Fluorescent-based nanosensors for selective detection of a wide range of biological macromolecules: A comprehensive review.

Microbial biofilms have caused serious concerns in healthcare, medical, and food industries because of their intrinsic resistance against conventional antibiotics and cleaning procedures and their capability to firmly adhere on surfaces for persistent contamination. These global issues strongly motivate researchers to develop novel methodologies to investigate the kinetics underlying biofilm formation, to understand the response of the biofilm with different chemical and physical treatments, and to identify biofilm-specific drugs with high-throughput screenings. Meanwhile microbial biofilms can also be utilized positively as sensing elements in cell-based sensors due to their strong adhesion on surfaces. In this perspective, we provide an overview on the connections between sensing and microbial biofilms, focusing on tools used to investigate biofilm properties, kinetics, and their response to chemicals or physical agents, and biofilm-based sensors, a type of biosensor using the bacterial biofilm as a biorecognition element to capture the presence of the target of interest by measuring the metabolic activity of the immobilized microbial cells. Finally we discuss possible new research directions for the development of robust and rapid biofilm related sensors with high temporal and spatial resolutions, pertinent to a wide range of applications.

https://oist.repo.nii.ac.jp/?action=repository_action_common_download&item_id=2602&item_no=1&attribute_id=22&file_no=1

Detection and Characterization of Bacterial Biofilms and Biofilm-Based Sensors.

The property of organic light-responsive materials is determined by their electronic excited states to a large extent, for instance, the radiative decay rate constants, redox potentials, and lifetimes. Tuning the excited state properties with external stimuli will lead to versatile functional materials; a representative example is the fluorescence molecular probes, in which the singlet excited states are controlled by the external stimuli, i.e., by interaction with the analytes. In comparison, controlling the triplet excited state with external stimuli has been rarely reported, although it is also crucial for the development of novel materials for targeted photodynamic therapy (PDT) reagents and phosphorescent molecular probes. The reported results show that the principles used in singlet excited state tuning are unable to be simply applied to the triplet excited state. In this review article, we summarized the recent results on controlling the triplet excited states by the external stimuli (chemical or light), and the application of the triplet state tuning in the chemical/light controllable triplet–triplet-annihilation upconversion (TTA UC). We discussed the methods for the control of the triplet states, as well as singlet excited state, for the purpose of controlling the TTA UC. Both successful and unsuccessful methods are discussed. This information is helpful for understanding the photophysical processes in which the triplet excited state is involved, and the development of novel external stimuli-responsive triplet photosensitizers.

Controlling the triplet states and their application in external stimuli-responsive triplet-triplet-annihilation photon upconversion: from the perspective of excited state photochemistry.

Integrated phosphorescence-based photonic biosensor (iPOB) for monitoring oxygen levels in 3D cell culture systems.

Typical ionophore-based nanosensors use Nile blue derived indicators called chromoionophores, which must contend with strong background absorption, autofluorescence, and scattering in biological samples that limit their usefulness. Here, we demonstrate potassium-selective nanosensors that utilize triplet-triplet annihilation upconversion to minimize potential optical interference in biological media and a pH-sensitive quencher molecule to modulate the upconversion intensity in response to changes in analyte concentration. A triplet-triplet annihilation dye pair (platinum(II) octaethylporphyrin and 9,10-diphenylanthracene) was integrated into nanosensors containing an analyte binding ligand (ionophore), charge-balancing additive, and a pH indicator quencher. The nanosensor response to potassium was shown to be reversible and stable for 3 days. In addition, the nanosensors are selective against sodium, calcium, and magnesium (selectivity coefficients in log10 units of -2.2 for calcium, -2.0 for sodium, and -2.4 for magnesium), three interfering ions found in biological samples. The lack of signal overlap between the upconversion nanosensors and GFP, a common biological fluorescent indicator, is demonstrated in confocal microscope images of sensors embedded in a bacterial biofilm.

Triplet-Triplet Annihilation Upconversion Based Nanosensors for Fluorescence Detection of Potassium.

