Analytical biochemistry最新文献_第2页

Development of a chemiluminescence enzyme immunoassay (CLEIA) for quantitating L1 protein in HPV vaccines

IF 2.6 4区生物学

Analytical biochemistry Pub Date : 2025-05-02 DOI: 10.1016/j.ab.2025.115889

ChengRui Fei , Huan Yang , SiJie Wang , WenZhi He , Xue Shen , Ying Zhang , YiHeng Jiang , Li Yang , XiaoJuan Li , Fan Wu , YaNan Wu , Qin Liu

{"title":"Development of a chemiluminescence enzyme immunoassay (CLEIA) for quantitating L1 protein in HPV vaccines","authors":"ChengRui Fei , Huan Yang , SiJie Wang , WenZhi He , Xue Shen , Ying Zhang , YiHeng Jiang , Li Yang , XiaoJuan Li , Fan Wu , YaNan Wu , Qin Liu","doi":"10.1016/j.ab.2025.115889","DOIUrl":"10.1016/j.ab.2025.115889","url":null,"abstract":"<div><div>The L1 protein serves as the principal capsid component of human papillomavirus (HPV). All globally commercialized HPV vaccines utilize virus-like particles (VLPs) formed through L1 protein self-assembly. Quantitative analysis of L1 protein concentration constitutes a critical parameter for evaluating the antigenic potency of HPV vaccines. In this study, we developed a chemiluminescent enzyme immunoassay (CLEIA) for precise quantification of L1 protein in bivalent HPV vaccines. Employing a sandwich immunoassay format, antigen-antibody complexes were immobilized on 96-well microplates using capture antibodies, followed by detection with HRP-conjugated secondary antibodies. Chemiluminescent signal amplification was achieved through enzymatic catalysis of luminol/hydrogen peroxide in the presence of enhancer molecules. Systematic optimization of experimental parameters yielded a validated methodology demonstrating excellent reproducibility (inter-assay CV < 4 %), accuracy (recovery rate 100.5 ± 2.8 % for 16L1 and 95.5 ± 6.4 % for 18L1), precision (inter-assay CV < 7 %) and sensitivity (limit of quantitation (LOQ) 0.0996 μg/mL for HPV16L1 and 0.1459 μg/mL for HPV18L1). This optimized assay provides a reliable analytical platform for quantitating L1 Protein in bivalent HPV vaccine.</div></div>","PeriodicalId":7830,"journal":{"name":"Analytical biochemistry","volume":"704 ","pages":"Article 115889"},"PeriodicalIF":2.6,"publicationDate":"2025-05-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143943313","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Triple DNAzyme cleavage mediated signal cascade for sensitive and reliable Kawasaki disease related microRNA analysis 三重DNAzyme切割介导的信号级联用于敏感可靠的川崎病相关microRNA分析

IF 2.6 4区生物学

Analytical biochemistry Pub Date : 2025-05-01 DOI: 10.1016/j.ab.2025.115887

Qunyan Ruan, Bina Zhao

引用次数: 0

TransCNN: A novel architecture combining transformer and TextCNN for detecting N4-acetylcytidine sites in human mRNA transnn：一种结合transformer和TextCNN的新型结构，用于检测人类mRNA中的n4 -乙酰胞苷位点

IF 2.6 4区生物学

Analytical biochemistry Pub Date : 2025-04-29 DOI: 10.1016/j.ab.2025.115882

Shengli Zhang, Kai Liu, Yujie Xu

{"title":"TransCNN: A novel architecture combining transformer and TextCNN for detecting N4-acetylcytidine sites in human mRNA","authors":"Shengli Zhang, Kai Liu, Yujie Xu","doi":"10.1016/j.ab.2025.115882","DOIUrl":"10.1016/j.ab.2025.115882","url":null,"abstract":"<div><div>N4-acetylcytidine (ac4C), a pivotal post-transcriptional RNA modification, is central to understanding transcriptional regulation and diverse biological processes. As a key determinant of RNA structural stability and functional regulation, ac4C has been strongly associated with multiple human diseases. We can obtain a better understanding of regulation mechanism of gene expression by identifying ac4C sites rapidly and precisely. However, existing predictive approaches are constrained by limitations in feature representation and sequence context modeling, necessitating the development of advanced methodologies. In this study, we introduce a novel architecture named TransCNN that integrates transformer and Text convolutional neural network (TextCNN) to predict ac4C sites. TransCNN demonstrates superior performance compared to existing models on both 10-fold cross-validation and independent dataset with the accuracy of 83.27 % and 82.89 %, respectively. The enhanced performance of TransCNN is attributed to the transformer's ability to extract adaptive features and TextCNN's capability to form both narrow and broad connections within the sequence. This study aims to contribute significantly to the field by advancing the understanding and prediction of RNA modifications. The datasets and code used in this study are available at <span><span>https://github.com/liukai23157/</span><svg><path></path></svg></span>TransCNN.</div></div>","PeriodicalId":7830,"journal":{"name":"Analytical biochemistry","volume":"703 ","pages":"Article 115882"},"PeriodicalIF":2.6,"publicationDate":"2025-04-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143898423","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Metabolomics approach using HR-MAS NMR spectroscopy for the assessment of metabolic profiles of uterine fibroids 代谢组学方法使用HR-MAS核磁共振光谱评估子宫肌瘤的代谢谱

