Detection Of Lipopolysaccharide Research Articles

NK-lysin antimicrobial peptide-functionalized nanoporous alumina membranes as biosensors for label-free bacterial endotoxin detection

We report an NK-lysin peptide-functionalized nanoporous anodized aluminum oxide (NAAO) based biosensor to detect bacterial endotoxin. Bovine NK-lysin-derived peptides show antimicrobial activity against bacterial pathogens, and bactericidal activity is primarily due to the membranolysis activity. Antimicrobial activity of NK-lysin NK2A was confirmed against a Gram-negative Mannheimia haemolytica and a Gram-positive Staphylococcus aureus. Electron microscopic examination showed the localization of NK2A conjugated silver nanoparticles, but not unconjugated silver nanoparticles used as control, to the bacterial outer membrane and cell wall. NK2A functionalized NAAO membranes were used in a previously developed four-electrode electrochemical configuration to detect the presence of Gram-negative bacterial lipopolysaccharides (LPS) and Gram-positive bacterial lipoteichoic acid (LTA) molecules. NK2A-functionalized NAAO biosensor could detect LPS with a detection limit of 10 ng/mL within an appreciable signal/noise ratio. Biosensors functionalized with a scrambled amino acid version of NK2A (Sc-NK2A) that lacks antimicrobial activity could not detect the presence of LPS. However, both NK2A and Sc-NK2A functionalized biosensors showed sensing signals with Gram-positive bacterial lipoteichoic acids. These results suggest that the specific binding of NK2A-LPS on the NAAO membrane surface is responsible for the observed biosensor signals. These findings suggest that NK2A-functionalized biosensors can be used for rapid and sensitive label-free LPS detection.

Ratiometric fluorescence probe integrated with smartphone for visually detecting lipopolysaccharide

A portable instrument-free detection method for lipopolysaccharide (LPS) analysis was developed based on dual-emission ratiometric fluorescence sensing system. Herein, red-emitting Au nanoclusters (Au NCs) were as reference probe, while blue-emitting fluorescent silica quantum dots (Si QDs) were as response probe. Additionally, the aptamer of LPS was covalently grafted to the surface of Si QDs in order to specific recognize the LPS. According to the changes of fluorescence intensityratio (FL ratio, I461 nm/I643 nm) with the concentrations of LPS, the linear equation was fitted with the range of 50–3000 ng/mL, and the limit of detection (LOD) was 29.3 ng/mL. As a practical application, this method was employed to analyze LPS in normal saline with the recovery rate of 97.7–103.8 %. The color picker platform in the smartphone was used to transform the detection picture to the process of Red, Green and Blue (RGB) for visual detection of LPS. The low-cost and easy-carry method reported here presents broad merits for the visually quantitative detection of LPS.

Cu2+-Mediated Aggregation of Gold Nanoparticles as an Optical Probe for the Detection of Endotoxin.

Endotoxins or lipopolysaccharides (LPS) present in the outer layer of Gram-negative bacteria (GNB) are responsible for bacterial toxicity. It is an environmental hazard that everyone is exposed to daily to various extents. Due to its potent toxicity, quantitative detection with very high sensitivity is essential in the food, medical, and pharmaceutical industries. Herein, we report an optical nanosensor for the rapid and sensitive detection of LPS and GNB based on the Cu2+-mediated aggregation of gold nanoparticles (Cu@AuNPs). The sensor detects LPS within a linear range of 20 ag/mL to 20 ng/mL with a lower detection limit of 0.2 ag/mL. The sensor could successfully recover spiked endotoxin in grape juice with a percentage error of ±0.2, confirming its application in the food industry. The sensor could also distinguish Gram-negative bacteria from Gram-positive bacteria, and the selectivity of the Cu@AuNP sensor toward GNB is utilized to detect Escherichia coli in wastewater. The rapid detection of E. coli without any pretreatment is a promising strategy in water analysis.

