Electroosmotic Perfusion-Microdialysis Probe Created by Direct Laser Writing for Quantitative Assessment of Leucine Enkephalin Hydrolysis by Insulin-Regulated Aminopeptidase in Vivo.

There are many processes that actively alter the concentrations of solutes in the extracellular space. Enzymatic reactions, either by soluble enzymes or membrane-bound ectoenzymes, and uptake or clearance are two such processes. Investigations of ectoenzymatic reactions in vivo is challenging, particularly in the brain. Studies using microdialysis have revealed some qualitative information about what enzymes may be present, but microdialysis is a sampling technique so it is not designed to control conditions such as a substrate concentration outside the probe. Micropush-pull perfusion has been used to determine which nitric oxide synthase enzymes are active in discrete regions of the rat retina. Ectopeptidases are a particularly important class of ectoenzymes. As far as it is known, the extracellular activity of active peptides in the brain is controlled by ectopeptidases. To understand ectopeptidase activity, we developed a physical probe and an accompanying method. The probe has a two-channel source that supplies substrate or substrate plus inhibitor using electroosmotic perfusion (EOP). It also has a microdialysis probe to collect products and unreacted substrate. The method provides quantitative estimates of substrate-to-product conversion and the influence of inhibitors on this process. The quantitative estimates are made possible by including a d-amino acid-containing peptide analog of the substrate in the substrate-containing solution infused. Quantitative analysis of substrate, substrate analog, and products is carried out by quantitative, online capillary liquid chromatography-tandem mass spectrometry. The electroosmotic perfusion-microdialysis probe and associated method were used to determine the effect of the selective inhibitor HFI-419 on insulin-regulated aminopeptidase (EC 3.4.11.3) in the rat neocortex.

Neighborhood Disadvantage and Hospital Quality Ratings in the Medicare Hospital Compare Program.

BACKGROUND: The Centers for Medicare and Medicaid Services provide nationwide hospital ratings that may influence reimbursement. These ratings do not account for the social risk of communities and may inadvertently penalize hospitals that service disadvantaged neighborhoods. OBJECTIVE: This study examines the relationship between neighborhood social risk factors (SRFs) and hospital ratings in Medicare's Hospital Compare Program. RESEARCH DESIGN: 2017 Medicare Hospital Compare ratings were linked with block group data from the 2015 American Community Survey to assess hospital ratings as a function of neighborhood SRFs. SUBJECTS: A total of 3608 Medicare-certified hospitals in 50 US states. MEASURES: Hospital summary scores and 7 quality group scores (100 percentile scale), including effectiveness of care, efficiency of care, hospital readmission, mortality, patient experience, safety of care, and timeliness of care. RESULTS: Lower hospital summary scores were associated with caring for neighborhoods with higher social risk, including a reduction in hospital score for every 10% of residents who reported dual-eligibility for Medicare/Medicaid [-3.3%; 95% confidence interval (CI), -4.7 to -2.0], no high-school diploma (-0.8%; 95% CI, -1.5 to -0.1), unemployment (-1.2%; 95% CI, -1.9 to -0.4), black race (-1.2%; 95% CI, -1.7 to -0.8), and high travel times to work (-2.5%; 95% CI, -3.3 to -1.6). Associations between neighborhood SRFs and hospital ratings were largest in the timeliness of care, patient experience, and hospital readmission groups; and smallest in the safety, efficiency, and effectiveness of care groups. CONCLUSIONS: Hospitals serving communities with higher social risk may have lower ratings because of neighborhood factors. Failing to account for neighborhood social risk in hospital rating systems may reinforce hidden disincentives to care for medically underserved areas in the United States.

Multiplicative On-Column Solute Focusing Using Spatially Dependent Temperature Programming for Capillary HPLC.

