Enhancement of the Detection Performance of Paper-Based Analytical Devices by Nanomaterials

Paper-based analytical devices (PADs), including lateral flow assays (LFAs), dipstick assays and microfluidic PADs (μPADs), have a great impact on the healthcare realm and environmental monitoring. This is especially evident in developing countries because PADs-based point-of-care testing (POCT) enables to rapidly determine various (bio)chemical analytes in a miniaturized, cost-effective and user-friendly manner. Low sensitivity and poor specificity are the main bottlenecks associated with PADs, which limit the entry of PADs into the real-life applications. The application of nanomaterials in PADs is showing great improvement in their detection performance in terms of sensitivity, selectivity and accuracy since the nanomaterials have unique physicochemical properties. In this review, the research progress on the nanomaterial-based PADs is summarized by highlighting representative recent publications. We mainly focus on the detection principles, the sensing mechanisms of how they work and applications in disease diagnosis, environmental monitoring and food safety management. In addition, the limitations and challenges associated with the development of nanomaterial-based PADs are discussed, and further directions in this research field are proposed.     

Electrochemistry on Paper‐based Analytical Devices: A Review

Even though they were introduced less than a decade ago, electrochemical paper‐based devices (ePADs) have attracted widespread attention because of their inherent advantages in many applications. ePADs combine the advantages of microfluidic paper‐based devices (low cost, ease of use, equipment free pumping, etc.) for sample handling and processing with the advantages of sensitive and selective detection provided by electrochemistry. As a result, ePADs provide simplicity, portability, reproducibility, low cost and high selectivity and sensitivity for analytical measurements in a variety of applications ranging from clinical diagnostics to environmental sensing. Herein, recent advances in ePAD development and application are reviewed, focusing on electrode fabrication techniques and examples of applications specially focused on environmental monitoring, biological applications and clinical assays. Finally, a summary and prospective directions for ePAD research are also provided.     

普适型纸芯片的研究与应用

纸基分析芯片(纸芯片)具有成本低、便携化、操作和后处理简单无污染等优点,在临床诊断、食品质量控制和环境监测等领域有着广阔的应用前景。然而,由于难以制作性能优异的疏脂性屏障,使得纸芯片在涉及有机溶剂和表面活性剂的分析检测中,其发展受到了限制。针对当前纸芯片开发研究中存在的灵敏度较低、对有机溶剂和表面活性剂敏感等难点问题,研究人员在滤纸基底上制作出了性能优异的疏水隔离图案和高粘附疏水表面,并验证了所制备的纸芯片对涉及有机溶剂和表面活性剂的分析检测具有普适性。本文对此类普适型纸芯片的研究与应用进行评述。     

阵列纸芯片比色法检测碱性磷酸酶

采用5-溴-4-氯-3-吲哚磷酸盐(BCIP)/氯化硝基四氮唑蓝(NBT)显色体系,构建了阵列纸芯片比色检测碱性磷酸酶(ALP)的方法.首先,借助烘干处理方式在光刻法制备的阵列纸芯片微孔中固定显色试剂,然后加入ALP进行显色反应,最后,采用凝胶成像仪和普通照相机成像,读取显色强度(灰度值)进行比色检测.详细考察了显色条件对检测结果的影响,探讨了人血清白蛋白对ALP检测的增色效应,在最佳实验条件下,ALP检测的线性范围为1.5～20 U/L,检出限(3 σ)为0.78 U/L(n=18),比文献报道中纸芯片上检测ALP方法的检出限低约两个数量级.本方法成功用于实际血清样品检测,测定结果与临床值一致.在此基础上,构建了双色阵列纸芯片,通过颜色的变化实现了ALP的可视化半定量检测.     

基于等离子体技术制作微流控纸芯片及其在血糖检测中的应用研究

滤纸经十八烷基三氯硅烷(OTS)疏水化处理以后,用等离子体区域降解滤纸纤维表面的OTS疏水单分子层,使滤纸的局部区域恢复亲水性,得到具有亲疏水图案化的微流控纸芯片. 考察了等离子体处理时间对滤纸表面亲水性、亲水深度(水溶液由滤纸表层下渗至内部的纵向深度)的影响. 优化模具的设计,依据对滤纸亲水深度的不同需求,设计了两种PMMA-PDMS复合片的组合模具. 初步探讨了该亲疏水性变化过程的化学机理. 将制得的纸芯片用于人体全血中血糖含量的测定,线性范围为1.7～17.7 mmol·L^-1,可满足血液样品中血糖的测定.     