IF 2.6 4区生物学

Analytical biochemistry Pub Date : 2025-04-29 DOI: 10.1016/j.ab.2025.115885

Senem Arda Düz , Akın Mumcu , Berat Doğan , Erdinç Sarıdoğan , Görkem Tuncay , Taylan Onat , Abdullah Karaer

{"title":"Metabolomics approach using HR-MAS NMR spectroscopy for the assessment of metabolic profiles of uterine fibroids","authors":"Senem Arda Düz , Akın Mumcu , Berat Doğan , Erdinç Sarıdoğan , Görkem Tuncay , Taylan Onat , Abdullah Karaer","doi":"10.1016/j.ab.2025.115885","DOIUrl":"10.1016/j.ab.2025.115885","url":null,"abstract":"<div><div>The aim of this study is to determine dysregulated metabolites and metabolic pathways in uterine fibroids and in the myometrial tissue from which uterine fibroids are derived. Fifteen (15) patients underwent hysterectomy because of uterine fibroids and 14 controls were included in this study. <sup>1</sup>H HR-MAS NMR spectroscopy data were obtained from uterine fibroid tissue, the adjacent healthy myometrial tissue from cases, and myometrial tissue from controls. PCA and PLS-DA score plots from multivariate statistical analysis of pre-processed spectral data demonstrated a distinction between cases and control groups. The levels of lactate, alanine, glutamate, glutamine, methionine, acetone, isocitrate, choline, glycerophosphocholine, phosphocholine, <em>o</em>-phosphoethanolamine, taurine, myo-inositol, <em>p</em>-methylhistidine, phenylacetate, ascorbate, glucose, and methylhistidine were significantly higher in uterine fibroid tissue compared to the neighboring healthy myometrial tissue. Additionally, when adjacent healthy myometrial tissue was compared to control myometrial tissue, significantly lower levels of valine, leucine, isoleucine, ethanol, arginine, N-acetyl tyrosine, acetone, <em>p</em>-methylhistidine, glucose, phenylacetate, myo-inositol, and alpha-glucose were observed. The study provides a foundational framework by revealing the metabolomic heterogeneity of uterine fibroids. Strategies should be developed to target the metabolic alterations that contribute to the growth of these common tumors.</div></div>","PeriodicalId":7830,"journal":{"name":"Analytical biochemistry","volume":"704 ","pages":"Article 115885"},"PeriodicalIF":2.6,"publicationDate":"2025-04-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143935883","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Proximity rolling circle amplification based generation of lighting-up aptamer for sensitive and label-free MicroRNA analysis 基于邻近滚动圆扩增的点亮适体的产生，用于敏感和无标记的MicroRNA分析

