論文 - 蛭田　勇樹

Current advances in separation chemistry for antibody purification and analysis

Deol S., Matsuda Y., Hiruta Y.

Analytical Sciences 41 （ 5 ） 653 - 666 2025年05月

研究論文（学術雑誌）, 共著, 最終著者, 責任著者, 査読有り,  ISSN  09106340

　概要を見る

Monoclonal antibodies (mAbs) have become essential in modern therapeutics, offering high specificity and efficacy in treating diseases such as cancer, autoimmune disorders, and infectious diseases. With the increasing demand for mAbs, robust analytical and purification technologies are critical to ensuring their safety, efficacy, and consistency. This review highlights recent advances in the application of HPLC for both mAb analysis and purification. In the analytical domain, techniques such as size exclusion chromatography (SEC), hydrophobic interaction chromatography (HIC), ion exchange chromatography (IEX), and reversed-phase chromatography (RPLC) are explored for their role in evaluating mAb structural attributes and post-translational modifications (PTMs). These methods are indispensable for ensuring stability and functionality while meeting stringent regulatory requirements. In the context of purification, Protein A affinity chromatography remains the gold standard for initial capture of mAb; however, challenges such as high cost and harsh elution conditions have prompted the development of alternative purification technologies. IEX, HIC, multimodal, and membrane chromatography are also critical in achieving high-purity mAb products through multistep workflows, including intermediate purification and polishing stages. This review emphasizes the central role of HPLC in addressing the complex challenges of mAb manufacturing and characterization. By integrating established and emerging chromatographic techniques, it provides insights into the future directions of therapeutic antibody development.

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Bioluminescence readout lateral flow immunoassay using nanobody targeting aflatoxin B1

Takahashi, S; Hiruta, Y; Citterio, D

ANALYST 150 （ 8 ） 1563 - 1570 2025年04月

研究論文（学術雑誌）, 共著, 査読有り,  ISSN  0003-2654

　概要を見る

Multiple signal detection methods are known for lateral flow immunoassays (LFIAs), with colorimetric approaches dominating the field. However, their limited sensitivity is a remaining challenge. Fluorescence-based signaling is regarded as a more sensitive method, but it comes at the cost of partial sacrifice of the user-friendliness of LFIAs due to the requirement of an excitation light source. In this context, bioluminescence providing an inherently high signal to noise ratio without the need of excitation light could be an attractive alternative. But only a few studies have demonstrated the application of bioluminescence signaling in LFIAs. This work aimed at the development of a simple bioluminescence-based LFIA for the detection of aflatoxin B1 (AFB1), used as a model target in a competitive LFIA format. Signal transduction was achieved by nanobody-nanoluciferase (Nluc) fusion proteins. These small-sized recombinant heavy-chain-only antibodies derived from the camelidae family directly linked with the Nluc enzyme produce high intensity glow-type bioluminescence in combination with the furimazine substrate. LFIA devices consisting of a sample pad, nitrocellulose membrane and absorbent pad with AFB1-BSA conjugate deposited at the test line on the nitrocellulose membrane, achieved an LOD of 0.26 ng mL⁻¹ for aqueous AFB1 solutions pre-mixed with Nanobody-Nluc and bioluminescence emission observed on an imaging system. More user-friendly LFIA devices with integrated conjugate pad and pre-deposited Nanobody-Nluc provided clear AFB1 concentration-dependent bioluminescence signals with low background and enabled readout with a standard digital camera, resulting in an LOD of 1.12 ng mL⁻¹. Finally, the LFIA strips have been applied in AFB1-spiked oat milk samples. The LOD of 4.09 ng mL⁻¹ achieved in the real sample matrix is well below the maximum allowable residual concentration of AFB1 in the U.S. (20 ng mL⁻¹).