Smartphone-Assisted Protein to Creatinine Ratio Determination on a Single Paper-Based Analytical Device

Proteinuria is a condition in which an excessive amount of protein is excreted in urine. It is, among others, an indicator of kidney disease or risk of cardiovascular disease. Rapid and reliable diagnosis and monitoring of proteinuria is of great importance for both patients and their physicians. For that reason, a paper-based sensor for proteinuria diagnosis was designed, optimized, and validated utilizing smartphone-assisted signal acquisition. In the first step, a few commonly employed protein assays were optimized and compared in terms of analytical performance on paper matrix. The tetrabromophenol blue method was selected as the one providing a sufficiently low limit of detection (39 mg·L⁻¹) on the one hand and appropriate long-term stability (up to 3 months) on the other hand. The optimized assay was employed for protein-to-creatinine ratio (PCR) determination on a single paper-based sensor. For both analytes the linear ranges were within the clinically relevant range. The analytical usefulness of the developed sensors was demonstrated by a PCR recovery study in artificial urine. The obtained PCR recoveries were from ca. 80 to 150%.     

Utilizing Paper‐Based Devices for Antimicrobial‐Resistant Bacteria Detection

Antimicrobial resistance (AMR), the ability of a bacterial species to resist the action of an antimicrobial drug, has been on the rise due to the widespread use of antimicrobial agents. Per the World Health Organization, AMR has an estimated annual cost of USD 34 billion in the US and is predicted to be the number one cause of death worldwide by 2050. One way AMR bacteria can spread, and by which individuals can contract AMR infections, is through contaminated water. Monitoring AMR bacteria in the environment currently requires that samples be transported to a central laboratory for slow and labor intensive tests. We have developed an inexpensive assay using paper‐based analytical devices (PADs) that can test for the presence of β‐lactamase‐mediated resistance. To demonstrate viability, the PAD was used to detect β‐lactam resistance in wastewater and sewage and identified resistance in individual bacterial species isolated from environmental water sources.     

Paper‐Based Ratiometric Fluorescence Analytical Devices towards Point‐of‐Care Testing of Human Serum Albumin

Monitoring of human serum albumin (HSA) in a point‐of‐care fashion is urgently needed in particular for elderly or chronically ill patients. Herein, a dual‐state emissive chalcone probe having the feature of aggregation‐induced emission was designed and synthesized. The concentration of HSA can be evaluated by the ratios of emission from probes in aggregated and monomeric state, which gives a visually discernible red‐to‐green color change. A simple, portable paper‐based analytical device have been fabricated by integration of the recognition probe in the detection pad and employed for HSA test using the whole blood samples. This paper‐based assay shows the analytical capability comparable to the standard testing methods but is in a point‐of‐care fashion, providing a promising tool for at‐home HSA detection and HSA‐related disease diagnosis.     

L-cysteine modified gold nanoparticles / paper chip system for rapid detection of chiral dopaEI北大核心CSCD

A new detection method for L-Dopa based on paper chips was established. The L-Cys-AuNPs were characterized by their size, zeta potential, and UV-visible absorption spectra. The system had the high selectivity for the colorimetric detection of L-Dopa with the color changing from red to blue. The results were recorded using a common cell phone and subsequently analyzed using Photoshop software. A ratiometric color intensity method was designed for the quantification analysis. The ratio of color intensity at red channel and blue channel (R / B) increases linearly with L-Dopa concentration in the range of 5 to 80 μmol / L (R = 0. 9944), with the limit of detection of 4 μmol / L. The spiked recoveries of samples were 98%-102%. The RSDs of inter-day and intraday were 3. 3% and 3. 8%, respectively. Real samples were detected, and the error between the determination and the labeled value was within 5%. The method can be used to detect L-Dopa in real samples. © 2022, Youke Publishing Co.,Ltd. All rights reserved.     

