning has been incorporated with Raman spectroscopy to facilitate the detection of MNPs and provide automation.

In electrochemistry, ions and electrons move together to carry charge and drive chemical reactions, which has resulted in the innovation of many technologies, such as energy storage batteries, biosensors, and fuel cells. Organic electrochemical transistors (OECTs) combine the principles of electrochemistry and organic electronics. Here, the current flow is governed by the coordinated movement of ions and electrons. Typically, ions from an electrolyte enter an organic channel and modify its doping state, which results in changed conductivity and overall electrical response. OECTs can operate efficiently at low voltages in aqueous environments. It exhibits improved dielectric properties and can integrate naturally well with biological systems. This enables them to be well-suited candidates for real-time biosensing, electrophysiological recording, and in vivo monitoring. Apart from this, their simple fabrication process and robustness on wet or flexible substrates further enhance their compatibility with biological applications. Thus, they offer superior performance compared to organic field-effect transistors (OFETs), which basically operate on interfacial charge modulation. OECTs can also amplify weak biological signals and track environmental parameters. It makes them a versatile tool for point-of-care (POC) diagnostics, wearable health monitoring, pollutant detection, and precision agriculture. However, there are concerns regarding stability, scalability, and environmental impact that may hinder the widespread adoption of OECTs. This review reports the use of green electrochemistry in OECT design, with particular emphasis on environmentally friendly materials, biodegradable polymers, non-toxic electrolytes, low energy manufacturing techniques, and translation opportunities, etc., which is consistent with the vision of a circular economy and reduced e-waste.

Abstract Introduction Tumor hypoxia diminishes antitumor immunity by stabilizing HIF-1α, promoting M2-like macrophage polarization, and impairing cytotoxic T-cell activity. To reverse these effects, we developed hemoglobin-loaded biomimetic nanoparticles (Hb-BNPs) using a modified nanoprecipitation strategy. The nanoparticles encapsulate hemoglobin within a polycaprolactone matrix and are cloaked with RBC/A549 membranes to provide immune camouflage and sustained oxygen release. This study evaluates their ability to reoxygenate the tumor microenvironment and restores immune and therapeutic responses in non-small-cell lung carcinoma (NSCLC). Methods Hb-BNPs were synthesized via modified nanoprecipitation and membrane cloaking. Physicochemical features were assessed by DLS, TEM. Hypoxia was induced in A549 cells, followed by Hb-BNP treatment. qRT-PCR, paclitaxel IC50 assays, 3D spheroids, LunX CAR-T cytotoxicity, and KANK1-transfection studies evaluated immunologic and therapeutic responses (all n = 3, ANOVA/t). Results Under hypoxia, HIF1A, VEGF, BNIP3, ENO1, HK1, PGK1 were upregulated by 5—8-fold (p < 0.01). After Hb-BNP reoxygenation, these genes were downregulated by 4—6-fold (p < 0.001), and dissolved oxygen increased 5.3-fold (p < 0.001). Oxygen recovery improved the immune function as LunX CAR-T cytotoxicity increased 2.3-fold (p < 0.01), and KANK1-driven transgene expression increased 2.7-fold (p < 0.05). Co-treatment with paclitaxel reduced IC50 from 25,612 ng/mL to 781 ng/mL (p < 0.0001) and enhanced 3D spheroid core cell death 2.4-fold (p < 0.01). Hb-BNPs did not elevate IL-6, TNF-α, or IL-8 in THP-1 macrophages, confirming immune tolerance. Conclusion Hb-BNPs reoxygenate hypoxic tumors, suppress HIF-1α signaling, and restore cytotoxic immune function without inducing inflammation. By normalizing oxygen balance, these biomimetic nanoparticles enhance CAR-T activity, gene transfection, and chemotherapy response, offering a scalable, immune-tolerant platform to overcome hypoxia-driven resistance in solid tumors. Funding Source N/A Topic Categories Tumor Immunology: Checkpoints, Prevention, and Treatment (TIPT)