The benefits of capillary liquid chromatography columns are truly realized when small, limited sample volumes require signal enhancement, but the available sample volume does not permit on-column focusing during injection onto a larger column. This dilemma is common when samples are naturally small or precious (such as in biological, forensic, art, and archeological investigations) and analyte concentrations are low. Signal enhancement by solvent-based focusing is effective with capillary columns, but it is limited to a single band-compression step and can only be achieved at the inlet. Here we evaluate multiplicative temperature-assisted solute focusing using a linear array of ten independently controlled 1.0 × 1.0 cm thermoelectric cooling elements (TECs) to generate dynamic temperature changes along the length of the column. The evaluation has two prongs: simulation and experimental. Simulation is required to understand the effect of a particular temperature change at a particular place and time on the column to determine optimal timing of temperature changes. Because the accuracy of the simulations is good, as long as the effect of temperature on retention factor is known, experimental conditions required to achieve a particular focusing objective can be estimated. We evaluated the capability of the technique to selectively focus only one of two solutes. This was achieved using three adjacent zones with temperature controlled by (upstream first) four, two, and one TECs. The three focusing steps occurring on column gave a 20-fold increase in peak height without solvent-based focusing for a solute with modest retention enthalpy.

Regional Epidemiology of Methicillin-Resistant Staphylococcus aureus Among Adult Intensive Care Unit Patients Following State-Mandated Active Surveillance.

Background: In 2007, Illinois became the first state in the United States to mandate active surveillance of methicillin-resistant Staphylococcus aureus (MRSA). The Illinois law applies to intensive care unit (ICU) patients; contact precautions are required for patients found to be MRSA colonized. However, the effectiveness of a legislated "search and isolate" approach to reduce MRSA burden among critically ill patients is uncertain. We evaluated whether the prevalence of MRSA colonization declined in the 5 years after the start of mandatory active surveillance. Methods: All hospitals with an ICU having ≥10 beds in Chicago, Illinois, were eligible to participate in single-day serial point prevalence surveys. We assessed MRSA colonization among adult ICU patients present at time of survey using nasal and inguinal swab cultures. The primary outcome was region-wide MRSA colonization prevalence over time. Results: All 25 eligible hospitals (51 ICUs) participated in serial point prevalence surveys over 8 survey periods (2008-2013). A total of 3909 adult ICU patients participated in the point prevalence surveys, with 432 (11.1%) found to be colonized with MRSA (95% confidence interval [CI], 10.1%-12.0%). The MRSA colonization prevalence among patients was unchanged during the study period; year-over-year relative risk for MRSA colonization was 0.97 (95% CI, .89-1.05; P = .48). Conclusions: MRSA colonization prevalence among critically ill adult patients did not decline during the time period following legislatively mandated MRSA active surveillance. Our findings highlight the limits of legislated MRSA active surveillance as a strategy to reduce MRSA colonization burden among ICU patients.

On-Column Dimethylation with Capillary Liquid Chromatography-Tandem Mass Spectrometry for Online Determination of Neuropeptides in Rat Brain Microdialysate.

We have developed a method for online collection and quantitation of neuropeptides in rat brain microdialysates using on-column dimethylation with capillary liquid chromatography-tandem mass spectrometry (cLC-MS2). This method addresses a number of the challenges of quantifying neuropeptides with cLC-MS. It is also a completely automated and robust method for the preparation of stable isotope labeled-peptide internal standards to correct for matrix effects and thus ensure accurate quantitation. Originally developed for tissue-derived proteomics samples ( Raijmakers et al. Mol. Cell. Proteomics 2008 , 7 , 1755 - 1762 ), the efficacy of on-column dimethylation for native peptides in microdialysate has not been demonstrated until now. We have modified the process to make it more amenable to the time scale of microdialysis sampling and to reduce the accumulation of nonvolatile contaminants on the column and, thus, loss of sensitivity. By decreasing labeling time, we have a temporal resolution of 1 h from sample loading to elution and our peptide detection limits are in the low pM range for 5 µL injections of microdialysate. We have demonstrated the effectiveness of this method by quantifying basal and potassium stimulated concentrations of the neuropeptides leu-enkephalin and met-enkephalin in the rat hippocampus. To our knowledge, this is the first report of quantitation of these peptides in the hippocampus using MS.

