[Research on Dry Chemical Detection TechnologyBased on Reflectance Photometry].

In response to the issues of insufficient stability and accuracy in dry chemical detection using reflectance photometry, caused by the divergence and multiple internal reflections of the reflected light signal from the sample and the multilayer dry film test strip, a dry chemical reflectance photometry detection system based on an integrating sphere is designed. Firstly, an integrating sphere device is incorporated to reduce signal divergence and loss, ensuring even detection of the sample's reflected light signal and improving detection stability. Secondly, Light Tools optical simulation analysis is performed, and an integrating sphere detection model is established. Thirdly, the Williams-Clapper equation is employed to correct the error in reflectance density caused by multiple internal reflections, enhancing detection accuracy. Experimental validation demonstrates that the developed integrating sphere-based dry chemical reflectance photometry detection system improves the stability and accuracy of the detection system.

Tripedal DNA Walker as a Signal Amplifier Combined with a Potential-Resolved Multicolor Electrochemiluminescence Strategy for Ultrasensitive Detection of Prostate Cancer Staging Indicators.

A multicolor electrochemiluminescence (ECL) biosensor based on a closed bipolar electrode (BPE) array was proposed for the rapid and intuitive analysis of three prostate cancer staging indicators. First, [Irpic-OMe], [Ir(ppy)2(acac)], and [Ru(bpy)3]2+ were applied as blue, green, and red ECL emitters, respectively, whose mixed ECL emission colors covered the whole visible region by varying the applied voltages. Afterward, we designed a simple Mg2+-dependent DNAzyme (MNAzyme)-driven tripedal DNA walker (TD walker) to release three output DNAs. Immediately after, three output DNAs were added to the cathodic reservoirs of the BPE for incubation. After that, we found that the emission colors from the anode of the BPE changed as a driving voltage of 8.0 V was applied, mainly due to changes in the interfacial potential and faradaic currents at the two poles of the BPE. Via optimization of the experimental parameters, cutoff values of such three indicators at different clinical stages could be identified instantly with the naked eye, and standard precision swatches with multiple indicators could be prepared. Finally, in order to precisely determine the prostate cancer stage, the multicolor ECL device was used for clinical analysis, and the resulting images were then compared with standard swatches, laying the way for accurate prostate cancer therapy.

Confined DNA tetrahedral molecular sieve for size-selective electrochemiluminescence sensing.

Size selectivity is crucial in highly accurate preparation of biosensors. Herein, we described an innovative electrochemiluminescence (ECL) sensing platform based on the confined DNA tetrahedral molecular sieve (DTMS) for size-selective recognition of nucleic acids and small biological molecule. Firstly, DNA template (T) was encapsulated into the inner cavity of DNA tetrahedral scaffold (DTS) and hybridized with quencher (Fc) labeled probe DNA to prepare DTMS, accordingly inducing Ru(bpy)32+ and Fc closely proximate, resulting the sensor in a "signal-off" state. Afterwards, target molecules entered the cavity of DTMS to realize the size-selective molecular recognition while prohibiting large molecules outside of the DTMS, resulting the sensor in a "signal-on" state due to the release of Fc. The rigid framework structure of DTS and the anchor of DNA probe inside the DTS effectively avoided the nuclease degradation of DNA probe, and nonspecific protein adsorption, making the sensor possess potential application prospect for size-selective molecular recognition in diagnostic analysis with high accuracy and specificity.

Self-Assembled Tetraphenylethene-Based Nanoaggregates with Tunable Electrochemiluminescence for the Ultrasensitive Detection of E. coli.