Polymeric adsorbents with peptide pendants as artificial receptors for β-lactam antibiotics by mimicking the binding sites of β-lactamases

A series of polymeric adsorbents with peptide pendants were designed as the artificial receptors of β-lactam antibiotics by mimicking the structures of binding site in β-lactamases. Crosslinked poly(N, N-dimethyl acrylamide) gel as a carrier was prepared by suspension copolymerization of N, N-dimethyl acrylamide and N. Ń-bisacryl-diaminoethane and then functionalized with ethylenediamine after partial hydrolysis. Using solid-phase peptide synthesis with symmetrical anhydride of protected amino acid step by step, various peptide pendants were respectively anchored onto the functionalized carrier. The adsorption properties of these peptide-containing adsorbents for β-lactam antibiotics such as ampicillin and cefotaxime were then studied. The results showed that only those adsorbents in which peptide chains contained more than one lysine residues could obviously adsorb both β-lactams and that static interaction as well as hydrogen bond played an important role during the adsorption. Project supported by the National Natural Science Foundation of China (No. 59493207).     

AIM‐1: An Antibiotic‐Degrading Metallohydrolase That Displays Mechanistic Flexibility

Antibiotic resistance has emerged as a major threat to global health care. This is largely due to the fact that many pathogens have developed strategies to acquire resistance to antibiotics. Metallo‐β‐lactamases (MBL) have evolved to inactivate most of the commonly used β‐lactam antibiotics. AIM‐1 is one of only a few MBLs from the B3 subgroup that is encoded on a mobile genetic element in a major human pathogen. Here, its mechanism of action was characterised with a combination of spectroscopic and kinetic techniques and compared to that of other MBLs. Unlike other MBLs it appears that AIM‐1 has two avenues available for the turnover of the substrate nitrocefin, distinguished by the identity of the rate‐limiting step. This observation may be relevant with respect to inhibitor design for this group of enzymes as it demonstrates that at least some MBLs are very flexible in terms of interactions with substrates and possibly inhibitors.     

In Vitro Selection of a DNA Aptamer Targeting Degraded Protein Fragments for Biosensing

Protein biomarkers often exist as degradation fragments in biological samples, and affinity agents derived using a purified protein may not recognize them, limiting their value for clinical diagnosis. Herein, we present a method to overcome this issue, by selecting aptamers against a degraded form of the toxin B protein, which is a marker for diagnosing toxigenic Clostridium difficile infections. This approach has led to isolation of a DNA aptamer that recognizes degraded toxin B, fresh toxin B, and toxin B spiked into human stool samples. DNA aptamers selected using intact recombinant toxin B failed to recognize degraded toxin B, which is the form present in stored stool samples. Using this new aptamer, we produced a simple paper‐based analytical device for colorimetric detection of toxin B in stool samples, or in the NAP1 strain of Clostridium difficile. The combined aptamer‐selection and paper‐sensing strategy can expand the practical utility of DNA aptamers in clinical diagnosis.     

Designing Functionally Substituted Pyridine-Carbohydrazides for Potent Antibacterial and Devouring Antifungal Effect on Multidrug Resistant (MDR) Strains

The emergence of multidrug-resistant (MDR) pathogens and the gradual depletion of available antibiotics have exacerbated the need for novel antimicrobial agents with minimal toxicity. Herein, we report functionally substituted pyridine carbohydrazide with remarkable antimicrobial effect on multi-drug resistant strains. In the series, compound 6 had potent activity against four MDR strains of Candida spp., with minimum inhibitory concentration (MIC) values being in the range of 16–24 µg/mL and percentage inhibition up to 92.57%, which was exceptional when compared to broad-spectrum antifungal drug fluconazole (MIC = 20 µg/mL, 81.88% inhibition). Substitution of the octyl chain in 6 with a shorter butyl chain resulted in a significant anti-bacterial effect of 4 against Pseudomonas aeruginosa (ATCC 27853), the MIC value being 2-fold superior to the standard combination of ampicillin/cloxacillin. Time-kill kinetics assays were used to discern the efficacy and pharmacodynamics of the potent compounds. Further, hemolysis tests confirmed that both compounds had better safety profiles than the standard drugs. Besides, molecular docking simulations were used to further explore their mode of interaction with target proteins. Overall results suggest that these compounds have the potential to become promising antimicrobial drugs against MDR strains.     