Bacterial bioﬁlms can form persistent infections on wounds and implanted medical devices and are associated with many chronic diseases, such as cystic fibrosis. These infections are medically difficult to treat, as biofilms are more resistant to antibiotic attack than their planktonic counterparts. An understanding of the spatial and temporal variation in the metabolism of biofilms is a critical component toward improved biofilm treatments. To this end, we developed oxygen-sensitive luminescent nanosensors to measure three-dimensional (3D) oxygen gradients, an application of which is demonstrated here with Pseudomonas aeruginosa biofilms. The method was applied here and improves on traditional one-dimensional (1D) methods of measuring oxygen profiles by investigating the spatial and temporal variation of oxygen concentration when bioﬁlms are challenged with antibiotic attack. We observed an increased oxygenation of biofilms that was consistent with cell death from comparisons with antibiotic kill curves for PAO1. Due to the spatial and temporal nature of our approach, we also identified spatial and temporal inhomogeneities in the biofilm metabolism that are consistent with previous observations. Clinical strains of P. aeruginosa subjected to similar interrogation showed variations in resistance to colistin and tobramycin, which are two antibiotics commonly used to treat P. aeruginosa infections in cystic fibrosis patients.IMPORTANCE Biofilm infections are more difficult to treat than planktonic infections for a variety of reasons, such as decreased antibiotic penetration. Their complex structure makes biofilms challenging to study without disruption. To address this limitation, we developed and demonstrated oxygen-sensitive luminescent nanosensors that can be incorporated into biofilms for studying oxygen penetration, distribution, and antibiotic efficacy-demonstrated here with our sensors monitoring antibiotic impacts on metabolism in biofilms formed from clinical isolates. The significance of our research is in demonstrating not only a nondisruptive method for imaging and measuring oxygen in biofilms but also that this nanoparticle-based sensing platform can be modified to measure many different ions and small molecule analytes.

Luminescent Nanosensors for Ratiometric Monitoring of Three-Dimensional Oxygen Gradients in Laboratory and Clinical Pseudomonas aeruginosa Biofilms.

Biofilms form complex structures that are ubiquitous in natural environments and can cause chronic infections which are difficult to treat. While much is known about biofilms, many questions remain about how biofilms mature and respond to internal and external stimuli. To understand the phenomena that occur within the biofilm matrix, sensing techniques capable of quantitatively resolving spatial and temporal dynamics of key analytes in the biofilm are needed. Biofilm spatial and temporal heterogeneity provide unique hurdles in fully assessing these intricacies. This review provides a detailed overview of methods that have been used to sense a variety of relevant analytes in biofilms. Electrochemical, optical, and other analytical techniques each possess distinct strengths and weaknesses depending on the application showing that biofilm sensing does not have a “one-size fits all” solution. The analyte being measured, the environment it is analyzed in, and the information that is needed determine whether known approaches may be appropriate or novel methods need to be developed. This review aims to help readers understand these techniques so that they can be applied to future projects for better understanding of biofilm elements and evolving sensing approaches.

A review of chemosensors and biosensors for monitoring biofilm dynamics

In clinical environments, many serious antibiotic-resistant infections are caused by biofilm-forming species. This presents issues when attempting to determine antimicrobial dosing as traditional antibiotic susceptibility tests (ASTs) are typically designed around planktonic bacteria and thus offer information that is not relevant to the biofilm phenotype present in the patient. Even the popular Calgary biofilm device may provide inaccurate minimum biofilm inhibitory concentrations (MBICs) and can be time- and material-intensive. In this work, we present a method utilizing oxygen-sensitive nanosensor technology to monitor the oxygen consumption dynamics of living biofilms as they are exposed to antibiotics. We incorporated our nanosensors into biofilms grown from P. aeruginosa strains of varying sensitivity to traditional classes of antibiotics. Through measuring nanosensor response under antibiotic administration we determined the concentrations able to cease biofilm metabolism. This method provides information on the MBIC as well as kinetic response information in a manner that requires fewer materials and is more reflective of biofilm behavior than a traditional AST.

Nanodiagnostics to monitor biofilm oxygen metabolism for antibiotic susceptibility testing

https://www.mdpi.com/2079-6374/10/9/120/pdf

Megan P. Jewell

Papers

Triplet-Triplet Annihilation Upconversion Based Nanosensors for Fluorescence Detection of Potassium.

Luminescent Nanosensors for Ratiometric Monitoring of Three-Dimensional Oxygen Gradients in Laboratory and Clinical Pseudomonas aeruginosa Biofilms.

A review of chemosensors and biosensors for monitoring biofilm dynamics

Nanodiagnostics to monitor biofilm oxygen metabolism for antibiotic susceptibility testing

LipiSensors: Exploiting Lipid Nanoemulsions to Fabricate Ionophore-Based Nanosensors.