Synthesis, Structure, and Acidity Constants of Ligated α-Boryl Acetic Acids.

Basic hydrolyses of various ligated α-boryl acetic acid esters provided the first ligated derivatives of the unknown compound boroacetic acid (BH2 CH2 CO2 H). Four monoacids (L-BH2 CH2 CO2 H) and one diacid (L-BH(CH2 CO2 H)2 ) were prepared with N-heterocyclic carbene, amine, and pyridine ligands (L). The stable acids were characterized by X-ray crystallography and acidity constant (pKa ) measurements. They rank among the least acidic of all known carboxylic acids. In turn, their conjugate bases are among the strongest of all carboxylates.

Numerical Modeling of Electroosmotic Push-Pull Perfusion and Assessment of Its Application to Quantitative Determination of Enzymatic Activity in the Extracellular Space of Mammalian Tissue.

Many sampling methods have been developed to measure the extracellular concentrations of solutes in the extracellular space of mammalian tissue, e.g., brain. However, few have been used to quantitatively study the various processes, such as enzymatic degradation, that determines the fate of these solutes. For a method to be useful in this pursuit, it must be able to (1) perfuse tissue and collect the perfusate for quantitative analysis of the solutes introduced and reaction products produced, (2) control the average residence time of the active solutes, and (3) have the appropriate spatial resolution for the process of interest. Our lab previously developed a perfusion technique based on electroosmosis (EO), called EO push-pull perfusion (EOPPP), that is in principle suitable to meet these needs. However, much like the case for other sampling methods that came before, there are parameters that are needed for quantitative interpretation of data but that cannot be measured easily (or at all). In this paper, we present a robust finite element model that provides a deep understanding of fluid dynamics and mass transport in the EOPPP method, assesses the general applicability of EOPPP to studying enzyme activity in the ECS, and grants a simple approach to data treatment and interpretation to obtain, for example, Vmax and Km for an enzymatic reaction in the extracellular space of the tissue. This model is a valuable tool in optimizing and planning experiments without the need for costly experiments.

Graphical Method for Choosing Optimized Conditions Given a Pump Pressure and a Particle Diameter in Liquid Chromatography.

The general limitations on liquid chromatographic performance in isocratic and gradient elution are now well understood. Many workers have contributed to this understanding and to developing graphical methods, or plots, to illustrate the capabilities of chromatographic systems over a wide range of values of operational parameters. These have been invaluable in getting a picture, in broad strokes, about the value of changing an operational parameter or the value of one separation approach over another. Here we present a plotting approach more appropriate for determining how to use chromatography most efficiently in one's own laboratory. The axes are linear: column length vertical and mobile phase velocity horizontal. In this coordinate system, straight lines with intercept zero correspond to different values of t0. Hyperbolas correspond to values of pressure as the product of length and velocity is proportional to pressure. For a given relationship between theoretical plate height and velocity (e.g., van Deemter), the number of theoretical plates as a function of column length and mobile phase velocity is a surface (z direction) to the x and y of velocity and length. By representing the surface as contours, a two-dimensional plot results. Any point along a constant pressure hyperbola represents the best one can do given the particle diameter, solute diffusion coefficient, and temperature. The user can quickly see how to use the pressure for speed or for more theoretical plates. Sets of such plots allow for comparisons among particle diameters or temperatures. Analogous plots of peak capacity for gradient elution are equally revealing. The plots lead instantly to understanding liquid chromatographic optimization at a practical level. They neatly illustrate the value (or not) of changing pump pressure, particle diameter, or temperature for fast or slow separations in either isocratic or gradient elution. They are illustrated with a focus on maximizing plate count with a given analysis time (isocratic), the effect of volume overload (isocratic), and separations of a limited number of peptides with a peak capacity coming from statistical peak overlap theory (gradient).