Activation of the Non-canonical Inflammasome in Mouse and Human Cells.

The non-canonical inflammasome is a signaling platform that allows for the detection of cytoplasmic lipopolysaccharides (LPS) in immune and non-immune cells. Upon detection of LPS, this inflammasome activates the signaling proteases caspase-4 and -5 (in humans) and caspase-11 (in mice). Inflammatory caspases activation leads to caspase self-processing and the cleavage of the pore-forming protein Gasdermin D (GSDMD). GSDMD N-terminal fragments oligomerize and form pores at the plasma membranes, leading to an inflammatory form of cell death called pyroptosis. Here, we describe a simple method to activate the non-canonical inflammasome in myeloid and epithelial cells and to measure its activity using cell death assay and immunoblotting.

Electrochemical biosensor for serogroup specific diagnosis of leptospirosis

A problem with the current leptospirosis diagnostic methods is the low sensitivity and specificity during the acute phase of illness. Rapid point-of-care (POC) assays with minimal sample utilization and low cost are desired in clinical practice. Here, we report for the first time lipopolysaccharide (LPS) based electrochemical biosensor that offers a rapid, highly sensitive, serogroup specific diagnosis of leptospirosis during the acute stage of infection and also to distinguish from other flu like infections. The proposed sensor is fabricated by the immobilization of LPS onto dodecanethiol (DT) modified gold electrode. Monolayer of DT is attached through covalent bond (Au-S) interaction onto the gold electrode. Thus, leptospiral antibodies from the human serum samples bind to the LPS present on self-assembled monolayer (SAM) of DT and showed a higher RCT value compared to SAM. The detection limit of the developed LPS sensor is estimated to be 100 nM. This biosensor is the first electrochemical sensing platform used for detection of LPS from Leptospira spp. This method is completely a solution-based diagnostic method and therefore it is rapid, simple, and sensitive; thus establishing a key technology towards a useful POC diagnostic strategy in serogroup level and hence an alternative to MAT.

Near-Infrared-Fluorescent Probe for Turn-On Lipopolysaccharide Analysis Based on PEG-Modified Gold Nanorods with Plasmon-Enhanced Fluorescence.

Lipopolysaccharide (LPS), as the major component of the outer membrane of Gram-negative bacteria, can trigger a variety of biological effects such as sepsis, septic shock, and even multiorgan failure. Herein, we developed a near-infrared-fluorescent probe for fluorescent turn-on analysis of LPS based on plasmon-enhanced fluorescence (PEF). Gold nanorods (Au NRs) modified polyethylene glycol (PEG) was used as PEF materials. Au NRs were prepared with different longitudinal surface plasmon resonance (LSPR), and their fluorescence enhancement was investigated. Three kinds of molecular weights (1000, 5000, and 10000) of polyethylene glycol (PEG) were employed to control the distance between the Au NRs and the fluorescence substances of cyanine 7 (Cy7). Experimental analysis showed that the enhancement was related to the spectral overlap between the plasmon resonance of Au NRs and the extinction/emission of fluorophore. The three-dimensional finite-difference time-domain (3D-FDTD) simulation further revealed that the enhancement was caused by local electric field enhancement. Furthermore, the probe was used for the ultrasensitive analysis of LPS with a detection limit of 3.85 ng/mL and could quickly distinguish the Gram-negative bacterium-Escherichia coli (E. coli) (with LPS in the membrane) from Gram-positive bacterium-Staphylococcus aureus (S. aureus) (without LPS), as well as quantitative determination of E. coli with a detection limit of 1.0 × 106 cfu/mL. These results suggested that the prepared probe has great potential for biomedical diagnosis and selective detection of LPS from different bacterial strains.

