An extraction free modified o-phthalaldehyde assay for quantifying residual protein and microbial biofilms on surfaces.

Biological contamination of surfaces in industry and healthcare is an important vector of disease transmission. Current assays for detecting surface-adherent contamination require extraction of biological soil. However, physical inaccessibility or poor solubility may limit recovery. Here, how the o-phthalaldehyde (OPA) protein assay can be modified to measure residual protein (modeled with bovine serum albumin) or biofilm on a surface without extraction is described. The assay limit of detection (LOD) for protein was 1.6 µg cm-2. The detection threshold for Staphylococcus epidermis biofilm was 117 µg cm-2. The clinical utility of the method was demonstrated for measurements taken from clinically used endoscopes. Since this method is more sensitive than extraction-based testing, clinical results should not be compared with conventional benchmarks. By enabling direct detection and quantification of soils in complex or hard-to-reach areas, this method has potential to improve the margin of safety in medical and industrial cleaning processes.

Hemoglobin assay for validation and quality control of medical device reprocessing.

Hemoglobin (Hb) is an important analyte in medicine, forensics, and research. One area of crucial need for real-world Hb quantitation is the validation and quality control (QC) of reprocessed medical device cleaning. Here, we show how a microplate reader and colorimetric blood test strips can be used to quantitate nanogram (ng) quantities of Hb in a 1-min assay. The assay had a linear range of 0-50 ng (0-370 ng on a log scale) for Hb, with a limit of detection (LOD) of 3.3 ng, which was ∼500-fold more sensitive than the micro-BCA reagent (LOD = 1.6 µg) and on the same order of magnitude as detection of labeled Hb with fluorescence (LOD = 1.9 ng). For validation of medical device cleaning, the assay was specific for Hb in the presence of artificial test soil and was unaffected by interferences from common cleaning reagents at 10 ppm. Lubricant and sodium dodecyl sulfate did not significantly affect the assay at 10 ppm but affected the assay at 1 % g/g. The method showed 100 % recovery of hemoglobin in extracted soils, with extraction from silicone having the greatest variability in recovery, while Teflon and stainless steel had <10 % RSD. The assay makes it possible for medical device companies and health-care providers to obtain crucially needed information on the cleanliness of reused devices.

The effects of non-ionic polymeric surfactants on the cleaning of biofouled hydrogel materials.

Block co-polymer surfactants have been used for cleaning hydrogel medical devices that contact the body (e.g., contact lenses) because of their biocompatibility. This work examined the relationship between concentration and detergency of two non-ionic polymeric surfactants (Pluronic F127 and Triton X-100) for cleaning protein soil, with anionic surfactants (sodium dodecyl sulfate and sodium laureth sulfate) as positive controls. Surface plasmon resonance was used to quantify removal of simulated tear soil from self-assembled monolayer surfaces, and a microplate format was used to study the removal of fluorescently labeled soil proteins from contact lenses. While detergency increased as a function of concentration for anionic surfactants, it decreased with concentration for the two polymeric surfactants. The fact that the protein detergency of some non-ionic polymeric surfactants did not increase with concentration above the critical micelle concentration could have implications for optimizing the tradeoff between detergency and biocompatibility.

The effect of fluorescent labels on protein sorption in polymer hydrogels.

Hydrogels are an increasingly important class of medical device materials that enable diverse and unique function, but can also be subject to significant biofouling and contamination. Although it is challenging to accurately quantify protein biofouling in hydrogels, spectroscopic detection of fluorescently labeled proteins is one method with the potential to provide direct, sensitive quantitation in transparent materials. Therefore, it is important to understand how fluorophores can affect protein-material interactions in hydrogels. This work uses an independent method, native ultraviolet fluorescence (native UV) of proteins, in conjunction with labeled protein fluorescence and the bicinchoninic acid assay (BCA), to assess the effect of fluorescent labels on protein sorption in polymer hydrogels. Bovine serum albumin (BSA) and lysozyme (LY) were labeled with two common but structurally different fluorophores and used as model biofouling proteins in three contact lens hydrogel materials. Native UV was used to directly measure both labeled and unlabeled protein sorption, while orthogonal measurements were performed with extrinsic fluorescence and BCA assay to compare with the native UV results. Sorption of labeled proteins was found to be <2-fold higher than unlabeled proteins on most protein-material combinations, while differences of up to 10-fold were observed for labeled BSA in more hydrophobic hydrogels. Fluorescence recovery after photobleaching (FRAP) also showed that the fluorescent label chemistry can significantly affect surface adsorption of sorbed proteins on the internal surfaces of hydrogels. This study reveals the complex nature of fluorophore-protein-material interactions and shows the potential of native UV for investigating unlabeled protein biofouling in hydrogels.

Analytical Chemistry in the Regulatory Science of Medical Devices.

In the United States, regulatory science is the science of developing new tools, standards, and approaches to assess the safety, efficacy, quality, and performance of all Food and Drug Administration-regulated products. Good regulatory science facilitates consumer access to innovative medical devices that are safe and effective throughout the Total Product Life Cycle (TPLC). Because the need to measure things is fundamental to the regulatory science of medical devices, analytical chemistry plays an important role, contributing to medical device technology in two ways: It can be an integral part of an innovative medical device (e.g., diagnostic devices), and it can be used to support medical device development throughout the TPLC. In this review, we focus on analytical chemistry as a tool for the regulatory science of medical devices. We highlight recent progress in companion diagnostics, medical devices on chips for preclinical testing, mass spectrometry for postmarket monitoring, and detection/characterization of bacterial biofilm to prevent infections.