Improving the Sensitivity, Resolution, and Peak Capacity of Gradient Elution in Capillary Liquid Chromatography with Large-Volume Injections by Using Temperature-Assisted On-Column Solute Focusing.

Capillary HPLC (cLC) with gradient elution is the separation method of choice for the fields of proteomics and metabolomics. This is due to the complementary nature of cLC flow rates and electrospray or nanospray ionization mass spectrometry (ESI-MS). The small column diameters result in good mass sensitivity. Good concentration sensitivity is also possible by injection of relatively large volumes of solution and relying on solvent-based solute focusing. However, if the injection volume is too large or solutes are poorly retained during injection, volume overload occurs which leads to altered peak shapes, decreased sensitivity, and lower peak capacity. Solutes that elute early even with the use of a solvent gradient are especially vulnerable to this problem. In this paper, we describe a simple, automated instrumental method, temperature-assisted on-column solute focusing (TASF), that is capable of focusing large volume injections of small molecules and peptides under gradient conditions. By injecting a large sample volume while cooling a short segment of the column inlet at subambient temperatures, solutes are concentrated into narrow bands at the head of the column. Rapidly raising the temperature of this segment of the column leads to separations with less peak broadening in comparison to solvent focusing alone. For large volume injections of both mixtures of small molecules and a bovine serum albumin tryptic digest, TASF improved the peak shape and resolution in chromatograms. TASF showed the most dramatic improvements with shallow gradients, which is particularly useful for biological applications. Results demonstrate the ability of TASF with gradient elution to improve the sensitivity, resolution, and peak capacity of volume overloaded samples beyond gradient compression alone. Additionally, we have developed and validated a double extrapolation method for predicting retention factors at extremes of temperature and mobile phase composition. Using this method, the effects of TASF can be predicted, allowing determination of the usefulness of this technique for a particular application.

Chicago Ebola Response Network (CERN): A Citywide Cross-hospital Collaborative for Infectious Disease Preparedness.

The 2014-2015 Ebola virus disease (EVD) epidemic and international public health emergency has been referred to as a "black swan" event, or an event that is unlikely, hard to predict, and highly impactful once it occurs. The Chicago Ebola Response Network (CERN) was formed in response to EVD and is capable of receiving and managing new cases of EVD, while also laying the foundation for a public health network that can anticipate, manage, and prevent the next black swan public health event. By sharing expertise, risk, and resources among 4 major academic centers, Chicago created a sustainable network to respond to the latest in a series of public health emergencies. In this respect, CERN is a roadmap for how a region can prepare to respond to public health emergencies, thereby preventing negative impacts through planning and implementation.

In Vivo Monitoring of Dopamine by Microdialysis with 1 min Temporal Resolution Using Online Capillary Liquid Chromatography with Electrochemical Detection.

Microdialysis is often applied to understanding brain function. Because neurotransmission involves rapid events, increasing the temporal resolution of in vivo measurements is desirable. Here, we demonstrate microdialysis with online capillary liquid chromatography for the analysis of 1 min rat brain dialysate samples at 1 min intervals. Mobile phase optimization involved adjusting the pH, buffer composition, and surfactant concentration to eliminate interferences with the dopamine peak. By analyzing electrically evoked dopamine transients carefully synchronized with the switching of the online LC sample valve, we demonstrate that our system has both 1 min sampling capabilities and bona fide 1 min temporal resolution. Evoked DA transients were confined to single, 1 min brain dialysate samples. After uptake inhibition with nomifensine (20 mg/kg i.p.), responses to electrical stimuli of 1 s duration were detected.

Electroosmotic perfusion of tissue: sampling the extracellular space and quantitative assessment of membrane-bound enzyme activity in organotypic hippocampal slice cultures.