Immunohistochemical Detection of Bacteria in the Resected Valves was Associated with Stromal Immune Checkpoint Protein Expression that may Contribute to Calcific Aortic Stenosis

Aortic stenosis (AS) is a disease characterized by narrowing of the aortic valve (AV) orifice. The purpose of this study was to clarify the significance of bacterial detection and clinicopathological factors, including valve-infiltrating immune cells and disease severity, in relation to AS. After obtaining the written informed consent form from 50 patients with AS, we performed immunohistochemical analysis of lipopolysaccharide (LPS) for gram-negative bacteria and lipoteichoic acid (LTA) for gram-positive bacteria on surgically resected-AVs. Moreover, we evaluated the relationships among the presence of bacteria, immune checkpoint protein PD-L1 expression, and immune cell infiltrations such as CD8-positive T lymphocytes, CD163-positive macrophages, and FOXP3-positive regulatory T cell (Treg) in resected-aortic valves. LPS detection in the resected-aortic valve tissues was significantly associated with stromal PD-L1 expression, valve calcification, and LTA existence in resected samples. We showed that the presence of LPS was significantly related to high PD-L1 expression only in calcified-AVs, not in non-calcified-AVs. Moreover, the high expression of PD-L1 in AS samples without LPS was significantly associated with positive infiltration of CD163-positive macrophages and FOXP3-positive Tregs. Immunohistochemical bacterial detection in resected-aortic valves was associated with PD-L1 accumulation and valve calcification. PD-L1 significantly accumulated only in calcified valves with LPS existence.

Fighting Against Bacterial Lipopolysaccharide-Caused Infections through Molecular Dynamics Simulations: A Review.

Lipopolysaccharide (LPS) is the primary component of the outer leaflet of Gram-negative bacterial outer membranes. LPS elicits an overwhelming immune response during infection, which can lead to life-threatening sepsis or septic shock for which no suitable treatment is available so far. As a result of the worldwide expanding multidrug-resistant bacteria, the occurrence and frequency of sepsis are expected to increase; thus, there is an urge to develop novel strategies for treating bacterial infections. In this regard, gaining an in-depth understanding about the ability of LPS to both stimulate the host immune system and interact with several molecules is crucial for fighting against LPS-caused infections and allowing for the rational design of novel antisepsis drugs, vaccines and LPS sequestration and detection methods. Molecular dynamics (MD) simulations, which are understood as being a computational microscope, have proven to be of significant value to understand LPS-related phenomena, driving and optimizing experimental research studies. In this work, a comprehensive review on the methods that can be combined with MD simulations, recently applied in LPS research, is provided. We focus especially on both enhanced sampling methods, which enable the exploration of more complex systems and access to larger time scales, and free energy calculation approaches. Thereby, apart from outlining several strategies for surmounting LPS-caused infections, this work reports the current state-of-the-art of the methods applied with MD simulations for moving a step forward in the development of such strategies.

Shigella OspC3 suppresses murine cytosolic LPS sensing

SummaryShigella flexneri, a cytosol-invasive gram-negative pathogen, deploys an array of type III-secreted effector proteins to evade host cell defenses. Caspase-11 and its human ortholog caspase-4 detect cytosolic lipopolysaccharide (LPS) and trigger gasdermin D-mediated pyroptosis to eliminate intra-cytoplasmic bacterial threats. However, the role of caspase-11 in combating S. flexneri is unclear. The Shigella T3SS effector OspC3 reportedly suppresses cytosolic LPS sensing by inhibiting caspase-4 but not caspase-11 activity. Surprisingly, we found that S. flexneri also uses OspC3 to inhibit murine caspase-11 activity. Mechanistically, we found that OspC3 binds only to primed caspase-11. Importantly, we demonstrate that S. flexneri employs OspC3 to prevent caspase-11-mediated pyroptosis in neutrophils, enabling bacteria to disseminate and evade clearance following intraperitoneal challenge. In contrast, S. flexneri lacking OspC3 is attenuated in a caspase-11- and gasdermin D-dependent fashion. Overall, our study reveals that OspC3 suppresses cytosolic LPS detection in a broad array of mammals.

