Barry R. Lutz

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Papers by Barry R. Lutz

Fluidic Considerations of Measuring Intracranial Pressure Using an Open External Ventricular Drain

Cureus

Measurement of intracranial pressure (ICP) during cerebrospinal fluid (CSF) drainage with an exte... more Measurement of intracranial pressure (ICP) during cerebrospinal fluid (CSF) drainage with an external ventricular drain (EVD) typically requires stopping the flow during measurement. However, there may be benefits to simultaneous ICP measurement and CSF drainage. Several studies have evaluated whether accurate ICP measurements can be obtained while the EVD is open. They report differing outcomes when it comes to error, and hypothesize several sources of error. This study presents an investigation into the fluidic sources of error for ICP measurement with concurrent drainage in an EVD. Our experiments and analytical model both show that the error in pressure measurement increases linearly with flow rate and is not clinically significant, regardless of drip chamber height. At physiologically relevant flow rates (40 mL/hr) and ICP set points (13.6-31.3 cmH 2 O or 10-23 mmHg), our model predicts an underestimation of 0.767 cmH 2 O (0.56 mmHg) with no observed data point showing error greater than 1.09 cmH 2 O (0.8 mmHg) in our experiment. We extrapolate our model to predict a realistic worst-case clinical scenario where we expect to see a mean maximum error of 1.06 cmH 2 O (0.78 mmHg) arising from fluidic effects within the drainage system for the most resistive catheter. Compared to other sources of error in current ICP monitoring, error in pressure measurement due to drainage flow is small and does not prohibit clinical use. However, other effects such as ventricular collapse or catheter obstruction could affect ICP measurement under continuous drainage and are not investigated in this study.

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Two-Dimensional Paper 1 Networks: Multi-Step Fluidic Programming Using a Fluid Source Well & Shaped Paper

Most laboratory assays take advantage of multi-step protocols to achieve high performance, but co... more Most laboratory assays take advantage of multi-step protocols to achieve high performance, but conventional paper-based tests (e.g., lateral flow tests) are generally limited to assays that can be carried out in a single fluidic step. We present a method to automate multi-step chemical delivery in a simple paper device via a single user activation step. A fluid well is used as a buffer source to feed a paper network; wicking causes the fluid level in the well to decrease, and different paper legs are disconnected in a timed sequence to allow sequential timed delivery of fluids to a detection zone.

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Point-Of-Care Immunoassay Cards for Sample-To-Result Differential Diagnosis of Acute Fever

We present air-driven microfluidic cards that perform sample processing and parallel immunoassays... more We present air-driven microfluidic cards that perform sample processing and parallel immunoassays for antigens and IgM antibodies. The disposable cards are based on a flow-through membrane immunoassay carried out on porous nitrocellulose, which provides rapid diffusion for short assay times and a high surface area for visual detection. Malarial antigen, Plasmodium falciparum HRPII antigen, showed a limit of detection of 20-30 ng/mL spiked into fetal bovine serum. Detection of IgM antibodies to Salmonella Typhi, from clinical plasma samples, correlated closely with bench-top tests. All reagents were stored dried on card; only sample and buffer were required to run the test.

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Novel Sample Processing Modules for Enhanced Paper-Based Diagnostics

One substantial limitation to conventional paper-based diagnostics is their lack of sample proces... more One substantial limitation to conventional paper-based diagnostics is their lack of sample processing capabilities. Here we demonstrate paper-based devices for mixing/dilution and specific protein depletion. The mixing/dilution module demonstrated here can be tuned to achieve a wide range of dilutions. The specific protein depletion module in this work used IgG as a model target and achieved ~90% depletion in buffer spiked samples. Both sample processing modules are compatible with lateral flow tests and add significant functionality without requiring additional steps from the end-user. Paper devices were composed of nitrocellulose (HFB135, 8-10 µm pore diameter, Millipore, Billerica, MA), with wicking pads (Millipore, Billerica, MA) placed at the distal end of the paper devices to control fluid flow. Paper materials were cut using a CO2 laser cutter (Universal Laser Systems, Scottsdale, AZ). The solutions used for mixing/dilution were 800 µM erioglaucine (blue dye), 1870 µM tartraz...