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Origami Paper-Based Immunoassay Device with CRISPR/Cas12a Signal Amplification

Suzuki, H; Tong, GD; Nath, P; Hiruta, Y; Citterio, D

ACS SENSORS 10 （ 3 ） 1811 - 1821 2025年03月

研究論文（学術雑誌）, 共著, 査読有り,  ISSN  2379-3694

　概要を見る

In clinical diagnosis, the determination of target proteins at low concentration levels is generally performed by immunoassays, such as the enzyme-linked immunosorbent assay (ELISA), which is a time-consuming process. To date, paper-based ELISA platforms enabling faster and less expensive analysis have been developed, but their important issue for clinical applications is the limited sensitivity compared to conventional ELISA. To address this challenge, this paper introduces a simple, rapid, and highly sensitive detection method for non-nucleic acid targets achieved by integrating the CRISPR/Cas12a system into paper-based ELISA. An origami-type paper-based device enabling simple assay operation has been designed, and the detection of targets on the paper substrates is based on observing the fluorescence signal induced by the CRISPR/Cas12a enzyme cleaving a probe single-stranded DNA (ssDNA) labeled with fluorophore and quencher (FQ reporter). To enhance sensitivity, antibodies labeled with a network of multiple DNA activating the CRISPR/Cas12a enzyme have been utilized as detection antibodies. As a result, the developed device successfully boosted the detection sensitivity for both human IgG and the hepatitis B virus surface antigen (HBsAg). In particular, the limit of detection (LOD) for HBsAg was estimated to be 12 pg/mL, representing over 10-fold higher sensitivity compared with commercially available HBsAg ELISA kits (LOD: 200 pg/mL). In addition, the fluorescence response toward porcine whole blood samples containing different HBsAg concentrations was also confirmed by capturing images with a smartphone, followed by quantitative data analysis. These results demonstrate the potential applicability of the proposed platform for clinical tests at the point of care.

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The latest developments of near-infrared fluorescent probes from NIR-I to NIR-II for bioimaging

Kuronuma, Y; Watanabe, R; Hiruta, Y

ANALYTICAL SCIENCES 41 （ 6 ） 737 - 757 2025年02月

研究論文（学術雑誌）, 共著, 最終著者, 責任著者, 査読有り,  ISSN  0910-6340

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Abstract: Near-infrared I (NIR-I: 650–950 nm) fluorescence imaging is a powerful tool for deep-tissue biological imaging, addressing the limitations of photon penetration depth in the visible-light region. Over the past decade, NIR imaging has extended to the near-infrared II (NIR-II: 1000–1700 nm) region, offering high-resolution and low background imaging by suppressing light scattering, and autofluorescence of tissues. Near-infrared fluorescent probes from NIR-I to NIR-II, with diverse functionalities, are increasingly utilized across biological fields to meet various detection needs and to explore physiological events in real time and spatial dimensions. This review discusses recent advancements in small-molecule NIR fluorescent dyes and probes, particularly those based on cyanine and rhodamine scaffolds, highlighting examples of their applications in bioimaging.

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Ratiometric Imaging for Quantification of Elevated Ca²⁺ in Neurons Using Synthetic Low-Affinity Fluorescent Probe

Kuronuma Y., Shindo Y., Kumada R., Sakama A., Citterio D., Oka K., Hiruta Y.

ACS Chemical Neuroscience 16 （ 4 ） 649 - 658 2025年02月

研究論文（学術雑誌）, 共著, 最終著者, 責任著者, 査読有り,  ISSN  1948-7193

　概要を見る

The availability of various calcium ion (Ca2+) fluorescent probes has contributed to revealing physiological events related to intracellular Ca2+. However, conventional probes face challenges for quantitatively and selectively visualizing high Ca2+ concentrations in cells induced by any stimuli, including biomolecules or electrical signal that disrupt Ca2+ homeostasis. In this report, we designed and synthesized a low-affinity ratiometric Ca2+ probe, KLCA-Fura, utilizing o-aminophenol-N,N-diacetate-O-methylene-methylphosphinate (APDAP) as a ligand, for which we recently demonstrated the suitability as a new low-affinity ligand for Ca2+. KLCA-Fura showed a blue shift in excitation wavelength with increasing Ca2+ concentration based on the intramolecular charge transfer (ICT). Its affinity for Ca2+ is lower than commercially available conventional Ca2+ probes. Furthermore, the selectivity for Ca2+ and the fluorescence intensity were considered sufficient to accurately detect Ca2+. The corresponding acetoxymethyl ester, KLCA-FuraAM, was synthesized for intracellular imaging and applied to Ca2+ quantification in neurons. KLCA-FuraAM enabled quantitative ratiometric monitoring of the two-step Ca2+ concentration increase induced by glutamate stimulation. While this two-step response was not clearly observed with a commercially available low-affinity ratiometric Ca2+ probe, Fura-FF, KLCA-FuraAM has demonstrated the potential to quantitatively visualize the behavior of high Ca2+ concentrations. The ratiometric low-affinity Ca2+ probe, KLCA-Fura, is expected to be a powerful tool for discovering new functions of Ca2+ in neurons.