A water-soluble and photostable aggregation-induced emission lumogen for imaging Gram-negative bacteria by supramolecular assembly

Detection of lipopolysaccharide (LPS) and Gram-negative bacteria is essential for bacterial diagnostics and healthcare monitoring. Herein, we designed an amphiphilic fluorescent probe (CS-PyB) with an aggregation-induced emission (AIE) feature and high photostability for fluorescence turn-on detection of LPS. The AIE-active probe CS-PyB binds with LPS via multiple intermolecular interactions to form highly emissive aggregates, resulting in a large fluorescence enhancement. CS-PyB is highly selective for LPS over other biological analytes and exhibits a low limit of detection (LOD = 40.7 nM) toward LPS. Besides, high-contrast discrimination of Gram-negative bacteria (E. coli) over Gram-positive bacteria (S. aureus) is successfully achieved using CS-PyB. Significantly, our LPS probe also possesses superior resistance to photobleaching. These remarkable performances render the photostable LPS probe CS-PyB as a robust candidate for long-term dynamic tracking of Gram-negative bacteria associated infections and diseases.

A mutant equipped with a regenerated disulphide for the missing His loop of a serine protease zymogen in the horseshoe crab coagulation cascade.

The lipopolysaccharide (LPS)-triggered coagulation cascade in horseshoe crabs is composed of three zymogens belonging to the trypsinogen family: prochelicerase C, prochelicerase B (proB) and the proclotting enzyme (proCE). Trypsinogen-family members contain three conserved disulphides located around the active site. While it is known that proB evolutionarily lost one of the disulphides, the His-loop disulphide, the roles of the missing His-loop disulphide in proB remain unknown. Here, we prepared a proB mutant, named proB-murasame, equipped with a regenerated His-loop disulphide. The activation rate by upstream α-chelicerase C for proB-murasame was indistinguishable from that for wild-type (WT) proB. The resulting protease chelicerase B-murasame exhibited an 8-fold higher kcat value for downstream proCE than WT chelicerase B, whereas the Km value of chelicerase B-murasame was equivalent to that of WT chelicerase B. WT serpins-1, -2 and -3, identified as scavengers for the cascade, had no reactivity against WT chelicerase B, whereas chelicerase B-murasame was inhibited by WT serpin-2, suggesting that WT chelicerae B may trigger as-yet-unsolved phenomena after performing its duty in the cascade. The reconstituted LPS-triggered cascade containing proB-murasame exhibited ∼5-fold higher CE production than that containing WT proB. ProB-murasame might be used as a high value-adding reagent for LPS detection.

Indoleamine 2,3 dioxygenase, age, and immune activation in people living with HIV

Immune activation complicates HIV despite antiretroviral therapy (ART). Indoleamine 2,3 dioxygenase (IDO) catabolizes tryptophan (T) to kynurenine (K), regulating immune activity, and IDO activity increases with age. This study examines...

Poly-l-lysine-Functionalized Green-Light-Emitting Carbon Dots as a Fluorescence Turn-on Sensor for Ultrasensitive Detection of Endotoxin.

Herein, we report a facile, ultrasensitive, and selective fluorescence turn-on sensing strategy based on green-light-emitting functional nanodots for the detection of bacterial lipopolysaccharide (LPS) endotoxin. In this protocol, first, the pure carbon dots (CDs) with a fairly high quantum yield were prepared by microwave-assisted pyrolysis of citric acid in the presence of urea. Subsequently, the carboxyl-group-rich surfaces of the CDs were allowed to conjugate with the poly-l-lysine (PLL) using an EDC-NHS amidization method to obtain the PLL-modified CDs (PLL-CDs). The LPS could specifically bind to the PLL at the PLL-CD surfaces, and this binding enabled an electron transfer from the phosphate groups of LPS to the carbon core through the PLL bridge, thus resulting in a fluorescence enhancement. Interestingly, this fluorescent turn-on sensor provided a detection limit of 68.3 fM in PBS (pH 7.4), which is the lowest ever reported among all of the synthetic assays for LPS detection. Furthermore, our fluorescent probe was able to show a remarkable selectivity toward LPS over a range of commonly known interfering substances. Thus, this study demonstrated the feasibility of using specific LPS binding to PLL to drive molecular recognition in aqueous medium and offered an effective fluorescence turn-on sensing strategy to detect bacterial endotoxin in diverse clinical and biological applications.