IF 2.6 4区生物学

Analytical biochemistry Pub Date : 2025-04-25 DOI: 10.1016/j.ab.2025.115884

Chaogang Yuan , Lifang Cao , Honglian Shen , Yuexiang Zhang

{"title":"Proximity rolling circle amplification based generation of lighting-up aptamer for sensitive and label-free MicroRNA analysis","authors":"Chaogang Yuan , Lifang Cao , Honglian Shen , Yuexiang Zhang","doi":"10.1016/j.ab.2025.115884","DOIUrl":"10.1016/j.ab.2025.115884","url":null,"abstract":"<div><div>The dysregulated expression of microRNA (miRNA) has been strongly linked to pneumonia. However, ultrasensitive miRNA detection remains technically challenging due to their short sequences and low abundance in biological samples. Herein, we developed a catalytic hairpin assembly -triggered proximity rolling circle transcription system for synthesizing malachite green (MG)-binding RNA aptamers, enabling label-free and highly sensitive detection of miRNA-21. In this system, target miRNA-21 initiates hairpin probe structural rearrangement, promoting miRNA-21 recycling and activating dual rolling circle transcription reactions. The resulting long RNA transcripts undergo partial hybridization, yielding numerous functional MG RNA aptamers. Upon binding to MG dye, these aptamers generate a robust fluorescence signal, allowing ultrasensitive miRNA-21 detection at concentrations as low as 0.51 fM. The assay exhibits high specificity, discriminating miRNA-21 from single-base mismatched sequences, and shows promise for monitoring miRNA-21 expression in clinical samples. By combining the signal amplification of rolling circle transcription with the high-affinity recognition of MG aptamers, this method achieves a low background, superior signal-to-noise ratio, and exceptional sensitivity. Furthermore, the strategy can be readily adapted to detect other trace biomarkers by modifying the target-recognition sequence.</div></div>","PeriodicalId":7830,"journal":{"name":"Analytical biochemistry","volume":"703 ","pages":"Article 115884"},"PeriodicalIF":2.6,"publicationDate":"2025-04-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143894737","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

nCas9-based method for rolling-circle DNA substrate generation 基于ncas9的滚圈DNA底物生成方法

IF 2.6 4区生物学

Analytical biochemistry Pub Date : 2025-04-25 DOI: 10.1016/j.ab.2025.115883

Nischal Sharma, Kelsey S. Whinn, Harshad Ghodke, Antoine M. van Oijen, Jacob S. Lewis, Lisanne M. Spenkelink

{"title":"nCas9-based method for rolling-circle DNA substrate generation","authors":"Nischal Sharma, Kelsey S. Whinn, Harshad Ghodke, Antoine M. van Oijen, Jacob S. Lewis, Lisanne M. Spenkelink","doi":"10.1016/j.ab.2025.115883","DOIUrl":"10.1016/j.ab.2025.115883","url":null,"abstract":"<div><div>Rolling-circle DNA replication is a DNA-duplication mechanism whereby circular DNA templates are continuously copied to produce long DNA products. It is widely used in molecular diagnostics, DNA sequencing, nanotechnology, and <em>in vitro</em> DNA replication studies. The efficiency of rolling-circle replication reaction heavily relies on the quality of the rolling-circle DNA template. Existing methods to create rolling-circle DNA substrates often rely on unique restriction sites and have limited control over replication fork topology and position. To address these limitations, we present a straightforward, customizable, and efficient strategy for producing rolling-circle DNA substrates with control over gap size and fork position. Our method relies on the use of nickase Cas9 (nCas9), which can be programmed to target specific DNA sequences using guide RNAs. In a one-pot reaction, we target nCas9 to four sites on an 18-kb plasmid to create 8–11-bp fragments. These fragments are removed and a flap oligo is ligated, to construct a fork with precisely controlled flap length and gap size. We demonstrate the application of this DNA substrate in an <em>in vitro</em> single-molecule rolling-circle DNA-replication assay. With our method, any plasmid DNA can be converted into a rolling-circle template, permitting generation of more physiologically-relevant DNA templates.</div></div>","PeriodicalId":7830,"journal":{"name":"Analytical biochemistry","volume":"703 ","pages":"Article 115883"},"PeriodicalIF":2.6,"publicationDate":"2025-04-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143881540","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Development of a competitive ELISA based on Brucella neotomae lipopolysaccharide for detecting brucellosis in livestock 基于新生布鲁氏菌脂多糖的竞争性酶联免疫吸附测定家畜布鲁氏菌病

IF 2.6 4区生物学

Analytical biochemistry Pub Date : 2025-04-23 DOI: 10.1016/j.ab.2025.115880

Guo-Hua Cai, Chao-Yue Guo, Kai-Xuan Guo, Jian-Dong Zhang, Huan-Chun Chen, Zheng-Fei Liu