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Creating coveted bioluminescence colors for simultaneous multi-color bioimaging

Hattori, M; Wazawa, T; Orioka, M; Hiruta, Y; Nagai, T

Science Advances 11 （ 4 ） eadp4750 2025年01月

研究論文（学術雑誌）, 共著, 査読有り,  ISSN  2375-2548

　概要を見る

Bioluminescence, an optical marker that does not require excitation by light, allows researchers to simultaneously observe multiple targets, each exhibiting a different color. Notably, the colors of the bioluminescent proteins must sufficiently vary to enable simultaneous detection. Here, we aimed to introduce a method that can be used to expand the color variation by tuning dual-acceptor bioluminescence resonance energy transfer. Using this approach, we could visualize multiple targets with up to 20 colors through single-shot acquisition using a color complementary metal-oxide semiconductor camera. Overall, this method enables simple and simultaneous observation of multiple biological targets and phenomena.

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3D-printed devices for multiplexed semi-quantitative competitive lateral flow immunoassays

Tong, GD; Misawa, K; Jarujamrus, P; Hiruta, Y; Citterio, D

ANALYTICAL METHODS 17 （ 6 ） 1206 - 1213 2025年01月

研究論文（学術雑誌）, 共著, 査読有り,  ISSN  1759-9660

　概要を見る

Lateral flow immunoassays (LFIAs) are widely used for the simple and rapid detection of various targets at the point of need. However, LFIAs enabling the simultaneous detection of multiple analytes and the possibility for naked-eye semi-quantitative analysis are facing various challenges, including the requirement of large sample volumes, low efficiency, and accuracy. This is particularly the case for the competitive immunoassay format targeting the detection of low molecular weight compounds, such as, for example, drugs. Due to limited space for multiple reaction zones on a single planar nitrocellulose membrane, conducting multiplexed tests requires the addition of more test strips, which consequently increases the size of the whole device. To overcome these spatial constraints, two 3D-printed devices fitting eight assay lanes of both backed and unbacked nitrocellulose membranes have been designed. For proof of concept, 8-OHdG, caffeine, and acetaminophen were used as model analytes. Inkjet printing was applied to deposit capture reagents in the form of text symbols while controlling the concentration thresholds for text readability to achieve an intuitive result expression. A comparably small sample volume of 350 mL was sufficient to simultaneously visually distinguish 4 concentration levels of caffeine (0, 4, 10, 175 ng mL-1) and acetaminophen (0, 4, 8, 12 ng mL-1) in mixed solutions without crosstalk. This study demonstrates the potential of 3D-printed LFIA devices for multiplex and semiquantitative analyte detection.

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Development of a Semiquantitative Barcode Readout Approach for Paper-Based Analytical Devices (PADs) for Enzymatic H₂O₂ and Glucose Detection