As an effective ECL emitter, tetraphenylethene (TPE)-based molecules have recently been reported with aggregation-induced electrochemiluminescence (AIECL) property, while it is still a big challenge to control its aggregation states and obtain uniform aggregates with intense ECL emission. In this study, we develop three TPE derivatives carrying a pyridinium group, an alkyl chain, and a quaternary ammonium group via the Menschutkin reaction. The resulting molecules exhibit significantly red-shifted FL and enhanced ECL emissions due to the tunable reduction of the energy gap between the highest occupied molecular orbitals (HOMOs) and the lowest unoccupied molecular orbitals (LUMOs). More importantly, the amphiphilicity of the as-developed molecules enables their spontaneous self-assembly into well-controlled spherical nanoaggregates, and the ECL intensity of nanoaggregates with 3 -CH2- (named as C3) is 17.0-fold higher compared to that of the original 4-(4-(1,2,2-triphenylvinyl)phenyl)pyridine (TPP) molecule. These cationic nanoaggregates demonstrate a high affinity toward bacteria, and an ECL sensor for the profiling of Escherichia coli (E. coli) was developed with a broad linear range and good selectivity in the presence of an E. coli-specific aptamer. This study provides an effective way to enhance the ECL emission of TPE molecules through their derivatization and a simple way to prepare well-controlled AIECL nanoaggregates for ECL application.

Potential-Resolved Ratiometric Aptasensor for Sensitive Acetamiprid Analysis Based on Coreactant-free Electrochemiluminescence Luminophores of Gd-MOF and "Light Switch" Molecule of [Ru(bpy)2dppz]2.

For conventional potential-resolved ratiometric electrochemiluminescence (ECL) systems, the introduction of multiplex coreactants is imperative. However, the undesirable interactions between different coreactants inevitably affect analytical accuracy and sensitivity. Herein, through the coordination of aggregation-induced emission ligands with gadolinium cations, the self-luminescent metal-organic framework (Gd-MOF) is prepared and serves as a novel coreactant-free anodic ECL emitter. By the intercalation of [Ru(bpy)2dppz]2+ with light switch effect into DNA duplex, one high-efficiency cathodic ECL probe is obtained using K2S2O8 as a coreactant. In the presence of acetamiprid, the strong affinity between the target and its aptamer induces the release of [Ru(bpy)2dppz]2+, resulting in a decreasing cathode signal and an increasing anode signal owing to the ECL resonance energy transfer from Gd-MOF to [Ru(bpy)2dppz]2+. In this way, an efficient dual-signal ECL aptasensor is constructed for the ratiometric analysis of acetamiprid, exhibiting a remarkably low detection limit of 0.033 pM. Strikingly, by using only one exogenous coreactant, the cross interference from multiple coreactants can be eliminated, thus improving the detection accuracy. The developed high-performance ECL sensing platform is successfully applied to monitor the residual level of acetamiprid in real samples, demonstrating its potential application in the field of food security.

Combined Experiments with in Vivo Fiber Photometry and Behavior Tests Can Facilitate the Measurement of Neuronal Activity in the Primary Somatosensory Cortex and Hyperalgesia in an Inflammatory Pain Mice Model.

The pain matrix, which includes several brain regions that respond to pain sensation, contribute to the development of chronic pain. Thus, it is essential to understand the mechanism of causing chronic pain in the pain matrix such as anterior cingulate (ACC), or primary somatosensory (S1) cortex. Recently, combined experiment with the behavior tests and in vivo calcium imaging using fiber photometry revealed the interaction between the neuronal function in deep brain regions of the pain matrix including ACC and the phenotype of chronic pain. However, it remains unclear whether this combined experiment can identify the interaction between neuronal activity in S1, which receive pain sensation, and pain behaviors such as hyperalgesia or allodynia. In this study, to examine whether the interaction between change of neuronal activity in S1 and hyperalgesia in hind paw before and after causing inflammatory pain was detected from same animal, the combined experiment of in vivo fiber photometry system and von Frey hairs test was applied. This combined experiment detected that amplitude of calcium responses in S1 neurons increased and the mechanical threshold of hind paw decreased from same animals which have an inflammatory pain. Moreover, we found that the values between amplitude of calcium responses and mechanical thresholds were shifted to negative correlation after causing inflammatory pain. Thus, the combined experiment with fiber photometry and the behavior tests has a possibility that can simultaneously consider the interaction between neuronal activity in pain matrix and pain induced behaviors and the effects of analgesics or pain treatments.

Metal-organic framework (MOF)-based biosensors for miRNA detection.