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A Two-Dimensional Paper Network for Comprehesive Dengue Detection at the Point of Care

The two-dimensional paper network (2DPN) is an inexpensive technology based on the strengths of o... more The two-dimensional paper network (2DPN) is an inexpensive technology based on the strengths of one-dimensional lateral flow rapid diagnostic tests. The 2DPN allows complex (bio)chemical processing to be performed without the need for external instrumentation, which is ideally suited to point-of-care testing for infectious disease in low resource settings. We have been exploring the use of 2DPNs for sensitive multiplexed immunoassays. We present progress in the development of 2DPN modules to be used in a multiplexed amplified test for dengue virus proteins and two types of antibodies against them.

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Rapid Near Point-of-Care Assay for HLA-B*57:01 Genotype Associated with Severe Hypersensitivity Reaction to Abacavir

AIDS Research and Human Retroviruses, 2021

doi: medRxiv preprint NOTE: This preprint reports new research that has not been certified by pee... more

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Harmony COVID-19: a ready-to-use kit, low-cost detector, and smartphone app for point-of-care SARS-CoV-2 RNA detection

ABSTRACTRNA amplification tests allow sensitive detection of SARS-CoV-2 infection, but their comp... more ABSTRACTRNA amplification tests allow sensitive detection of SARS-CoV-2 infection, but their complexity and cost are prohibitive for expanding COVID-19 testing. We developed “Harmony COVID-19”, a point-of-care test using inexpensive consumables, ready-to-use reagents, and a simple device that processes up to 4 samples simultaneously. Our lyophilized reverse-transcription, loop-mediated isothermal amplification (RT-LAMP) can detect as little as 15 SARS-CoV-2 RNA copies per reaction, and it can report as early as 17 min for samples with high viral load (2 x 105 RNA copies per reaction). Analysis of RNA extracted from clinical nasal specimens (n = 101) showed 95% concordance with RT-PCR, including 100% specificity in specimens positive for other viruses and bacteria. Analysis of contrived samples in the nasal matrix showed detection of 92% or 100% in samples with ≥20 or ≥100 particles per reaction, respectively. Usability testing showed 95% accuracy by healthcare workers operating the ...

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Implementation of an interactive mobile application to pilot a rapid assay to detect HIV drug resistance mutations in Kenya

IntroductionUsability is an overlooked aspect of implementing lab-based assays, particularly nove... more IntroductionUsability is an overlooked aspect of implementing lab-based assays, particularly novel assays in low-resource-settings. Esoteric instructions can lead to irreproducible test results and patient harm. To address these issues, we developed a software application based on “Aquarium”, a laboratory-operating system run on a computer tablet that provides step-by-step digital interactive instructions, protocol management, and sample tracking. Aquarium was paired with a near point-of-care HIV drug resistance test, “OLA-Simple”, that detects mutations associated with virologic failure. In this observational study we evaluated the performance of Aquarium in guiding untrained users through the multi-step laboratory protocol with little supervision.MethodsA feasibility study was conducted in a laboratory at Coptic Hope Center in Nairobi, Kenya. To evaluate the training by Aquarium software, twelve volunteers who were unfamiliar with the kit performed the test on blinded samples (2 b...

format_quoteAll 12 technicians completed the OLA-Simple workflow successfully, achieving interpretation accuracy rates of 90% and 97%.format_quote

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Ultrasensitive hybridization capture: Reliable detection of <1 copy/mL short cell-free DNA from large-volume urine samples

PLOS ONE, 2021

Urine cell-free DNA (cfDNA) is a valuable non-invasive biomarker with broad potential clinical ap... more Urine cell-free DNA (cfDNA) is a valuable non-invasive biomarker with broad potential clinical applications, but there is no consensus on its optimal pre-analytical methodology, including the DNA extraction step. Due to its short length (majority of fragments <100 bp) and low concentration (ng/mL), urine cfDNA is not efficiently recovered by conventional silica-based extraction methods. To maximize sensitivity of urine cfDNA assays, we developed an ultrasensitive hybridization method that uses sequence-specific oligonucleotide capture probes immobilized on magnetic beads to improve extraction of short cfDNA from large-volume urine samples. Our hybridization method recovers near 100% (95% CI: 82.6–117.6%) of target-specific DNA from 10 mL urine, independent of fragment length (25–150 bp), and has a limit of detection of ≤5 copies of double-stranded DNA (0.5 copies/mL). Pairing hybridization with an ultrashort qPCR design, we can efficiently capture and amplify fragments as short a...