A Modified Vancomycin Molecule Confers Potent Inhibitory Efficacy against Resistant Bacteria Mediated by Metallo-β-Lactamases

Multidrug-resistant bacterial infections mediated by metallo-β-lactamases (MβLs) have grown into an emergent health threat, and development of novel antimicrobials is an ideal strategy to combat the infections. Herein, a novel vancomycin derivative V_b was constructed by conjugation of triazolylthioacetamide and vancomycin molecules, characterized by reverse-phase high performance liquid chromatography (HPLC) and confirmed by matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS). The biological assays revealed that V_b effectively inhibited S. aureus and methicillin-resistant S. aureus (MRSA), gradually increased the antimicrobial effect of β-lactam antibiotics (cefazolin, meropenem and penicillin G) and exhibited a dose-dependent synergistic antibacterial effect against eight resistant strains tested, which was confirmed by the time-kill curves determination. Most importantly, V_b increased the antimicrobial effect of meropenem against the clinical isolates EC08 and EC10 and E. coli producing ImiS and CcrA, resulting in a 4- and 8-fold reduction in MIC values, respectively, at a dose up to 32 μg/mL. This work offers a promising scaffold for the development of MβLs inhibitors, specifically antimicrobials for clinically drug-resistant isolates.     

A Synergistic Capture Strategy for Enhanced Detection and Elimination of Bacteria

Despite the advanced detection and sterilization techniques available today, the sensitive diagnosis and complete elimination of bacterial infections remain a significant challenge. A strategy is reported for efficient bacterial capture (ca. 90 %) based on the synergistic effect of the nanotopography and surface chemistry of the substrate on bacterial attachment and adhesion. The outstanding bacterial‐capture capability of the functionalized nanostructured substrate enables rapid and highly sensitive bacterial detection down to trace concentrations of pathogenic bacteria (10 colony‐forming units mL^?1). In addition, this synergistic biocapture substrate can be used for efficient bacterial elimination and shows great potential for clinical antibacterial applications.     

Combating Multidrug‐Resistant Bacteria: Current Strategies for the Discovery of Novel Antibacterials

The introduction of effective antibacterial therapies for infectious diseases in the mid‐20th century completely revolutionized clinical practices and helped to facilitate the development of modern medicine. Many potentially life‐threatening conditions became easily curable, greatly reducing the incidence of death or disability resulting from bacterial infections. This overwhelming historical success makes it very difficult to imagine life without effective antibacterials; however, the inexorable rise of antibiotic resistance has made this a very real and disturbing possibility for some infections. The ruthless selection for resistant bacteria, coupled with insufficient investment in antibacterial research, has led to a steady decline in the efficacy of existing therapies and a paucity of novel structural classes with which to replace them, or complement their use. This situation has resulted in a very pressing need for the discovery of novel antibiotics and treatment strategies, the development of which is likely to be a key challenge to 21st century medicinal chemistry.     

A β‐Lactamase‐Imprinted Responsive Hydrogel for the Treatment of Antibiotic‐Resistant Bacteria

Antibiotics play important roles in infection treatment and prevention. However, the effectiveness of antibiotics is now threatened by the prevalence of drug‐resistant bacteria. Furthermore, antibiotic abuse and residues in the environment cause serious health issues. In this study, a stimuli‐responsive imprinted hydrogel was fabricated by using β‐lactamase produced by bacteria for deactivating antibiotics as the template molecule. The imprinted hydrogel could initially trap β‐lactamase excreted by drug‐resistant bacteria, thus making bacteria sensitive to antibiotics. After the bactericidal treatment, the “imprinted sites” on the hydrogel could be reversibly abolished with a temperature stimulus, which resulted in the reactivation of β‐lactamase to degrade antibiotic residues. We also present an example of the use of this antibacterial design to treat wound infection.     

A Self‐Immobilizing and Fluorogenic Probe for β‐Lactamase Detection

The spread of antibiotic resistance in pathogenic bacteria has become one of the major concerns to public health. Improved monitoring of drug resistance is of high importance for infectious disease control. One of the major mechanisms for bacteria to overcome treatment of antibiotics is the production of β‐lactamases, which are enzymes that hydrolyze the β‐lactam ring of the antibiotic. In this study, we have developed a self‐immobilizing and fluorogenic probe for the detection of β‐lactamase activity. This fluorogenic reagent, upon activation by β‐lactamases, turns on a fluorescence signal and, more importantly, generates a covalent linkage to the target enzymes or the nearby proteins. The covalent labeling of enzymes was confirmed by SDS‐PAGE analysis and MALDI‐TOF mass spectrometry. The utility of this structurally simple probe was further confirmed by the fluorescent labeling of a range of β‐lactamase‐expressing bacteria.