MicroRNAs (miRNAs) play several crucial roles in the physiological and pathological processes of the human body. They are considered as important biomarkers for the diagnosis of various disorders. Thus, rapid, sensitive, selective, and affordable detection of miRNAs is of great importance. However, the small size, low abundance, and highly similar sequences of miRNAs impose major challenges to their accurate detection in biological samples. In recent years, metal-organic frameworks (MOFs) have been applied as promising sensing materials for the fabrication of different biosensors due to their distinctive characteristics, such as high porosity and surface area, tunable pores, outstanding adsorption affinities, and ease of functionalization. In this review, the applications of MOFs and MOF-derived materials in the fabrication of fluorescence, electrochemical, chemiluminescence, electrochemiluminescent, and photoelectrochemical biosensors for the detection of miRNAs and their detection principle and analytical performance are discussed. This paper attempts to provide readers with a comprehensive knowledge of the fabrication and sensing mechanisms of miRNA detection platforms.

Evaluation of size-exclusion chromatography, multi-angle light scattering detection and mass photometry for the characterization of mRNA.

The recent approval of messenger ribonucleic acid (mRNA) as vaccine to combat the COVID-19 pandemic has been a scientific turning point. Today, the applicability of mRNA is being demonstrated beyond infectious diseases, for example in cancer immunotherapy, protein replacement therapy and gene editing. mRNA is produced by in vitro transcription (IVT) from a linear DNA template and modified at the 3' and 5' ends to improve translational efficiency and stability. Co-existing impurities such as RNA fragments and double-stranded RNA (dsRNA), amongst others, can drastically impact mRNA quality and efficacy. In this study, size-exclusion chromatography (SEC) is evaluated for the characterization of IVT-mRNA. The effect of mobile phase composition (ionic strength and organic modifier), pH, column temperature and pore size (300 Å, 1000 Å, and 2000 Å) on the separation performance and structural integrity of IVT-mRNA varying in size is described. Non-replicating, self-amplifying (saRNA), temperature degraded, and ribonuclease (RNase) digested mRNA, the latter to characterize the 3' poly(A) tail, were included in the study. Beyond ultraviolet (UV) detection, refractive index (RI) and multi-angle light scattering (MALS) detection were implemented to accurately determine molecular weight (MW) of mRNA. Finally, mass photometry is introduced as a complementary methodology to study mRNA under native conditions.

Comprehensive Impurity Profiling of mRNA: Evaluating Current Technologies and Advanced Analytical Techniques.

In vitro transcription (IVT) of mRNA is a versatile platform for a broad range of biotechnological applications. Its rapid, scalable, and cost-effective production makes it a compelling choice for the development of mRNA-based cancer therapies and vaccines against infectious diseases. The impurities generated during mRNA production can potentially impact the safety and efficacy of mRNA therapeutics, but their structural complexity has not been investigated in detail yet. This study pioneers a comprehensive profiling of IVT mRNA impurities, integrating current technologies with innovative analytical tools. We have developed highly reproducible, efficient, and stability-indicating ion-pair reversed-phase liquid chromatography and capillary gel electrophoresis methods to determine the purity of mRNA from different suppliers. Furthermore, we introduced the applicability of microcapillary electrophoresis for high-throughput (<1.5 min analysis time per sample) mRNA impurity profiling. Our findings revealed that impurities are mainly attributed to mRNA variants with different poly(A) tail lengths due to aborted additions or partial hydrolysis and the presence of double-stranded mRNA (dsRNA) byproducts, particularly the dsRNA 3'-loop back form. We also implemented mass photometry and native mass spectrometry for the characterization of mRNA and its related product impurities. Mass photometry enabled the determination of the number of nucleotides of different mRNAs with high accuracy as well as the detection of their size variants [i.e., aggregates and partial and/or total absence of the poly(A) tail], thus providing valuable information on mRNA identity and integrity. In addition, native mass spectrometry provided insights into mRNA intact mass, heterogeneity, and important sequence features such as poly(A) tail length and distribution. This study highlights the existing bottlenecks and opportunities for improvement in the analytical characterization of IVT mRNA, thus contributing to the refinement and streamlining of mRNA production, paving the way for continued advancements in biotechnological applications.

Portable, intelligent MIECL sensing platform for ciprofloxacin detection using a fast convolutional neural networks-assisted Tb@Lu2O3 nanoemitter.