Manmana, Y; Kinugasa, S; Hiruta, Y; Citterio, D

ANALYTICAL CHEMISTRY 97 （ 3 ） 1500 - 1506 2025年01月

研究論文（学術雑誌）, 共著, 査読有り,  ISSN  0003-2700

　概要を見る

The integration of barcode technology with smartphones on paper-based analytical devices (PADs) presents a promising approach to bridging manual detection with digital interpretation and data storage. However, previous studies of 1D barcode approaches have been limited to providing only a “yes/no” response for analyte detection. Herein, a method of using barcode readout for semiquantitative signal detection on PADs has been achieved through the integration of barcode technology with a distance-based measurement concept on PADs. To demonstrate the feasibility of this concept, a PAD fabrication strategy incorporating barcodes was explored, using the enzymatic reaction between horseradish peroxidase (HRP), 3,3′-diaminobenzidine (DAB), and H2O2 as a model system. The enzyme-catalyzed polymerization of DAB to polyDAB in the presence of hydrogen peroxide results in the appearance of color observable by the naked eye inside a paperfluidic channel, with the color-changed length depending on the H2O2 concentration. At the same time, the barcode pattern displayed as a result of this distance-based color evolution overlaid with a paper-based barcode layer can be read using a smartphone application. Parameters affecting the signal readout performance were studied. The developed device can be used to detect H2O2 concentrations in the range of 0.25 to 10 mM within 90 min with 79.6% of barcode signals correctly readable. Additionally, results from different smartphone models showed a consistent reading performance (78.4-79.6%). Finally, the quantification of glucose levels in artificial urine samples was demonstrated. This developed PAD signaling strategy offers end-users more simplicity and can be used as a standalone device or in conjunction with other digital devices.

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Amplification-free CRISPR/Cas based dual-enzymatic colorimetric nucleic acid biosensing device

Tong, GD; Nath, P; Hiruta, Y; Citterio, D

Lab on a Chip 25 （ 4 ） 536 - 545 2025年01月

研究論文（学術雑誌）, 共著, 査読有り,  ISSN  1473-0197

　概要を見る

Nucleic acid testing (NAT) is widely considered the gold standard in analytical fields, with applications spanning environmental monitoring, forensic science and clinical diagnostics, among others. However, its widespread use is often constrained by complicated assay procedures, the need for specialized equipment, and the complexity of reagent handling. In this study, we demonstrate a fully integrated 3D-printed biosensensing device employing a CRISPR/Cas12a-based dual-enzymatic mechanism for highly sensitive and user-friendly nucleic acid detection. A plastic probe stick was designed to host small-sized gold nanoparticles, enhancing enzyme labeling density. Alkaline phosphatase (ALP) was then conjugated via single-stranded DNA, requiring only a single enzyme substrate addition to generate a simple visual signal change. This approach eliminates the need for amplification or centrifugation steps, achieving a limit of detection (LOD) as low as 10 pM – among the highest sensitivities reported for amplification-free colorimetric nucleic acid detection. Furthermore, we developed a device that incorporates this probe stick, integrates all necessary reagents, and features a smartphone-compatible accessory for quantitative analysis. This allows end-users to perform visual or quantitative DNA analysis with simple operations, achieving a visual detection limit of approximately 100 pM, comparable to other CRISPR-based non-amplified nucleic acid detection methods. Additionally, the system successfully distinguished perfectly matched from mismatched nucleic acid sequences, demonstrating its specificity and versatility. Although certain design limitations affected the sensitivity of the integrated device compared to the probe stick alone, the simplicity and portability of this device make it a promising tool for rapid nucleic acid screening in clinical diagnostics, environmental monitoring, and food safety control. This study paves the way for the development of practical biosensors for point-of-care testing (POCT) applications.

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Development of Column Packing Materials Using Functional Polymers and Application to Pharmaceutical Analysis

Hiruta, Y

CHROMATOGRAPHY 46 （ 1 ） 11 - 18 2025年

研究論文（学術雑誌）, 単著, 筆頭著者, 最終著者, 責任著者, 査読有り,  ISSN  1342-8284

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IgG-Binding Peptidomimetic Mixed-Charge Polymer-Modified Resins for Chromatographic Purification of Antibodies