This review covers recent advances in sampling fluid from the extracellular space of brain tissue by electroosmosis (EO). Two techniques, EO sampling with a single fused-silica capillary and EO push-pull perfusion, have been developed. These tools were used to investigate the function of membrane-bound enzymes with outward-facing active sites, or ectoenzymes, in modulating the activity of the neuropeptides leu-enkephalin and galanin in organotypic-hippocampal-slice cultures (OHSCs). In addition, the approach was used to determine the endogenous concentration of a thiol, cysteamine, in OHSCs. We have also investigated the degradation of coenzyme A in the extracellular space. The approach provides information on ectoenzyme activity, including Michaelis constants, in tissue, which, as far as we are aware, has not been done before. On the basis of computational evidence, EO push-pull perfusion can distinguish ectoenzyme activity with a ~100 µm spatial resolution, which is important for studies of enzyme kinetics in adjacent regions of the rat hippocampus.

Integrated electroosmotic perfusion of tissue with online microfluidic analysis to track the metabolism of cystamine, pantethine, and coenzyme A.

We have developed an approach that integrates electroosmotic perfusion of tissue with a substrate-containing solution and online microfluidic analysis of products, in this case thiols. Using this approach we have tracked the metabolism of cystamine, pantethine and CoA in the extracellular space of organotypic hippocampal slice cultures (OHSCs). Currently, little is known about coenzyme A (CoA) biodegradation and even less is known about the regulation and kinetic characteristics for this sequential multienzyme reaction. We found that the steady state percentage yields of cysteamine from cystamine and pantethine during the transit through OHSCs were 91% ± 4% (SEM) and 0.01%-0.03%, respectively. The large difference in the yields of cysteamine can be used to explain the drugs' different toxicities and clinical effectiveness against cystinosis. The kinetic parameters of the enzyme reaction catalyzed by the ectoenzyme pantetheinase are KM,C/α = 4.4 ± 1.1 mM and Vmax,C = 29 ± 3 nM/s, where α is the percentage yield of pantethine to pantetheine through disulfide exchange. We estimate that the percentage yield of pantethine to pantetheine through disulfide exchange is approximately 0.5%. Based on the formation rate of cysteamine in the OHSCs, we obtained the overall apparent Michaelis constant and maximum reaction rate for sequential, extracellular CoA degradation in an in situ environment, which are K'M = 16 ± 4 µM, V'max = 7.1 ± 0.5 nM/s. Kinetic parameters obtained in situ, although difficult to measure, are better representations of the biochemical flux in the living organism than those from isolated enzymes in vitro.

An in situ measurement of extracellular cysteamine, homocysteine, and cysteine concentrations in organotypic hippocampal slice cultures by integration of electroosmotic sampling and microfluidic analysis.

We demonstrate an all-electric sampling/derivatization/separation/detection system for the quantitation of thiols in tissue cultures. Extracellular fluid collected from rat organotypic hippocampal slice cultures (OHSCs) by electroosmotic flow through an 11 cm (length) × 50 µm (i.d.) sampling capillary is introduced to a simple microfluidic chip for derivatization, continuous flow-gated injection, separation, and detection. With the help of a fluorogenic, thiol-specific reagent, ThioGlo-1, we have successfully separated and detected the extracellular levels of free reduced cysteamine, homocysteine, and cysteine from OHSCs within 25 s in a 23 mm separation channel with a confocal laser-induced fluorescence (LIF) detector. Attention to the conductivities of the fluids being transported is required for successful flow-gated injections. When the sample conductivity is much higher than the run buffer conductivities, the electroosmotic velocities are such that there is less fluid coming by electroosmosis into the cross from the sample/reagent channel than is leaving by electroosmosis into the separation and waste channels. The resulting decrease in the internal fluid pressure in the injection cross pulls flow from the gated channel. This process may completely shut down the gated injection. Using a glycylglycine buffer with physiological osmolarity but only 62% of physiological conductivity and augmenting the conductivity of the run buffers solved this problem. Quantitation is by standard additions. Concentrations of cysteamine, homocysteine, and cysteine in the extracellular space of OHSCs are 10.6 ± 1.0 nM (n = 70), 0.18 ± 0.01 µM (n = 53), and 11.1 ± 1.2 µM (n = 70), respectively. This is the first in situ quantitative estimation of endogenous cysteamine in brain tissue. Extracellular levels of homocysteine and cysteine are comparable with other reported values.