Environmental pollution caused by ciprofloxacin is a major problem of global public health. A machine learning-assisted portable smartphone-based visualized molecularly imprinted electrochemiluminescence (MIECL) sensor was developed for the highly selective and sensitive detection of ciprofloxacin (CFX) in food. To boost the efficiency of electrochemiluminescence (ECL), oxygen vacancies (OVs) enrichment was introduced into the flower-like Tb@Lu2O3 nanoemitter. With the specific recognition reaction between MIP as capture probes and CFX as detection target, the ECL signal significantly decreased. According to, CFX analysis was determined by traditional ECL analyzer detector in the concentration range from 5 × 10-4 to 5 × 102 µmol L-1 with the detection limit (LOD) of 0.095 nmol L-1 (S/N = 3). Analysis of luminescence images using fast electrochemiluminescence judgment network (FEJ-Net) models, achieving portable and intelligent quick analysis of CFX. The proposed MIECL sensor was used for CFX analysis in real meat samples and satisfactory results, as well as efficient selectivity and good stability.

A multi-modal biosensing platform based on Ag-ZnIn2S4@Ag-Pt nanosignal probe-sensitized UiO-66 for ultra-sensitive detection of penicillin.

We designed a multi-modal biosensing platform for versatile detection of penicillin based on a unique Ag-ZnIn2S4@Ag-Pt signal probe-sensitized UiO-66 metal-organic framework. Firstly, a large number of Ag-ZnIn2S4 quantum dots (AZIS QDs) were attached to Ag-Pt NPs, preparing a new multi-signal probe AZIS QDs@Ag-Pt NPs with excellent photoelectrochemistry (PEC), electrochemiluminescence (ECL), and fluorescence (FL) signals. Moreover, the AZIS QDs@Ag-Pt NPs signal probe can well match the energy level of UiO-66 metal-organic framework (MOF) with good photoelectric property, which can reverse the PEC current of UiO-66 to reduce false positives in detection. When penicillin was present, it bound to its aptamer to release the multifunctional signal probes, which can generate PEC, ECL, and PL signals, thus realizing ultrasensitive detection of penicillin by multi-signals. This work creates a novel three-signal QDs probe, which makes a great contribution to multi-mode photoelectric sensing analysis. The LOD of this work (3.48 fg·mL-1) was much lower than the MRLs (Maximum Residue Levels) established by the EU (4 ng·mL-1). The newly developed multi-mode biosensor has good practical application values in various biological detection, food assay, and early disease diagnosis.

Rigidifying AIEgens in covalent organic framework nanosheets for electrochemiluminescence enhancement: TABE-PZ-CON as a novel emitter for microRNA-21 detection.

Enhancing electrochemiluminescence (ECL) properties of luminophores is a hot direction in the current ECL field. Herein, we found that covalent rigidification of the aggregation-induced emission luminogens (AIEgens) TABE (TABE = tetra-(4-aldehyde-(1,1-biphenyl))ethylene) into covalent organic framework nanosheets (TABE-PZ-CON, PZ = piperazine) could result in stronger ECL emission than those of TABE aggregates and TABE monomers. We termed the interesting phenomenon "covalent rigidification-triggered electrochemiluminescence (CRT-ECL) enhancement". The superior ECL performance of TABE-PZ-CON not only because massive TABE luminogens were covalently assembled into the rigid TABE-PZ-CON network, which limited the intramolecular motions of TABE and hampered the radiationless transition, but also because the ultrathin porous TABE-PZ-CON significantly reduced the transportation distance of ions, electrons, and coreactants, which enabled the electrochemical excitation of more TABE luminogens and thus enhanced the ECL efficiency. Bearing in mind the exceptional ECL performance of TABE-PZ-CON, it was utilized as a high-efficient ECL indicator in combination with the DNA walker and duplex-specific nuclease-assisted target recycling amplification strategies to design an "off-on" ECL biosensor for the ultrasensitive assay of microRNA-21, exhibiting a favorable response range (100 aM-1 nM) with an ultralow detection limit of 17.9 aM. Overall, this work offers a valid way to inhibit the intramolecular motions of AIEgens for ECL enhancement, which gives a new vision for building high-performance AIEgen-based ECL materials, thus offering more chances for assembling hypersensitive ECL biosensors.

Analysis of Protein Complex Formation at Micromolar Concentrations by Coupling Microfluidics with Mass Photometry.