{"title":"Development of a competitive ELISA based on Brucella neotomae lipopolysaccharide for detecting brucellosis in livestock","authors":"Guo-Hua Cai, Chao-Yue Guo, Kai-Xuan Guo, Jian-Dong Zhang, Huan-Chun Chen, Zheng-Fei Liu","doi":"10.1016/j.ab.2025.115880","DOIUrl":"10.1016/j.ab.2025.115880","url":null,"abstract":"<div><div>Brucellosis, a global zoonotic threat, requires efficient diagnostic tools for effective surveillance. Commercial competitive enzyme-linked immunosorbent assay (cELISA) predominantly utilizes smooth lipopolysaccharides (S-LPS) extracted from <em>B. abortus</em> and <em>B. melitensis</em> as key antigens for brucellosis serodiagnosis. However, culturing pathogens requires facilities with high biosafety, which is operationally complex and economically demanding. In this study, we developed a cELISA using LPS extracted from <em>B. neotomae</em>, which can be handled more facilely in biosafety level 2 conditions, and analyzed clinical adaptability of the cELISA. The optimized cELISA demonstrated lower detection limits, which was 2–4 times more analytically sensitive than commercial kit by detecting sera against <em>B. melitensis</em> and <em>B. abortus</em>. No cross-reactivity was observed with sera infected with other bacteria, including <em>E. coli</em>, <em>Salmonella</em>, <em>Y. enterocolitica</em>, and <em>M. tuberculosis</em>. The diagnostic sensitivity and specificity of the cELISA were 100 % (40/40) and 100 % (40/40), respectively. The coefficients of variation were less than 10 %. Moreover, compared to the commercial kit, the developed ELISA achieved agreement of 92.51 % across 427 sera from vaccinated livestock, and agreement of 96.98 % across 696 sera from non-vaccinated livestock. In conclusion, the cELISA exhibits excellent sensitivity, specificity and repeatability, indicating its potential for brucellosis diagnosis in livestock.</div></div>","PeriodicalId":7830,"journal":{"name":"Analytical biochemistry","volume":"703 ","pages":"Article 115880"},"PeriodicalIF":2.6,"publicationDate":"2025-04-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143877194","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Real-time monitoring method of microbial growth using a simple pressure-based respiration detection system 利用简单的压力呼吸检测系统实时监测微生物生长的方法

IF 2.6 4区生物学

Analytical biochemistry Pub Date : 2025-04-22 DOI: 10.1016/j.ab.2025.115879

Nara Shin , Jinok Oh , Yebin Han , Gaeun Lim , Jeong Chan Joo , Woo-Young Jeon , Jungoh Ahn , Hee Taek Kim , Shashi Kant Bhatia , Yung-Hun Yang

{"title":"Real-time monitoring method of microbial growth using a simple pressure-based respiration detection system","authors":"Nara Shin , Jinok Oh , Yebin Han , Gaeun Lim , Jeong Chan Joo , Woo-Young Jeon , Jungoh Ahn , Hee Taek Kim , Shashi Kant Bhatia , Yung-Hun Yang","doi":"10.1016/j.ab.2025.115879","DOIUrl":"10.1016/j.ab.2025.115879","url":null,"abstract":"<div><div>Dry cell weight (DCW) and optical density (OD) measurement methods provide useful data for assessing microbial growth. However, their sampling process is labor-intensive and time-consuming. Therefore, we aimed to evaluate a method for measuring microbial growth through continuous CO<sub>2</sub> measurement under aerobic conditions using a pressure-based respiration detection system, which is traditionally used in anaerobic environments and applies measurement of reduced pressure by capturing CO<sub>2</sub> with KOH. The pressure reduction rate, OD, and DCW values were compared during <em>Ralstonia eutropha</em> H16 culture, which revealed a correlation of R<sup>2</sup> of 0.99 between the pressure reduction and DCW and a change of DCW (g/L) per pressure (1 mbar) of −0.02 g/L. It showed theoretical limit of detection at 14.67 mbar corresponding to 0.0428 g/L of DCW and theoretical limit of quantification at 48.9 mbar as lower limits. When the pressure-based method was applied to compare carbon source utilization and growth of different strains, such as <em>E. coli</em> sp., <em>Pseudomonas</em> sp., <em>Burkholderia</em> sp., and <em>Bacillus</em> sp., it showed a high correlation with DCW. Overall, these results demonstrate that the pressure-based respiration detection system is a reliable tool for microbial growth monitoring and offers significant advantages by providing real-time data with less labor.</div></div>","PeriodicalId":7830,"journal":{"name":"Analytical biochemistry","volume":"703 ","pages":"Article 115879"},"PeriodicalIF":2.6,"publicationDate":"2025-04-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143869448","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Development of a high-sensitivity double kinetic assay for creatinine using the enzymatic cycling method and its application to serum samples 酶循环法测定肌酐高灵敏度双动力学分析方法的建立及其在血清样品中的应用