Model for Porosity Changes Occurring during Ultrasound-Enhanced Transcorneal Drug Delivery.

Ultrasound-enhanced drug delivery through the cornea has considerable therapeutic potential. However, our understanding of how ultrasound enhances drug transport is poor, as is our ability to predict the increased level of transport for given ultrasound parameters. Described here is a computational model for quantifying changes in corneal porosity during ultrasound exposure. The model is calibrated through experiments involving sodium fluorescein transport through rabbit cornea. Validation was performed using nylon filters, for which the properties are known. It was found that exposure to 800-kHz ultrasound at an intensity 2 W/cm2 for 5 min increased the porosity of the epithelium by a factor of 5. The model can be useful for determining the extent to which ultrasound enhances the amount of drug transported through biological barriers, and the time at which a therapeutic dose is achieved at a given location, for different drugs and exposure strategies.

A contact-lens-on-a-chip companion diagnostic tool for personalized medicine.

We present a novel, microfluidic platform that integrates human tears (1 µL) with commercial contact lens materials to provide personalized assessment of lens care solution performance. This device enabled the detection of significant differences in cleaning and disinfection outcomes between subjects and between biofilms vs. planktonic bacteria.

Interactions of Staphylococcus aureus with ultrasoft hydrogel biomaterials.

Ultrasoft biomaterials-polymers, gels, and human soft tissues with an elastic modulus less than ∼100 kPa-are increasingly used in medical devices. While bacterial interactions (adhesion and biofilm formation) have been extensively studied on stiffer materials, little is known about how bacteria colonize ultrasoft materials as a nidus for infection. The goal of this work was to determine how material properties of ultrasoft hydrogels used for dermal fillers might affect pathogenesis of associated infections. We first synthesized a range of polyacrylamide hydrogels (PAAm) with moduli similar to clinically used dermal fillers and characterized the rheological, morphological and porous properties. We then developed a novel microfabricated insert to contain the PAAm in a flow system for quantification of bacterial adhesion and biofilm formation. The rate of adhesion and numbers of adherent Staphylococcus aureus on the surface of PAAm both decreased as the modulus increased. Adhesion was reduced by 3 logs (from 93 × 10(4)/cm(2) to 0.083 × 10(4)/cm(2)) with increasing modulus (from 17 Pa to 654 Pa). However, the number of bacteria in the bulk was the highest within the stiffest gels. This trend was further amplified in subsequent biofilm studies, where interfacial coverage of biofilm decreased as the modulus increased, while the fraction of biofilm in the bulk was the highest within the stiffest gel. The results show significant differences in bacterial colonization of PAAm based on material properties, and reveal how the injection process may unexpectedly create discontinuities that provide a microenvironmental niche for bacterial colonization.

Streamline based design guideline for deterministic microfluidic hydrodynamic single cell traps.

A prerequisite for single cell study is the capture and isolation of individual cells. In microfluidic devices, cell capture is often achieved by means of trapping. While many microfluidic trapping techniques exist, hydrodynamic methods are particularly attractive due to their simplicity and scalability. However, current design guidelines for single cell hydrodynamic traps predominantly rely on flow resistance manipulation or qualitative streamline analysis without considering the target particle size. This lack of quantitative design criteria from first principles often leads to non-optimal probabilistic trapping. In this work, we describe an analytical design guideline for deterministic single cell hydrodynamic trapping through the optimization of streamline distributions under laminar flow with cell size as a key parameter. Using this guideline, we demonstrate an example design which can achieve 100% capture efficiency for a given particle size. Finite element modelling was used to determine the design parameters necessary for optimal trapping. The simulation results were subsequently confirmed with on-chip microbead and white blood cell trapping experiments.

A smartphone controlled handheld microfluidic liquid handling system.

Microfluidics and lab-on-a-chip technologies have made it possible to manipulate small volume liquids with unprecedented resolution, automation and integration. However, most current microfluidic systems still rely on bulky off-chip infrastructures such as compressed pressure sources, syringe pumps and computers to achieve complex liquid manipulation functions. Here, we present a handheld automated microfluidic liquid handling system controlled by a smartphone, which is enabled by combining elastomeric on-chip valves and a compact pneumatic system. As a demonstration, we show that the system can automatically perform all the liquid handling steps of a bead-based HIV1 p24 sandwich immunoassay on a multi-layer PDMS chip without any human intervention. The footprint of the system is 6 × 10.5 × 16.5 cm, and the total weight is 829 g including battery. Powered by a 12.8 V 1500 mAh Li battery, the system consumed 2.2 W on average during the immunoassay and lasted for 8.7 h. This handheld microfluidic liquid handling platform is generally applicable to many biochemical and cell-based assays requiring complex liquid manipulation and sample preparation steps such as FISH, PCR, flow cytometry and nucleic acid sequencing. In particular, the integration of this technology with read-out biosensors may help enable the realization of the long-sought Tricorder-like handheld in vitro diagnostic (IVD) systems.