Mass photometry is a versatile mass measurement technology that enables the study of biomolecular interactions and complex formation in solution without labels. Mass photometry is generally suited to analyzing samples in the 100 pM-100 nM concentration range. However, in many biological systems, it is necessary to measure more concentrated samples to study low-affinity or transient interactions. Here, we demonstrate a method that effectively expands the range of sample concentrations that can be analyzed by mass photometry from nanomolar to tens of micromolar. In this protocol, mass photometry is combined with a novel microfluidics system to investigate the formation of protein complexes in solution in the micromolar concentration range. With the microfluidics system, users can maintain a sample at a desired higher concentration followed by dilution to the nanomolar range - several milliseconds prior to the mass photometry measurement. Due to the speed of the dilution, data is obtained before the equilibrium of the sample has shifted (i.e., dissociation of the complex). The technique is applied to measure interactions between an immunoglobulin G (IgG) antibody and the neonatal Fc receptor, showing the formation of high-order complexes that were not quantifiable with static mass photometry measurements. In conclusion, the combination of mass photometry and microfluidics makes it possible to characterize samples in the micromolar concentration range and is proficient in measuring biomolecular interactions with weaker affinities. These capabilities can be applied in a range of contexts - including the development and design of biotherapeutics - enabling thorough characterization of diverse protein-protein interactions.

Dual-microRNA-Controlled Electrochemiluminescence Biosensor for Breast Cancer Diagnosis and Supplemental Identification of Breast Cancer Metastasis.

Breast cancer remains the most frequently diagnosed cancer globally, and the metastasis of this malignancy is the primary cause of mortality in breast cancer patients. Hence, prompt diagnosis and timely detection of metastatic breast cancer are critical for effective therapeutic intervention. Both progression and metastasis of this malignancy are closely associated with aberrant expression of specific microRNAs (miRNAs) and enzymes. To facilitate breast cancer diagnosis and concomitant identification of metastatic breast cancer, we have engineered an innovative electrochemiluminescence (ECL)-based sensing platform integrated with enzyme-free DNA amplification circuits for dual functionality. Specifically, microRNA-21 (miR-21) is employed as a biomarker for breast cancer, and miR-21 induces the quenching of the ECL signal from luminophores via a strategically designed catalytic three-hairpin assembly (CTHA) circuit. Subsequently, miR-105 levels are measured via toehold-mediated strand displacement reactions (TSDR). Here, miR-105 restores the initially quenched ECL signal, enabling the assessment of the metastatic propensity. Our experimental data demonstrate that the devised ECL biosensor offers broad linear detection ranges and low detection limits for both miR-21 and miR-105. Importantly, our novel platform was also successfully validated by using cellular and serum samples. This biosensor not only discriminates breast cancer cell lines MCF-7 and MDA-MB-231 from nonbreast cancer cells like HepG2, TPC-1, and HeLa, but it also distinguishes between malignant MCF-7 and metastatic MDA-MB-231 cells. Consequently, our novel approach holds significant promise for clinical applications and precise cancer screening.

Automated Mass Photometry of Adeno-Associated Virus Vectors from Crude Cell Extracts.

Mass photometry (MP) is a fast and simple analysis method for the determination of the proportions of subpopulations in an AAV sample. It is label-free and requires minimal sample volumes between 5-10 µL, which makes it a promising candidate over orthogonal techniques such as analytical ultracentrifugation (AUC), cryo-transmission electron microscopy (Cryo-TEM) or charge-detection mass spectrometry (CDMS). However, these methods are limited in their application to purified samples only. Here we developed a purification step based on single-domain monospecific antibody fragments immobilised on either a poly(styrene-divinylbenzene) resin or on magnetic beads prior to MP analysis that allows the quantification of empty, partially filled, full and overfull AAV vectors in crude cell extracts. This is aimed at identifying potentially promising harvest conditions that yield large numbers of filled AAV vectors during the early stages of the viral vector development platform, e.g., the type of transfection reagent used. Furthermore, we provide a direct comparison of the automated and manual handling of the mass photometer with respect to the quantities of AAV subspecies, molar mass of the capsid and payload, and highlight the differences between the "buffer-free" sample measurement and the "buffer-dilution" mode. In addition, we provide information on which candidates to use for calibration and demonstrate the limitations of the mass photometer with respect to the estimation of the capsid titer.