Deura, K; Sakama, A; Moriwaki, Y; Citterio, D; Hiruta, Y

ACS APPLIED MATERIALS & INTERFACES 16 （ 49 ） 67468 - 67476 2024年11月

研究論文（学術雑誌）, 共著, 最終著者, 責任著者, 査読有り,  ISSN  1944-8244

　概要を見る

The process of antibody purification using Fc affinity ligands such as protein A, G, and L faces several challenges including high cost, low stability, and loss of antibody activity due to harsh elution conditions. Here, we describe a chromatographic purification of antibodies utilizing a pH-responsive mixed-charge polymer that mimics the IgG-binding peptide (Z34C) derived from the B domain of protein A. The protein A mimetic resins were prepared by modifying the surface of a TOYOPEARL, methacrylate resin with a polymer that mimics the amino acid sequence of Z34C and the functions of histidine and acidic and neutral amino acids using histamine methacrylamide (HisMA), methacrylic acid, and neutral monomers. The therapeutic monoclonal antibody (mAb), rituximab, was retained on the column at pH 7 and eluted under mildly acidic conditions at pH 5 using a protein A mimetic resin (HisMA20-EEMA) optimized for antibody interaction. The injected antibodies were selectively captured on the column by hydrophobic and electrostatic interactions with the protein A mimetic polymer under neutral conditions and eluted by electrostatic repulsion under acidic conditions. The HisMA20-EEMA column successfully purified mAbs from mixtures with BSA, mouse ascites fluid, and hybridoma cell culture supernatant. In addition, the HisMA20-EEMA column consistently achieved 90% antibody recovery in 100 consecutive purifications from cell culture supernatant. The antibody purification method presented in this study is low cost, highly durable, easy to synthesize, and allows for mild elution conditions. The results demonstrate that the approach of mimicking IgG-binding peptides with mixed-charge polymers is useful for the development of column packing materials for antibody purification.

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Sensor Arrays for Electrochemical Detection of PCR-Amplified Genes Extracted from Cells Suspended in Environmental Waters

Aoki, H; Kawaguchi, M; Kumakura, Y; Kamo, H; Miura, K; Hiruta, Y; Simizu, S; Citterio, D

SENSORS 24 （ 22 ）  2024年11月

研究論文（学術雑誌）, 共著, 査読有り

　概要を見る

Ecological surveys of living things based on DNAs from environmental samples are attractive. However, despite simplicity of water sampling from the target environment, it is still necessary to transport the samples to the laboratory for DNA analysis based on skillful next-generation sequencers. To perform DNA-oriented surveys based on a simple protocol without any special training, we demonstrated, in this study, the detection of genes from cell-containing environmental waters using gene sensor arrays that require no DNA labeling and no external indicators. Cell-suspended PBS or river water were used as models of environmental waters containing living things, and DNA samples were prepared by PCR amplification. Ferrocene-terminated probes were synthesized and immobilized on an electrode array to develop a sensor array. The sensor array showed a large response to a target DNA complementary to the probe and no response to a mismatched DNA, indicating sequence-specific detection. For DNA samples prepared from the cells in PBS, they showed good responses similar to those for the target DNA. They also significantly detected DNA samples from the cells in river water at a general environmental concentration (38 cells mL−1) with 28-fold larger responses than those for 0 cells mL−1.

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Paper-based analytical device for point-of-care nucleic acid quantification combining CRISPR/Cas12a and a personal glucose meter

Tanifuji, Y; Tong, GD; Hiruta, Y; Citterio, D

ANALYST 149 （ 19 ） 4932 - 4939 2024年09月

研究論文（学術雑誌）, 査読有り,  ISSN  0003-2654

　概要を見る

Although CRISPR-based nucleic acid detection has great potential in point-of-care testing due to its simplicity, it has been rarely integrated into paper-based analytical devices (PADs), which are attractive platforms to simplify assays. This work introduces a CRISPR-assisted nucleic acid quantification approach integrated into a PAD with signal readout by a personal glucose meter (PGM). Retention of magnetic beads by filter paper and pre-deposition of all required reagents by freeze-drying stabilized with trehalose enabled the indirect quantification of human papilloma virus (HPV) DNA through a PGM readout without complicated user intervention and complex reagent handling. The calculated limit of detection was 57 pM, which is comparable with other amplification-free CRISPR-based assays detecting nucleic acids. The fully integrated device exhibited good storage stability for up to 4 weeks, suggesting its applicability toward practical point-of-care nucleic acid quantification.