Hemoglobin A0 (HbA0) is a physiologically important tetrameric protein consisting of two alpha (alpha) and two beta (beta) subunits that resides inside red blood cells and carries oxygen to the cells. In this study, the interaction of HbA0 protein with anionic sodium dodecyl sulfate (SDS) has been investigated by small-angle neutron scattering (SANS), UV-visible spectroscopy, fluorescence spectroscopy, and circular dichroism (CD) spectroscopy. Previous studies on the HbA0-surfactant system using DLS and spectroscopic techniques only confirmed the unfolding of HbA0 without deciphering the exact process and differentiating between the subunit dissociation and extended structure formation. Our spectroscopic results reveal disruption of the heme pocket, loss of Tyr -> Trp energy transfer, and spectral shifts indicative of methemoglobin formation. Utilizing the advantages of SANS, we have explored the subunit dissociation of the tetramer units of HbA0 and their different forms (monomer, dimer, etc.) with the sequential addition of SDS. The results reveal that the addition of SDS at low concentrations leads to the dissociation of the tetramer structure into dimers and monomers, driven by molecular-level binding of SDS monomers to hydrophobic and electrostatic patches at the subunit interfaces rather than by micellization. At higher SDS concentrations, a fraction of nonbound SDS forms free micelles that stabilize the dissociated monomers without promoting further unfolding. In the presence of electrolytes, electrostatic screening enhances protein-surfactant interactions and promotes the formation of extended rod-like complexes. These findings establish a detailed structural pathway for SDS-induced remodelling of HbA0 and highlight the distinct behaviour of multimeric proteins compared to single-chain globular proteins.

Abstract Introduction Macrophage polarization plays a pivotal role in regulating immune responses against infections and cancer. Cyanobacteria such as Spirulina and Leptolyngbya are rich in immunomodulatory metabolites; however, their mechanistic roles in macrophage polarization remain unexplored. This study investigates whether cyanobacterial extracts and their green-synthesized gold nanoparticles (AuNPs) modulate human macrophage polarization. Methods Methanolic extracts of Spirulina and Leptolyngbya and their AuNPs were characterized by UV—Vis, DLS, TEM, and FTIR. THP-1 macrophages were treated with extracts or AuNPs. Expression of M1/M2 gene markers (TNF, IL1β, IL8, CXCL10, CD163, TGFβ, and STAB1) was analyzed by qRT-PCR, while surface markers (CD80 and CD86) were assessed using both qRT-PCR and flow cytometry. Agonist—antagonist assays were conducted to trace the potential pathway. Statistical significant difference was observed using Student t-test and ANOVA (n = 3). Results THP-1 macrophages treated with extracts or AuNPs exhibited significant upregulation of M1 markers TNF-α (p < 0.01), IL-8 (p < 0.01), and IL-1β (p < 0.01), along with a moderate increase in CXCL10 (p < 0.05). Conversely, there was significant downregulation of M2 markers CD163 (p < 0.01), TGFβ (p < 0.05), and STAB1 (p < 0.01). Algal extract-induced polarization occurred via the TLR2 pathway, as it was significantly reversed by TLR2 blockade via a specific antagonist. In contrast, algal-derived AuNPs retained their M1-polarizing capability even after TLR2 inhibition, suggesting activation of TLR2-independent pathways. Conclusion The immunostimulatory response induced by cyanobacterial extracts is partly dependent on TLR-2 signaling. However, extracts AuNps appear to evade TLR2-mediated signaling by internalizing into the macrophages, suggesting activation of alternative pathways leading to M1 polarization. The M1 polarizing ability of extracts and AuNPs highlights their potential as promising therapeutic candidates for cancer immunotherapy. Funding Source n/a Topic Categories Vaccines and Immunotherapy (VAC)