Screening for juvenile idiopathic arthritis associated uveitis with laser flare photometry in the pediatric rheumatology office: a prospective observational study.

BACKGROUND: Juvenile Idiopathic Arthritis (JIA) Associated Uveitis (JIA-U) remains one of the most serious complications of JIA in children. Historically, pediatric JIA is diagnosed by an Optometrist or Ophthalmologist; however, barriers to scheduling increase wait times that may delay diagnosis and treatment. The purpose of this study was to evaluate laser flare photometry (LFP) use to diagnose JIA-U in the Pediatric Rheumatology clinic for patients with JIA. METHODS: This prospective, observational study assessed pediatric patients diagnosed with JIA without a previous history of uveitis between January 2020 and September 2022. All patients underwent at least one evaluation of both eyes using a Kowa FM-600 laser flare photometer during a routine Rheumatology appointment, as well as a standard slit lamp examination (SLE) by optometry or ophthalmology during routine clinical care. Data collected at patient visits included demographics, JIA characteristics, treatment, LFP readings, and anterior chamber (AC) cell grade score utilizing the SUN grading system. Data were summarized using descriptive analyses and the uveitis false positive rate was calculated. RESULTS: The study cohort included 58 pediatric patients diagnosed with JIA. The mean age was 8.4 years (1.2-16.3 years) at diagnosis and 11.9 (4.8-16.5 years) at enrollment. The mean duration of disease at time of enrollment was 42 months (range; 0-157 months). Participants were predominantly female (n = 43, 74.1%) and white/Caucasian race (n = 37, 63.8%). The most common JIA subtypes included persistent oligoarticular JIA (n = 19, 32.8%), and RF negative polyarticular JIA (n = 12, 20.7%). There were 12 ANA positive patients (20.7%). At enrollment, 16 patients (27.6%) were not on medications, with 20 (34.5%) on methotrexate, 20 (34.5%) on adalimumab, 6 (10.3%) on tocilizumab, and 5 (8.6%) on etanercept. During the study period, no eye exams detected active uveitis based on SLE with a SUN grade over 0. However, of the 135 LFP readings, 131 (97.0%) were normal, yielding a false positive rate of 3% (95% CI: 0.8%, 7.4%). CONCLUSIONS: LFP is a non-invasive tool that can be utilized in the pediatric rheumatology clinic to evaluate for JIA-U. There is a low false positive rate of LFP when compared with standard slit lamp exam.

Porous Complex-Mediated Dual Emission of Porphyrins for the Electrochemiluminescence Bioassay.

Although porphyrins make up a promising class of electrochemiluminescence (ECL) luminophors, their aggregation-caused quenching (ACQ) characteristics lead to inferior ECL efficiency (ΦECL). Furthermore, current application of porphyrins is limited to cathodic emission. This work creatively exploited a cage-like porous complex (referred to as SWU-1) as the microreactor to recede the ACQ effect while modulating dual ECL emission of meso-tetra(4-carboxyphenyl)porphine (TCPP), which self-assembled with SWU-1 to form TCPP@SWU-1 nanocapsules (TCPP@SWU-1 NCs). As the microreactor, SWU-1 not only effectively constrained TCPP aggregation to improve electron-hole recombination efficiency but also improved stability of anion and cation radicals, thus significantly enhancing the dual emission of TCPP. Compared with TCPP aggregates, the resulting TCPP@SWU-1 NCs exhibited significantly enhanced anodic and cathodic emission, and their ΦECL was increased by 8.7-fold and 3.9-fold, respectively. Furthermore, black hole quencher-2 (BHQ2) can simultaneously quench anodic and cathodic signals. TCPP@SWU-1 NCs coupling BHQ2 conveniently achieved an ECL ratio detection of miRNA-126, and the limit of detection (S/N = 3) was 4.1 aM. This work pioneered the development of the cage-like porous complex SWU-1 as the microreactor to alleviate defects of the ACQ effect and mediate dual emission of TCPP. The coupling of dual-emitting TCPP@SWU-1 NCs and dual-function moderator BHQ2 created a novel single-luminophor-based ratio system for bioanalysis and provided a promising ECL analysis approach for miRNA-126.