In vivo monitoring of serotonin in the striatum of freely moving rats with one minute temporal resolution by online microdialysis-capillary high-performance liquid chromatography at elevated temperature and pressure.

Online monitoring of serotonin in striatal dialysate from freely moving rats was carried out for more than 16 h at 1 min time resolution using microdialysis coupled online to a capillary HPLC system operating at about 500 bar and 50 °C. Several aspects of the system were optimized toward robust, in vivo online measurements. A two-loop, eight-port rotary injection valve demonstrated better consistency of continuous injections than the more commonly used two-loop, 10-port valve. A six-port loop injector for introducing stimulating solutions (stimulus injector) was placed in-line between the syringe pump and microdialysis probe. We minimized solute dispersion by using capillary tubing (75 µm inside diameter, 70 cm long) for the probe inlet and outlet. In vitro assessment of concentration dispersion during transport with a 30 s time resolution showed that the dispersion standard deviation for serotonin was well within the desired system temporal resolution. Each 30 or 60 s measurement reflects the integral of the true time response over the measurement time. We have accounted for this mathematically in determining the concentration dispersion during transport. The delay time between a concentration change at the probe and its detection is 7 min. The timing of injections from the stimulus injector and the cycle time for the HPLC monitoring of the flow stream were controlled. The electrochemical detector contained a 13 µm spacer to minimize detector dead volume. During in vivo experiments, retention time and separation efficiency were stable and reproducible. There was no statistically significant change over 5.5 h in the electrochemical detector sensitivity factor for serotonin. Dialysate serotonin concentrations change significantly in response to a 120 mM K(+) stimulus. Release of serotonin evoked by a 10 min, 120 mM K(+) stimulation, but not for other K(+) stimuli, exhibited a reproducible, oscillating profile of dialysate serotonin concentration versus time. Infusion of fluoxetine, a serotonin uptake inhibitor, increased dialysate serotonin concentrations and stimulated release magnitude. Transient serotonin increases were observed in response to the stress associated with unexpected handling. This system is simple, efficient, reliable, and suitable for the study of serotonin neurochemistry associated with emotion and behavior.

Probing enzymatic activity inside single cells.

We report a novel approach for determining the enzymatic activity within a single suspended cell. Using a steady-state microfluidic delivery device and timed exposure to the pore-forming agent digitonin, we controlled the plasma membrane permeation of individual NG108-15 cells. Mildly permeabilized cells (~100 pores) were exposed to a series of concentrations of fluorescein diphosphate (FDP), a fluorogenic alkaline phosphatase substrate, with and without levamisole, an alkaline phosphatase inhibitor. We generated quantitative estimates for intracellular enzyme activity and were able to construct both dose-response and dose-inhibition curves at the single-cell level, resulting in an apparent Michaelis contant Km of 15.3 µM ± 1.02 (mean ± standard error of the mean (SEM), n = 16) and an inhibition constant Ki of 0.59 mM ± 0.07 (mean ± SEM, n = 14). Enzymatic activity could be monitored just 40 s after permeabilization, and five point dose-inhibition curves could be obtained within 150 s. This rapid approach offers a new methodology for characterizing enzyme activity within single cells.

Electroosmotic Perfusion, External Microdialysis: Simulation and Experiment.