format_quoteShort PCR amplicon lengths significantly boost detection sensitivity, achieving 100% for 25 bp fragments and 56% for Epstein-Barr virus at reduced lengths.format_quote

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SwabExpress: An end-to-end protocol for extraction-free COVID-19 testing

Structured BackgroundThe urgent need for massively scaled clinical testing for SARS-CoV-2, along ... more Structured BackgroundThe urgent need for massively scaled clinical testing for SARS-CoV-2, along with global shortages of critical reagents and supplies, has necessitated development of streamlined laboratory testing protocols. Conventional nucleic acid testing for SARS-CoV-2 involves collection of a clinical specimen with a nasopharyngeal swab in transport medium, nucleic acid extraction, and quantitative reverse transcription PCR (RT-qPCR) (1). As testing has scaled across the world, the global supply chain has buckled, rendering testing reagents and materials scarce (2). To address shortages, we developed SwabExpress, an end-to-end protocol developed to employ mass produced anterior nares swabs and bypass the requirement for transport media and nucleic acid extraction.MethodsWe evaluated anterior nares swabs, transported dry and eluted in low-TE buffer as a direct-to-RT-qPCR alternative to extraction-dependent viral transport media. We validated our protocol of using heat treatme...

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OLA-Simple: A software-guided HIV-1 drug resistance test for low-resource laboratories

EBioMedicine, 2019

Background: HIV drug resistance (HIVDR) testing can assist clinicians in selecting treatments. Ho... more Background: HIV drug resistance (HIVDR) testing can assist clinicians in selecting treatments. However, high complexity and cost of genotyping assays limit routine testing in settings where HIVDR prevalence has reached high levels. Methods: The oligonucleotide ligation assay (OLA)-Simple kit was developed for detection of HIVDR against first-line non-nucleoside/nucleoside reverse transcriptase inhibitors and validated on 672 codons (168 specimens) from subtypes A, B, C, D, and AE. The kit uses dry reagents to facilitate assay setup, lateral flow devices for visual HIVDR detections, and in-house software with an interface for guiding users and analyzing results. Findings: HIVDR analysis of specimens by OLA-Simple compared to Sanger sequencing revealed 99.6 § 0.3% specificity and 98.2 § 0.9% sensitivity, and compared to high-sensitivity assays, 99.6 § 0.6% specificity and 86.2 § 2.5% sensitivity, with 2.6 § 0.9% indeterminate results. OLA-Simple was performed more rapidly compared to Sanger sequencing (<4 h vs. 35À72 h). Forty-one untrained volunteers blindly tested two specimens each with 96.8 § 0.8% accuracy. Interpretation: OLA-Simple compares favorably with HIVDR genotyping by Sanger and sensitive comparators. Instructional software enabled inexperienced, first-time users to perform the assay with high accuracy. The reduced complexity, cost, and training requirements of OLA-Simple could improve access to HIVDR testing in low-resource settings and potentially allow same-day selection of appropriate antiretroviral therapy.

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Threshold-Based Quantification in a Multiline Lateral Flow Assay via Computationally Designed Capture Efficiency

Analytical chemistry, Jan 4, 2018

Lateral flow assays (LFAs) are widely used for yes/no detection of analytes, but they are not wel... more Lateral flow assays (LFAs) are widely used for yes/no detection of analytes, but they are not well-suited for quantification. We show that the sensitivity of the test line in a lateral flow assay can be tuned to appear at a specific sample concentration by varying the density of capture molecules at the test line and that when test lines tuned for different responses are combined into a single test strip, lines appear at specific thresholds of sample concentration. We also developed a model based on mass-action kinetics that accurately described test line signal and shape over a wide matrix of capture molecules and sample concentrations in single-line strips. The model was used to design a three-line test strip with lines designed to appear at logarithmically spaced sample concentrations, and the experiments showed a remarkable match to predictions. The response of this "graded ladder bar" format is due to the effect of test line concentration on capture efficiency at each...