Mercury (Hg) poses a significant environmental and health threat, necessitating highly sensitive and rapid detection methods beyond costly conventional techniques. This study presents a novel electrochemical biosensor for Hg2+ ions, leveraging the specific binding properties of the metalloregulatory protein, i.e., mercuric resistance operon regulatory protein (MerR), covalently immobilized on an Indium Tin Oxide (ITO) surface. We used electrochemically inert methoxy Polyethylene Glycol amino (MPA) as a blocking agent, avoiding the specificity issues associated with traditional Bovine Serum Albumin (BSA). Our MerR/ITO biosensor demonstrated exceptional specificity and achieved a limit of detection (LOD) of 0.77 nM. This performance represents a significant advancement for electrochemical mercury biosensors, particularly considering its robust and direct protein based detection mechanism. This approach offers a highly effective solution for precise mercury monitoring.

Macrophages play a crucial role in innate immunity, regulating inflammatory responses, maintaining tissue homeostasis, and providing immune defense against microbial infections and tumor pathogenesis. Reprogramming macrophages toward the desired functional states, M1/M2 phenotype, is an emerging strategy in immunotherapy, particularly in the context of tumor-associated macrophages (TAMs). Spirulina, a blue-green alga, is rich in bioactive compounds and used as a health supplement. In this study, we explored the immunomodulatory potential of bioactive compounds derived from the commercially available Spirulina and its gold nanoformulations (Sp-GNPs) for the first time using THP-1 monocytic cells. Comprehensive analysis through various biophysical techniques, such as spectroscopy, chromatography, DLS, Zeta potential, FTIR, and electron microscopy, revealed that the methanolic extract of Spirulina is rich in bioactive compounds, including fatty acids, amino acids, glycerol derivatives, and diterpenoids, whereas Sp-GNP exhibited high colloidal stability and surface capping by native biomolecules. Spirulina extract significantly upregulated the expression of M1 signature markers, including TNF-α, IL-1β, IL-8, iNOS, CD80 and CD86, whereas M2-associated markers (IL-10, TGF-β, and STAB-1) remained unchanged in THP-1 macrophages. Intriguingly, Spirulina-derived gold nanoparticles (Sp-GNPs) enhanced the major M1-associated markers such as TNF-α, IL-1β, and IL-8, while simultaneously suppressing IL-10 and STAB-1, indicating their immunostimulatory effects. Toll-like receptor-2 (TLR-2) blockade revealed that Spirulina extract appears to stimulate the NF-κB/MYD88/TLR-2 axis for M1 polarization, and Sp-GNPs induced NF-κB activation via a TLR-2-independent pathway, highlighting a divergent signaling mechanism driven by gold nanoparticles. These findings highlight the potential of Spirulina and its gold nanoformulations as immunomodulators for reprogramming the innate immune system and suggest promising translational value mainly in cancer therapy.

Post-traumatic epilepsy (PTE) is the consequence of traumatic brain damage (TBI), which poses an important health risk for the human population. The underlying mechanism of PTE is complex and appears to be linked with various cellular processes, including oxidative stress and neuroinflammation. Over the years, it has been recognized that most of the available antiseizure medicines are ineffective in preventing PTE. As a result, there is an urgent need to search for alternate treatment options. Naringenin is a flavonoid with multiple pharmacological properties and has shown beneficial effects in several health issues, including neurological disorders. In this study, we investigated the effect of naringenin-loaded polycaprolactone nanoparticles (NarNPs) on neuroinflammatory response in the PTE model. NarNPs were produced using the nanoprecipitation method, and their physicochemical properties were examined by DLS, TEM and AFM. To induce epilepsy in rats, FeCl3 was injected intracortically, and naringenin (NAR), both free NAR and NarNPs, were administered orally, 15 days post-surgery. Epileptic seizures were observed by electroencephalography (EEG) patterns and spectral power analysis of γ-waves. Immunofluorescence analysis of GFAP, IBA1 and TNF-α was performed to examine the disease-modifying potential of NarNPs. Our findings demonstrated that NarNPs marginally reduced epileptiform seizure activity in epileptic rats. Further, the study reported that NarNPs lowered the expression of GFAP, IBA1, and TNF-α. Overall, our data imply that NarNPs have antiseizure and disease-modifying potential by attenuating glial activation and TNF-α production in PTE rats.