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Lateral flow immunoassay device to detect staphylococcal enterotoxin B (SEB) in durian candy

Relox, PE Jr; Israel, KAC; Hiruta, Y; Citterio, D

ANALYTICAL SCIENCES 41 （ 1 ） 3 - 11 2024年09月

研究論文（学術雑誌）, 査読有り,  ISSN  0910-6340

　概要を見る

Staphylococcal enterotoxin B (SEB), a potent enterotoxin produced by Staphylococcus aureus, has been implicated in incidences of Staphylococcal food poisoning in the Philippines. The use of lateral flow immunoassay devices to detect this toxin in solid food samples, like durian candy, at the point of sampling is constrained by the requirement for sample purification (e.g. centrifugation). This problem is also true with the other applications of LFIA devices on food samples. To overcome this challenge, a lateral flow immunoassay (LFIA) device capable of detecting SEB in unpurified durian candy sample was developed in this study. A modified LFIA device was assembled with three layers of glass fiber pads functioning as sample pads instead of a conventional cellulose fiber pad. Unlike with the cellulose fiber pad, the glass fiber sample pads acted as filter and allowed the flow of a 1:5 dilution of durian candy. The LFIA device applied to spiked 1:5 diluted durian candy samples achieved a visual limit of detection of 5 ng/mL for SEB, which is twofold lower than reported for previous LFIA devices designed to detect SEB in food samples.

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Rational design of pH-responsive near-infrared spirocyclic cyanines: the effects of substituents and the external environment

Sakama, A; Seo, H; Hara, J; Shindo, Y; Ikeda, Y; Oka, K; Citterio, D; Hiruta, Y

CHEMICAL COMMUNICATIONS 60 （ 46 ） 5984 - 5987 2024年06月

研究論文（学術雑誌）, 最終著者, 責任著者, 査読有り,  ISSN  1359-7345

　概要を見る

pH-responsive spirocyclic cyanine dyes were designed and synthesized. The equilibrium constant for cyclization (pKcycl) could be rationally controlled by changing the nucleophilic moiety and the side chains. Encapsulation in polymeric micelles inhibited the H-aggregation of the dye, and the pKcycl could be shifted according to the amphiphilic polymer employed.

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Basic evaluation of the CRISPR/Cas system stability for application to paper-based analytical devices

Tanifuji Y., Suzuki H., Tong G., Hiruta Y., Citterio D.

Analytical Methods 16 （ 25 ） 4143 - 4149 2024年05月

研究論文（学術雑誌）, 査読有り,  ISSN  17599660

　概要を見る

Despite the promising features of the CRISPR/Cas system for application to point-of-care nucleic acid tests, there are only a few reports on its integration into paper-based analytical devices (PADs) for the purpose of assay simplification. In most cases, paper platforms have only been used for the final signal readout in an assay otherwise performed in a test tube. Therefore, there is very limited information on the suitability of the CRISPR/Cas system for on-device reagent storage. To fill this gap, the current work primarily investigated the influence of various factors, including the type of paper, reagent drying method, effect of stabilizers, and storage condition on the storage stability of reagents necessary for CRISPR-based assays on paper substrates, by comparing the fluorescence signal emitted by the trans-cleavage of the dsDNA-activated Cas12a complex. The results obtained in the form of fluorescence signals emitted after trans-cleavage of a ssDNA probe through a dsDNA-activated Cas12a complex on paper substrates showed that CRISPR-related reagents spontaneously dried at room temperature on BSA blocked paper retained over 70% of their initial activity when stored at −20 °C for 28 days, independent of the type of paper substrates, which was improved by the addition of sucrose as a stabilizer. In addition, reagents dried on paper substrates under the optimized conditions exhibited stronger heat tolerance at temperatures above 65 °C compared to their corresponding solutions. This work is expected to contribute to the future development of fully integrated PADs relying on CRISPR/Cas systems for point-of-care applications requiring no additional reagent handling.

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Simplified capture, extraction, and amplification of cellular DNA from water samples

Kawaguchi M., Aoki H., Kamo H., Miura K., Hiruta Y., Simizu S., Citterio D.