Abstract P576: Plasma Bacterial Lipopolysaccharide Associates With Carotid Atherosclerosis, a Cause of Large Vessel Stroke

Introduction: Inflammation and infection are associated with cerebrovascular diseases including stroke due to carotid atherosclerotic plaques. C-reactive protein (CRP), an acute-phase protein, is upregulated in the plasma of patients with carotid atherosclerotic plaques. However, little is known about whether bacterial molecules trigger inflammation or play a role in patients with carotid atherosclerotic plaques. Recently, it has been recognized that inflammation associated with atherosclerosis and morbidity and mortality in cardiovascular diseases may be due to lipopolysaccharide (LPS) that is found in the outer wall of all Gram-negative bacteria. These findings prompted this study to explore whether plasma levels of LPS and LPS-binding protein (LBP) are elevated and correlated with CRP levels in patients with asymptomatic carotid plaques (ACP). We also compared LBP levels in patients with ACP to large vessel (LV) strokes due to carotid plaques and to matched controls. Methods: Patients (n = 30) with ACP, LV stroke due to carotid atherosclerosis and age-, sex- matched healthy controls gave consent and had their blood drawn. Plasma was processed for LPS, LBP and CRP detection using separate ELISA for each. Results: Plasma LBP level in ACP (22.7 ± 2.92 μg/ml) was similar to LV stroke (21.6 ± 1.56 μg/ml, p = 0.74, ACP vs LV) but greater than controls (13.6 ± 1.43 μg/ml, p = 0.011, ACP vs controls). In ACP patients, plasma LPS level (159.5 ± 30.5 μg/ml) was greater than controls (42.6 ± 11.7 μg/ml, p = 0.001); plasma CRP levels (20.2 ± 6.2 μg/ml) was higher than controls (5.3 ± 2.1 μg/ml, p = 0.011). There was a positive correlation between LPS levels and LBP levels (r = 0.86, p < 0.00001), LPS levels and CRP levels (r = 0.82, p = 0.00001), and LBP levels and CRP levels (r = 0.89, p < 0.00001) in ACP cases. Conclusions: Plasma LPS, LBP and CRP associate with asymptomatic carotid plaques suggesting a pro-inflammatory state exists in patients with asymptomatic carotid plaques, a cause of large vessel stroke. LPS is postulated to directly upregulate both CRP and LBP. Elevated LBP in large vessel stroke patients suggests a Gram-negative bacteria associated post-stroke inflammatory state.

Electrochemical aptasensor for ultrasensitive detection of lipopolysaccharide using silver nanoparticles decorated titanium dioxide nanotube/functionalized reduced graphene oxide as a new redox nanoprobe.

Anovel and relatively simple signal-off electrochemical aptasensor was constructed for highly sensitive detection of lipopolysaccharide (LPS). For the first time, silver nanoparticles (AgNPs) decorated titanium dioxide nanotube (TNT) was conjugated with polydiallyldimethylammonium chloride (PDDA) functionalized reduced graphene oxide (rGO) to form a new nanohybrid of Ag-TNT/P-rGO. This nanohybrid with a large specific surface area exhibited excellent electrochemical activity, which not only served as the sensing platform to immobilize LPS binding aptamer (LBA) but wasalso employed as the redox probe to monitor the change of the electrochemical signal. The electrochemical signal responses were measured by cyclic voltammetry (CV) in the potential range-0.3 to 0.5V at a scan rate of 0.1V/s. The proposed aptasensor exhibited acceptable stability, reproducibility, and specificity for LPS detection with a wide linear range from 17fg/mL to 100ng/mL. Thelimit of detection (LOD) was 5fg/mL. Furthermore, the prepared aptasensor showed acceptable recovery ranging from 96% to 103%, and the RSD varied between 1.4% and 8.5% for determining LPS in real samples.Graphical abstract.