Efficient recovery of high-quality DNA is critical for molecular diagnostics, yet conventional extraction methods remain labor-intensive, time-consuming, and frequently affected by inhibitory substances. Herein, a pH-responsive magnetic DNA isolation platform was developed using chitosan-coated iron oxide nanoparticles (IONP@Chitosan). The chitosan-coated magnetic nanohybrids were synthesized via co-precipitation followed by surface coating with chitosan and characterized using FTIR, SEM-EDS, TGA, and DLS, confirming successful surface functionalization. Under acidic conditions, protonated amino groups (-NH 3 + ) facilitated electrostatic interaction with the negatively charged phosphate backbone of genomic DNA, whereas alkaline conditions enabled efficient DNA desorption. In binding experiments, IONP@Chitosan adsorbed > 90% of genomic DNA with ~ 76% recovery during elution, substantially exceeding the performance of uncoated iron oxide nanoparticles (< 40% adsorption, ~ 8% recovery). From 1 mL bacterial cultures, the platform yielded 8.5 µg and 7.6 µg of genomic DNA from Escherichia coli and Staphylococcus aureus , respectively, with acceptable purity. DNA quality and integrity were validated by agarose gel electrophoresis, UV-visible spectroscopy, and successful PCR amplification of the uidA and nuc genes. Notably, DNA-nanoparticle complexes were directly compatible with PCR, enabling successful amplification without prior elution and supporting the feasibility of simplified molecular workflows. These findings suggest that chitosan-functionalized magnetic nanomaterials may serve as promising chaotropic salt-free platforms for genomic DNA isolation with potential utility in molecular diagnostic applications.

Hypoxic tumors remain a significant clinical challenge due to their resistance to conventional therapies, highlighting the need for targeted diagnostic and therapeutic strategies. In this study, we report the design of an iridium-based prodrug (Ir-NO₂) incorporating a p-nitrobenzyl moiety that functions as a nitroreductase (NTR)-responsive trigger, exploiting elevated NTR levels in hypoxic tumor environments. The prodrug preferentially accumulates in mitochondria and undergoes bioreduction by NTR to release the active species, Ir-NH₂. Ir-NO₂ exhibits significant cytotoxicity against 2D cancer cell lines, including human breast (MCF-7) and triple-negative breast (MDA-MB-231) cells, with IC₅₀ values ranging from 3.12 to 8.34 μM under normoxic conditions (48–72 h, MTT assay) and 12.21 μM (MCF-7) and 16.03 μM (MDA-MB-231) under hypoxia (48 h). In contrast, the clinically approved drug cisplatin shows substantially lower potency (IC₅₀ ≥ 50 μM in most cases), although enhanced activity was observed in MCF-7 cells at 72 h (IC₅₀ = 21.83 μM). Importantly, Ir-NO₂ demonstrates efficient penetration and significant antitumor activity in 3D multicellular tumor spheroids (MCTSs) of MCF-7 cells, achieving an IC₅₀ of 7.66 μM, indicative of improved performance in physiologically relevant models. Mechanistic studies reveal that Ir-NO₂ induces apoptotic cell death through mitochondrial membrane depolarization and increased intracellular reactive oxygen species (ROS) generation.