Analytical Sciences （Analytical Sciences）  40 （ 3 ） 501 - 510 2024年03月

研究論文（学術雑誌）, 査読有り,  ISSN  09106340

　概要を見る

DNA analysis in water samples is attracting attention in various fields. However, conventional methods for DNA analysis require a work-intensive and time-consuming sample pre-treatment. In this study, a simplified pre-treatment method for analyzing DNA in water samples was evaluated. The process consists of filtration, DNA extraction, and amplification, which can be achieved within a short time. In the filtration process, two types of filters, firstly a tissue paper (Kimwipe) and then a glass filter (GF/F), were used in sequence. The first large pore size filter enabled a reduction in filtration time by removing large particulate matter impurities present in river water matrix. Cells spiked into 1 L of river water were recovered at more than 90% within approximately 5 min filtration time. Also, DNA was extracted from the captured cells directly on the surface of the filter in only 5 min. Thus, DNA collection and extraction from a water sample can be completed within about 10 min. Furthermore, PCR amplification was performed directly from DNA-attached filter sections, which greatly reduced the number of required pre-treatment steps. Finally, we succeeded in establishing a simple and fast on-site pre-treatment system by using a hand-driven syringe filtration method. This pre-treatment system is expected to offer the possibility for the future establishment of a rapid and easy DNA analysis method applicable to various types of water samples. Graphical abstract: (Figure presented.)

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Evaluation of separation performance for eggshell-based reversed-phase HPLC columns by controlling particle size and application in quantitative therapeutic drug monitoring (vol 15, pg 1790, 2023)

Yoshii, T; Nakano, K; Okuda, T; Citterio, D; Hiruta, Y

ANALYTICAL METHODS 16 （ 9 ） 1416 - 1416 2024年02月

研究論文（学術雑誌）, 最終著者, 責任著者,  ISSN  1759-9660

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Current advances in the development of bioluminescent probes toward spatiotemporal trans-scale imaging

Sakama A., Orioka M., Hiruta Y.

Biophysics and physicobiology 21 2024年

研究論文（学術雑誌）, 最終著者, 責任著者, 査読有り

　概要を見る

Bioluminescence imaging has recently attracted great attention as a highly sensitive and non-invasive analytical method. However, weak signal and low chemical stability of the luciferin are conventional drawbacks of bioluminescence imaging. In this review article, we describe the recent progress on the development and applications of bioluminescent probes for overcoming the aforementioned limitations, thereby enabling spatiotemporal trans-scale imaging. The detailed molecular design for manipulation of their luminescent properties and functions enabled a variety of applications, including in vivo deep tissue imaging, long-term imaging, and chemical sensor.

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Nonsequential and Sequential Dual Detection Probe Lateral Flow Immunoassay Device for Staphylococcal Enterotoxin B Detection

Relox, PE; Israel, KAC; Hiruta, Y; Citterio, D

MINDANAO JOURNAL OF SCIENCE AND TECHNOLOGY 22   235 - 259 2024年

ISSN  2244-0410

　概要を見る

Staphylococcal enterotoxins (SEs) are among the leading causes of foodborne illnesses in the Philippines. Rapid and sensitive detection of these toxins in food, particularly staphylococcal enterotoxin B (SEB) - the most potent type of SE - is imperative. Rapid toxin detection can be achieved using a lateral flow immunoassay (LFIA) device. However, low sensitivity has been a persistent issue with LFIA devices. To develop an LFIA device with improved sensitivity, this study investigated the exclusive use of antibodies as biorecognition molecules in a dual-detection probe LFIA device format and the configuration of the LFIA device to release the probes nonsequentially and sequentially. The LFIA device was fitted with two layers of glass fiber sample pads, loaded separately with the two detection probes, and partially separated by a Scotch tape film. For the nonsequential LFIA device, the two sample pads were pretreated with blocking buffers containing the same sugar concentration, whereas for the sequential LFIA device, the two sample pads were pretreated with blocking buffers containing different sugar concentrations. When tested against a filtered, homogenized durian candy sample, the nonsequential LFIA device generated higher signal intensity than the sequential LFIA device. This result suggests that aggregation of the detection probes was superior to sequential binding in enhancing the signal of the LFIA device for SEB detection. The sensitivity of the nonsequential LFIA device at 5 ng/mL was two-fold higher compared to previous POCT LFIA devices for SEB detection in food samples.

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