Nanoplasmonic Biosensing Approach for Endotoxin Detection in Pharmaceutical Field

The outer membrane of Gram-negative bacteria contains bacterial endotoxins known as Lipopolysaccharides (LPS). Owing to the strong immune responses induced in humans and animals, these large molecules have a strong toxic effect that can cause severe fever, hypotension, shock, and death. Endotoxins are often present in the environment and medical implants and represent undesirable contaminations of pharmaceutical preparations and medical devices. To overcome the limitations of the standard technique, novel methods for early and sensitive detection of LPS will be of crucial importance. In this work, an interesting approach for the sensitive detection of LPS has been realized by exploiting optical features of nanoplasmonic transducers supporting Localized Surface Plasmon Resonances (LSPRs). Ordered arrays of gold nano-prisms and nano-disks have been realized by nanospheres lithography. The realized transducers have been integrated into a simple and miniaturized lab-on-a-chip (LOC) platform and functionalized with specific antibodies as sensing elements for the detection of LPS. Interactions of specific antibodies anchored on protein A-modified sensor chips with the investigated analyte resulted in a spectral shift in the plasmonic resonance peak of the transducers. A good linear relationship between peak shifts and the LPS concentration has been demonstrated for the fabricated nano-structures with a detection limit down to 5 ng/mL. Integration with a proper microfluidic platform demonstrates the possibility of yielding a prototypal compact device to be used as an analytical test for quality determination of pharmaceutical products.

Bacterial lipopolysaccharide as negative predictor of gemcitabine efficacy in advanced pancreatic cancer \u2013 translational results from the AIO-PK0104 Phase 3 study

BackgroundGram-negative bacteria mediated gemcitabine resistance in pre-clinical models. We determined if intratumoural lipopolysaccharide (LPS) detection by immunohistochemistry is associated with outcome in advanced pancreatic ductal adenocarcinoma (PDAC) treated with gemcitabine and non-gemcitabine containing 1st-line chemotherapy.MethodsWe examined LPS on tumour tissue from 130 patients treated within the randomised AIO-PK0104 trial and a validation cohort (n = 113) and analysed the association of LPS detection to patient outcome according to treatment subgroups.ResultsIn 24% of samples from the AIO-PK0104 study LPS was detected; in LPS-positive patients median OS was 4.4 months, compared to 7.3 months with LPS negative tumours (HR 1.732, p = 0.010). A difference in OS was detected in 1st-line gemcitabine-treated patients (n = 71; HR 2.377, p = 0.002), but not in the non-gemcitabine treatment subgroup (n = 59; HR 1.275, p = 0.478). Within the validation cohort, the LPS positivity rate was 23%, and LPS detection was correlated with impaired OS in the gemcitabine subgroup (n = 94; HR 1.993, p = 0.008) whereas no difference in OS was observed in the non-gemcitabine subgroup (n = 19; HR 2.596, p = 0.219).ConclusionsThe detection of intratumoural LPS as surrogate marker for gram-negative bacterial colonisation may serve as a negative predictor for gemcitabine efficacy in advanced PDAC.Clinical trial registryThe Clinical trial registry identifier is NCT00440167.