Porous substrates are crucial for suspended lipid bilayers, yet the influence of pore dimensions on bilayer stability and function remains underexplored. Understanding how these complementary platforms affect bilayer properties is essential for developing reliable biomimetic systems, and studying transmembrane proteins. We fabricated pores at different scales (micro- and nanoscale) by employing an aluminium-based substrate, using manual puncturing for micropores and anodization for nanopores. This approach was designed to address the intricate role of porous substrates in modulating the bilayer suspension, stability and function of 1-palmitoyl-2-oleoyl-glycero-3-phosphocholine (POPC) lipid bilayers. Micropores provide optical access for real-time imaging of bilayer coverage and defects, while nanopores offer stronger mechanical stability, as confirmed by impedance spectroscopy with giga-seal resistance (∼2 GΩ). Using this stable platform, aquaporin, a water-channel protein, was reconstituted into lipid bilayers using an n-octyl-β-d glucopyranoside (NOG) detergent-mediated method. Dynamic light scattering, zeta potential, and impedance analyses confirmed successful insertion, and forward osmosis assays demonstrated functional water transport. The study represents a comparative framework, demonstrating the trade-off between accessibility and stability and providing design principles for future bilayer-based biomimetic and protein reconstitution platforms.

Advanced Glycation End Products (AGEs) form as a result of non-enzymatic addition of reducing sugars to proteins, producing a chemically heterogeneous set of compounds with altered physicochemical properties. Generally, AGEs are considered pro-cancerous in nature due to their reactivity with extracellular, intracellular and membranous components of the cancerous-tissue microenvironment. Despite many studies on AGE-Cancer association, direct evidence linking glycation product preparations to endocrine therapy remains limited. In this study, Hemoglobin Advanced Glycation End Products (Hb-AGEs) have been generated in-vitro by incubating a mixture of hemoglobin and fructose at physiological (37 °C) and accelerated (60 °C) conditions. The end products have been characterized by biophysical and biochemical techniques followed by a functional testing in MCF-7 breast cancer cells. We show that the Hb-AGEs formed in our study contain reducing and radical scavenging (antioxidant) properties which are enhanced under accelerated (at 60 °C) conditions. Functionally, co-treatment of MCF-7 breast cancer cells with Hb-AGEs and tamoxifen protected the cells from tamoxifen toxicity. Given the chemical complexity of the reaction products and lack of mechanistic cellular assays, the observed modulation in tamoxifen response may reflect multiple non-exclusive mechanisms. Rather than simple antioxidant neutralization of the drug-response, these mechanisms may also include altered drug bioavailability, cell-membrane related changes and other downstream stress pathways. Nevertheless, our findings from this study provide a preliminary biochemical and functional framework to further investigate how glycation products may influence chemotherapeutic response in-vitro.

Traditional systemic drug delivery methods are often limited by faster clearance rates and high dose requirements. While innovative drug delivery systems, such as liposomes, polymersomes, and solid lipid nanoparticles etc., offer the ability to carry hydrophilic and hydrophobic drugs. But the lack of specificity in the current delivery systems leads to reduced therapeutic efficacy. To overcome these, functionalization with small molecules like peptides, aptamers, and antibodies is done. But they introduce new immunogenicity, stability, and complexity challenges, making these methods costly and less practical. To address these issues, circulatory cell-based drug delivery systems have gained attention as a promising alternative. Unlike red blood cells (RBCs), which have the Rh factor that can complicate, platelets are devoid of this factor and possess inherent advantages. Their natural properties, including non-immunogenicity, hemostasis, pathogen interaction, tissue regeneration, and wound healing, make them highly effective for targeted drug delivery. Furthermore, platelets can form leaky junctions at tumor sites, enhancing localized drug delivery and improving therapeutic outcomes. Thus, platelet-based drug delivery systems are gaining popularity. Moreover, the lower production costs make platelets an efficient and promising tool for a smart drug delivery system. This review highlights the significant role of platelets in advancing targeted drug delivery and their potential to revolutionize therapeutic strategies.