IF 2.6 4区生物学

Analytical biochemistry Pub Date : 2025-04-21 DOI: 10.1016/j.ab.2025.115877

Eisaku Hokazono , Yasunori Haraguchi , Ai Higashinakao, Akito Mominoki, Takeshi Uchiumi, Yuzo Kayamori

{"title":"Development of a high-sensitivity double kinetic assay for creatinine using the enzymatic cycling method and its application to serum samples","authors":"Eisaku Hokazono , Yasunori Haraguchi , Ai Higashinakao, Akito Mominoki, Takeshi Uchiumi, Yuzo Kayamori","doi":"10.1016/j.ab.2025.115877","DOIUrl":"10.1016/j.ab.2025.115877","url":null,"abstract":"<div><div>Estimated glomerular filtration rate (eGFR) is often used as a measure of renal function in clinical practice owing to its simplicity. Serum creatinine levels are essential for eGFR calculation. Although the Jaffé method is widely used for creatinine measurement, it exhibits low specificity and sensitivity. Development of various enzyme methods has increased its specificity; however, its sensitivity is still insufficient for accurate eGFR calculation. To overcome this issue, we developed a highly sensitive assay for creatinine detection using enzymatic cycling reaction in this study. Our method consisted of two steps: Endogenous ammonia elimination and creatinine analysis. NADH derived from creatinine changed the water-soluble tetrazolium salt-8 color in the presence of the electron carrier, 1-methoxy-5-methylphenazinium methylsulfate. The cycling oxidation–reduction of 1-methoxy-5-methylphenazinium methylsulfate to NADH facilitated the coloration of water-soluble tetrazolium salt-8, aiding in creatinine measurement with high sensitivity. The within-run reproducibility of the developed method was good (<1.76 % at each concentration tested), with a detection limit of 1.00 μmol/L, making it approximately 9 times more sensitive than the Jaffé method. Notably, its correlation with the high-performance liquid chromatography method was excellent (<em>r</em> = 0.976). Overall, this study successfully developed a new, rapid, simple, and highly sensitive method for creatinine analysis.</div></div>","PeriodicalId":7830,"journal":{"name":"Analytical biochemistry","volume":"703 ","pages":"Article 115877"},"PeriodicalIF":2.6,"publicationDate":"2025-04-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143906673","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Highly-sensitive peptide array using peptides immobilized on microbeads: Application to cow's milk allergy analysis 微珠固定多肽的高敏感多肽阵列：在牛奶过敏分析中的应用

IF 2.6 4区生物学

Analytical biochemistry Pub Date : 2025-04-19 DOI: 10.1016/j.ab.2025.115865

Hideo Tashiro , Tomoko Tashiro , Fumiya Yamaide , Taiji Nakano , Yuzaburo Inoue , Yuki Takase , Erika Sawano , Naoki Shimojo

{"title":"Highly-sensitive peptide array using peptides immobilized on microbeads: Application to cow's milk allergy analysis","authors":"Hideo Tashiro , Tomoko Tashiro , Fumiya Yamaide , Taiji Nakano , Yuzaburo Inoue , Yuki Takase , Erika Sawano , Naoki Shimojo","doi":"10.1016/j.ab.2025.115865","DOIUrl":"10.1016/j.ab.2025.115865","url":null,"abstract":"<div><h3>Analysis</h3><div>with peptide microarrays containing linear epitopes of allergenic proteins is expected to provide information on the clinical status of the patient, but peptide arrays are still limited to research use. We thus aimed at developing a simple and sensitive peptide array that operates with more cost-effective ECL detection, so that it can be routinely used in clinical practice.</div><div>For this purpose, instead of directly immobilizing the peptides onto the microarray surface as in the previous reports, we developed a two-step immobilization technique using microbeads. Peptides biotinylated at the N-terminal are first bound to microbeads with streptavidin (sAV) on the surface, followed by immobilization of the peptide-bound beads onto the microarray substrate using the photoreactive crosslinker. In this way, we were able to overcome the limitations of direct immobilization in increasing the amount and accessibility of peptides and greatly enhance the sensitivity so that ECL detection became possible. In the present study, we analyzed sera from cow's milk allergy (CMA) patients with our peptide array containing 20 peptides from αS1-casein. The results showed that IgE epitope patterns of patients could be visualized individually, and confirmed that the pattern is unique to each patient.</div></div>","PeriodicalId":7830,"journal":{"name":"Analytical biochemistry","volume":"703 ","pages":"Article 115865"},"PeriodicalIF":2.6,"publicationDate":"2025-04-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143878629","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0