Highly sensitive detection of lipopolysaccharide based on collaborative amplification of dual enzymes

In this work, a novel electrochemical biosensor is developed for facile and highly sensitive detection of lipopolysaccharide (LPS) based on collaboration of dual enzymes for multiple-stages signal amplification. Through ingenious design, the specific recognition of target LPS is transformed to the exonuclease III (Exo III)-assisted interface DNA cycling collaborated with the terminal deoxynucleotidyl transferase (TdT)-catalyzed DNA extension, finally inducing significant electrochemical signal concerned with the concentration of LPS. This paper mainly discusses the detection principle, optimization of key factors, and the analytical performance of the biosensor. With the efficient signal amplification, the biosensor shows high sensitivity with a good linearity and a low limit of detection of 1 pg mL−1 for LPS. Moreover, the developed biosensor can clearly discriminate LPS from interferents and show high specificity for LPS detection. This biosensor has also been successfully employed to measure LPS in real food samples, suggesting potential opportunity for application in food safety detection.

TRPV4 Mediates Acute Bladder Responses to Bacterial Lipopolysaccharides.

Urinary tract infections (UTI) affect a large proportion of the population, causing among other symptoms, more frequent and urgent micturition. Previous studies reported that the gram-negative bacterial wall component lipopolysaccharides (LPS) trigger acute epithelial and bladder voiding responses, but the underlying mechanisms remain unknown. The cation channel TRPV4 is implicated in the regulation of the bladder voiding. Since TRPV4 is activated by LPS in airway epithelial cells, we sought to determine whether this channel plays a role in LPS-induced responses in urothelial cells (UCs). We found that human-derived UCs display a fast increase in intracellular Ca2+ concentration upon acute application of Escherichia coli LPS. Such responses were detected also in freshly isolated mouse UCs, and found to be dependent on TRPV4, but not to require the canonical TLR4 signaling pathway of LPS detection. Confocal microscopy experiments revealed that TRPV4 is dispensable for LPS-induced nuclear translocation of NF-κB in mouse UCs. On the other hand, quantitative RT PCR determinations showed an enhanced LPS-induced production of proinflammatory cytokines in TRPV4-deficient UCs. Cystometry experiments in anesthetized wild type mice revealed that acute intravesical instillation of LPS rapidly increases voiding frequency. This effect was not observed in TRPV4-deficient animals, but was largely preserved in Tlr4 KO and Trpa1 KO mice. Our results suggest that activation of TRPV4 by LPS in UCs regulates the proinflammatory response and contributes to LPS-induced increase in voiding frequency. These findings further support the concept that TRP channels are sensors of LPS, mediating fast innate immunity mechanisms against gram-negative bacteria.

Chitosan Stabilized Silver Nanoparticles for the Electrochemical Detection of Lipopolysaccharide: A Facile Biosensing Approach for Gram-Negative Bacteria.

Negatively charged lipopolysaccharide (LPS), a major endotoxin and component of the outer membrane of several Gram-negative bacteria, provides a useful biomarker for the indirect detection of these pathogens. For instance, Escherichia coli (E. coli) is a pathogenic bacterium that causes infections in almost all age groups, and has been implicated in food and water contamination. Current diagnostic and detection methods tend to be labor-intensive or expensive, necessitating the need for an easy, sensitive, rapid, and low-cost method. We report on the synthesis and use of positively charged chitosan stabilized silver nanoparticles (Chi-AgNPs) as a sensitive electrochemical nanobiosensor for the detection of LPS. Chi-AgNPs were synthesized through a facile, single step protocol, and characterized for size, charge, and morphology. Glassy carbon electrodes modified with Chi-AgNPs resulted in an enhancement of signal in the presence of both LPS and E. coli. Detection was accomplished over a large concentration range (several orders of magnitude) of 0.001–100 ng/mL and 10–107 CFU/mL. The biosensors can reliably detect LPS and E. coli at very low concentrations. Chi-AgNPs have potential as low cost, sensitive nanobiosensors for Gram-negative bacteria due to strong electrostatic interaction with LPS present in their outer membranes.