Information about the rates of hydrolysis of neuropeptides by extracellular peptidases can lead to a quantitative understanding of how the steady-state and transient concentrations of neuropeptides are controlled. We have created a small microfluidic device that electroosmotically infuses peptides into, through, and out of the tissue to a microdialysis probe outside the head. The device is created by two-photon polymerization (Nanoscribe). Inferring quantitative estimates of a rate process from the change in concentration of a substrate that has passed through tissue is challenging for two reasons. One is that diffusion is significant, so there is a distribution of peptide substrate residence times in the tissue. This affects the product yield. The other is that there are multiple paths taken by the substrate as it passes through tissue, so there is a distribution of residence times and thus reaction times. Simulation of the process is essential. The simulations presented here imply that a range of first order rate constants of more than 3 orders of magnitude is measurable and that 5-10 min is required to reach a steady state value of product concentration following initiation of substrate infusion. Experiments using a peptidase-resistant d-amino acid pentapeptide, yaGfl, agree with simulations.

Nanocomposite Teflon AF 2400 films as tunable platforms for selective transport.

The performance of polymeric film-based sensors, separations, including extractions, depends on solute transport rates and selectivity. The membrane's chemical composition, its state (e.g, crystalline, glassy, rubbery), and its fractional free volume are all important in defining performance attributes. Other properties of films important in sensors are robustness in the environment, chemical inertness and biocompatibility, thermal stability, and optical transparency. With the long-term goal of selective transport/extraction based on molecular recognition, we have focused on fluorous media such as Teflon AF 2400. We present a novel approach to create nanocomposite Teflon AF 2400 films with the polymer in different states to facilitate permeation and fluorous selectivity in liquid phase transport. Films cast from stable suspensions of the fluorocarbon polymer Teflon AF 2400 (T(g) ∼ 240 °C), fluoroalkylsilane-modified solid, low polydispersity silica nanoparticles (FNPs: 116 nm diameter), and with or without a plasticizer (perfluorotripentylamine, FC-70) are macroscopically homogeneous. The nanocomposite films with glassy polymer absorb considerable solvent, CHCl(3), when in contact with solutions. Thus, the films are very permeable to solutes (toluene and octafluorotoluene) from CHCl(3) solution with poor selectivity for the fluorinated solute. Plasticized Teflon AF nanocomposite films show very low solvent sorption, improved fluorocarbon/hydrocarbon selectivity, and excellent transport rates. This is an unprecedented example demonstrating the effect of a plasticizer to create polymer nanocomposites with different chemical and barrier properties. The state of the polymer in the nanocomposites dictates chemical properties. The chemical properties dictate the transport behavior. In all cases, the films are dimensionally and thermally quite stable, making them ideal materials for applications in separations and sensors.

Iontophoresis from a micropipet into a porous medium depends on the ζ-potential of the medium.

Iontophoresis uses electricity to deliver solutes into living tissue. Often, iontophoretic ejections from micropipets into brain tissue are confined to millisecond pulses for highly localized delivery, but longer pulses are common. As hippocampal tissue has a ζ-potential of approximately -22 mV, we hypothesized that, in the presence of the electric field resulting from the iontophoretic current, electroosmotic flow in the tissue would carry solutes considerably farther than diffusion alone. A steady state solution to this mass transport problem predicts a spherically symmetrical solute concentration profile with the characteristic distance of the profile depending on the ζ-potential of the medium, the current density at the tip, the tip size, and the solute electrophoretic mobility and diffusion coefficient. Of course, the ζ-potential of the tissue is defined by immobilized components of the extracellular matrix as well as cell-surface functional groups. As such, it cannot be changed at will. Therefore, the effect of the ζ-potential of the porous medium on ejections is examined using poly(acrylamide-co-acrylic acid) hydrogels with various magnitudes of ζ-potential, including that similar to hippocampal brain tissue. We demonstrated that nearly neutral fluorescent dextran (3 and 70 kD) solute penetration distance in the hydrogels and OHSCs depends on the magnitude of the applied current, solute properties, and, in the case of the hydrogels, the ζ-potential of the matrix. Steady state solute ejection profiles in gels and cultures of hippocampus can be predicted semiquantitatively.