Ti3C2 MXene-based nanozyme as coreaction accelerator for enhancing electrochemiluminescence of glucose biosensing.

Delamination of the exfoliated multilayer MXenes with electro-catalysts, not only leads to increasing surface area for high electrochemiluminescent (ECL) signal tracer loading but also provides highly sensitive achievements in a coreaction accelerator manner. To this end, herein, we used bromophenol blue (BPB)-delaminated multilayer Ti3C2 MXene as both a coreaction accelerator to promote the electrochemiluminescent (ECL) reaction rate of luminol (LUM) and the co-reactant H2O2 and a substrate for retaining high loading of glucose oxidase (GOx)-conjugated polyethylene imine (PEI) along with luminophore species into more open structure of Ti3C2 MXene for sensitive detection of glucose. In the presence of glucose, in situ generating H2O2 product through a GOx-catalyzed process could produce abundant •OH radicals via the peroxidase-like activity of the BPB@Ti3C2 in the LUM ECL reaction. Moreover, decreasing the distance between the high-content LUM into the BPB@Ti3C2 and the generated •OH, minimizes the decomposition of highly active •OH, providing a superb ECL signal. Last, the proximity of incorporated GOx into the delaminated Ti3C2 MXene near the electrode allows efficient electron transfer between the electrode and enzyme. The integration of such amplifying effects endowed high sensitivity and excellent selectivity for glucose with a low limit of detection of 0.02 µM in the wide range of 0.01 µM-40,000 µM, enabling the feasibility of the glucose analysis in human serum samples. Overall, the enhanced ECL based on the BPB@Ti3C2 opens a new horizon to develop highly sensitive MXene-based ECL toward the field of biosensors.

Measuring Protein-Protein Interactions and Quantifying Their Dissociation Constants with Mass Photometry.

Protein-protein interactions underlie most biological processes, and determining the affinity and abundance of binding partners for each interaction is often a challenging task because these interactions often involve multiple ligands and binding sites. Standard methods for determining the affinity of protein interactions often require a large amount of starting material in addition to potentially disruptive labeling or immobilization of the binding partners. Mass photometry is a bioanalytical technique that measures the mass of single biomolecules in solution, quickly and with minimal sample requirements. This article describes how mass photometry can be used to determine the mass distribution of binding partners, the complexes they form, the relative abundance of each species, and, accordingly, the dissociation constant (KD ) of their interactions. © 2024 Refeyn Ltd. Current Protocols published by Wiley Periodicals LLC. Basic Protocol: Using mass photometry to measure protein-protein binding and quantify the KD of this interaction.

Measuring macular pigment optical density using reflective images of confocal scanning laser system.

PURPOSE: To develop a method to measure the macular pigment optical density (MPOD) using scanning laser ophthalmoscopic images in young adults and children. STUDY DESIGN: Cross-sectional study. METHODS: Blue light reflectance fundus images of 32 healthy subjects were used. A profile of the linear reflectance changes across the center of the fovea on a grayscale fundus image was generated. The ratio of the macula-to-periphery reflection was designated as the peak value of the MPOD (MPOD[FR]) based on established fundamentals. In the MPOD profile, the basal width of the pixels at MPOD < 0 (wMP) and width at one-half value of the MPOD[FR] (wMP0.5) were determined. The MOPD at eccentricity of 0.5° was measured by heterochromatic flicker photometry (MPOD[HFP]), and the correlation between the MPOD[FR] and MPOD[HFP] was evaluated. RESULTS: The MPOD[FR] ranged from 0.17 to 0.73 with a mean of 0.40 ± 0.13. The wMP ranged from 88 to 173 pixels with a mean of 121.7 ± 24.2 pixels, and the wMP0.5 ranged from 38 to 83 pixels with a mean of 54.1 ± 10.3 pixels. A significant correlation was found between the MPOD[FR] and MPOD[HFP] (r = 0.41, P = 0.02). CONCLUSIONS: This simplified method can provide accurate and reliable values of the MPOD comparable to heterochromatic flicker photometry. Obtaining the fundus images in this fast and easy way should be suitable for children thus enabling clinicians to determine the MPODs for children.