Biomolecules and excipients can modulate membrane properties, initiate a string of events, and regulate their functionality. To elucidate these phenomena ex vivo, we prepared a three-component lipid model system that consisted of varying proportions of cholesterol, sphingomyelin, and 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC). To decipher the membrane dynamics, we included a nonmembranous protein, hemoglobin, in our study. Our initial investigation revealed the formation of self-assembled structures and phase separation in a pure POPC-based synthetic membrane when hemoglobin was reconstituted via a detergent-mediated method. A slight unfolding helped the protein adapt to the huge hydrophobic stress of the lipid environment and led to the formation of these self-assembled structures. To identify an optimal lipid composition that mimics the biological membrane, we employed three varying proportions of lipid mixtures: POPC, sphingomyelin, and cholesterol. We examine events like the formation of lipid bilayers, supramolecular structures, and phase separation using techniques like FRAP, FCS, AFM, Z-stacking, and rheology. We observed the variation in condensate formation and its distribution within the membrane, which differs upon an increase in concentration of sphingomyelin and cholesterol. Such membrane behavior is important for raft formation and signaling.

Cancer is a multifactorial and complex disease, typically managed through a combination of therapies including surgery, chemotherapy, and radiation; however, it remains a leading cause of mortality worldwide, with solid tumors such as lung, breast, and pancreatic cancers continuing to pose significant therapeutic challenges. A key contributor to treatment failure in these malignancies is tumor hypoxia, an oxygen-deficient microenvironment that reprograms cancer cell metabolism, promotes immune evasion, and fosters drug resistance. Although hemoglobin (Hb) is a natural oxygen carrier, its therapeutic application is limited due to structural instability, oxidative degradation, and dose-dependent cytotoxicity, as well as its tendency to elicit inflammatory responses when administered in free form. To address this, we develop hemoglobin-loaded polycaprolactone nanoparticles (PCL-Hb NPs) for targeted oxygen delivery within hypoxic tumors. Hemoglobin is encapsulated in a stable core-shell structure, retaining its oxygen-binding capacity and structural integrity as confirmed by biophysical and spectroscopic analysis. Oxygen release assays confirm sustained and reversible oxygen diffusion. In vitro studies demonstrated that PCL-Hb nanoparticles are biocompatible, showing no cytotoxicity in HFL1 normal lung fibroblast and A549 lung cancer cells and no inflammatory activation in THP-1 macrophages. Flow cytometry and confocal microscopy revealed efficient cellular uptake both under normoxic and hypoxic conditions. Under hypoxic conditions, PCL-Hb reduced the expression of hypoxia-responsive genes and effectively reverses resistance to paclitaxel. Simultaneous treatment of PCL-Hb with paclitaxel significantly enhances drug efficacy in 2D hypoxic cells and 3D spheroids, restoring sensitivity and improving therapeutic response. These findings underscore the potential of PCL-Hb nanoparticles as an oxygen delivery system to alleviate tumor hypoxia and overcome hypoxia-induced chemotherapy resistance in solid tumors.

Cancer is a predominant cause of death among the human population. The conventional treatment options for this devastating disease are limited to chemo and radiation therapy, which can cause systemic toxicity due to off-target release. To address these challenges, an innovative therapeutic approach of biomimetic nanoformulations can be adopted. Biomimetic nanoformulations have the characteristics of synthetic nanoformulations and functional properties of cellular membranes. Cell-membrane-based biomimetic nanoformulations (CMBNs) are a type of biomimetic nanoformulation that holds promise to treat cancer due to their precise targeting capabilities. This review highlights the types of biomimetic nanoformulations, including the development strategies and application of CMBNs in drug delivery and immunotherapy. The membrane extraction and fusion processes of CMBNs are also highlighted, along with the challenges and limitations of their large-scale production and clinical applications. Furthermore, we shed light on the complex surface chemistry of various cell membranes and their functional components, as well as the different types of cores used for the synthesis of CMBNs. The application of CMBNs in targeted drug delivery has also been explored, along with the technical hurdles and optimization of the design and functionality of these nanoformulations. Overall, the review emphasized that CMBNs hold promise for revolutionising drug delivery and immune therapies in cancer.