scholarly journals An open-source software analysis package for Microspheres with Ratiometric Barcode Lanthanide Encoding (MRBLEs)

2018 ◽  
Author(s):  
Björn Harink ◽  
Huy Nguyen ◽  
Kurt Thorn ◽  
Polly Fordyce
Keyword(s):  
Open Source ◽  
Solid Phase ◽  
Sample Volume ◽  
Small Sample ◽  
Fluorescent Material ◽  
Software Analysis ◽  
Peptide Motifs ◽  
Reduce Measurement Error ◽  
Analysis Package ◽  
Spectrally Encoded

AbstractMultiplexed bioassays, in which multiple analytes of interest are probed in parallel within a single small volume, have greatly accelerated the pace of biological discovery. Bead-based multiplexed bioassays have many technical advantages, including near solution-phase kinetics, small sample volume requirements, many within-assay replicates to reduce measurement error, and, for some bead materials, the ability to synthesize analytes directly on beads via solid-phase synthesis. To allow bead-based multiplexing, analytes can be synthesized on spectrally encoded beads with a 1:1 linkage between analyte identity and embedded codes. Bead-bound analyte libraries can then be pooled and incubated with a fluorescently-labeled macromolecule of interest, allowing downstream quantification of interactions between the macromolecule and all analytes simultaneously via imaging alone. Extracting quantitative binding data from these images poses several computational image processing challenges, requiring the ability to identify all beads in each image, quantify bound fluorescent material associated with each bead, and determine their embedded spectral code to reveal analyte identities. Here, we present a novel open-source Python software package (the mrbles analysis package) that provides the necessary tools to: (1) find encoded beads in a bright-field microscopy image; (2) quantify bound fluorescent material associated with bead perimeters; (3) identify embedded ratiometric spectral codes within beads; and (4) return data aggregated by embedded code and for each individual bead. We demonstrate the utility of this package by applying it towards analyzing data generated via multiplexed measurement of calcineurin protein binding to MRBLEs (Microspheres with Ratiometric Barcode Lanthanide Encoding) containing known and mutant binding peptide motifs. We anticipate that this flexible package should be applicable to a wide variety of assays, including simple bead or droplet finding analysis, quantification of binding to non-encoded beads, and analysis of multiplexed assays that use ratiometric, spectrally encoded beads.

scholarly journals An open-source software analysis package for Microspheres with Ratiometric Barcode Lanthanide Encoding (MRBLEs)

PLoS ONE ◽  
2019 ◽  
Vol 14 (3) ◽  
pp. e0203725 ◽  
Author(s):  
Björn Harink ◽  
Huy Nguyen ◽  
Kurt Thorn ◽  
Polly Fordyce
Keyword(s):  
Open Source ◽  
Open Source Software ◽  
Software Analysis ◽  
Analysis Package

scholarly journals Development and Performance Evaluation of a Tandem Mass Spectrometry Assay for 4 Adrenal Steroids

2006 ◽  
Vol 52 (8) ◽  
pp. 1559-1567 ◽  
Author(s):  
Mark M Kushnir ◽  
Alan L Rockwood ◽  
William L Roberts ◽  
Elizabeth G Pattison ◽  
William E Owen ◽  
...  
Keyword(s):  
Solid Phase ◽  
Multiple Reaction Monitoring ◽  
Sample Volume ◽  
Reference Intervals ◽  
Small Sample ◽  
Tandem Mass ◽  
Adrenal Steroids ◽  
Adult Males ◽  
Butyl Ether ◽  
Adrenal Steroid

Abstract Background: Congenital adrenal hyperplasia is a group of autosomal recessive disorders caused by a deficiency of 1 of 4 enzymes required for the synthesis of glucocorticoids, mineralocorticoids, and sex hormones. Analysis of 11-deoxycortisol (11DC), 17-hydroxyprogesterone (17OHP), 17-hydroxypregnenolone (17OHPr), and pregnenolone (Pr) in blood allows detection of these enzyme defects. Methods: The steroids were extracted from 200 μL of serum or plasma by solid-phase extraction, derivatized to form oximes, and extracted again with methyl t-butyl ether. Instrumental analysis was performed on an API 4000 tandem mass spectrometer with electrospray ionization in positive mode and multiple reaction-monitoring acquisition. Results: The limits of detection were 0.025 μg/L for 11DC, 17OHP, and Pr and 0.10 μg/L for 17OHPr. The method was linear to 100 μg/L for 11DC, 17OHP, and Pr, respectively, and to 40 μg/L for 17OHPr. Within- and between-run (total) imprecision (CVs) were <7.1% and 11%, respectively. Reference intervals for children in Tanner stages 1 through 5 and adult males and females for 17OHP, 11DC, Pr, and 17OHPr were established. Prepared samples were stable for >72 h. Conclusions: The detection limit and selectivity of this method and its small sample volume requirement allow analysis of endogenous concentrations of adrenal steroids in serum or plasma from children and adults. The method thus has an important potential role in the evaluation of the status of 4 of the enzymes involved in adrenal steroid biosynthesis.

The Use of Solid Phase Microextraction (SPME) to Monitor for Major Organoleptic Compounds Produced by Chrysophytes in Surface Waters

1999 ◽  
Vol 40 (6) ◽  
pp. 251-256 ◽  
Author(s):  
Susan B. Watson ◽  
Brian Brownlee ◽  
Trevor Satchwill ◽  
E. McCauley
Keyword(s):  
Surface Waters ◽  
Solid Phase Microextraction ◽  
Solid Phase ◽  
Small Sample ◽  
Good Recovery ◽  
Culture Material ◽  
Treatment Facilities ◽  
Efficient Extraction ◽  
Cost Efficient ◽  
Related Compounds

An efficient extraction method is needed to measure trace levels of taste and odour compounds in surface waters. This is usually accomplished by costly and involved analytical procedures. We have developed a simpler alternative, using a commercially available microextraction apparatus (SPME). With this technique we successfully monitored trace levels of some target organoleptics (unsaturated aldehydes e.g. heptadienal, nonadienal, and related compounds) which commonly cause aquatic taste and odour. We identified these compounds in culture material, and analyzed for them during the development of odourous chrysophyte blooms in two ponds. Preliminary work has also found a good recovery of some important off-flavour terpenoids (e.g. geosmin and MIB). SPME is labour and cost efficient, and therefore appealing to water treatment facilities for detection and monitoring. In addition, SPME requires only small sample volumes, and is therefore suitable for culture work.

scholarly journals Universal Stokes’s nanomechanical viscometer

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Komal Chaudhary ◽  
Pooja Munjal ◽  
Kamal P. Singh
Keyword(s):  
Electric Field ◽  
Real Time ◽  
Sample Volume ◽  
Capillary Waves ◽  
Small Sample ◽  
Medical Applications ◽  
Rapid Measurement ◽  
Single Lens ◽  
Universal Applicability ◽  
Small Sample Volume

AbstractAlthough, many conventional approaches have been used to measure viscosity of fluids, most methods do not allow non-contact, rapid measurements on small sample volume and have universal applicability to all fluids. Here, we demonstrate a simple yet universal viscometer, as proposed by Stokes more than a century ago, exploiting damping of capillary waves generated electrically and probed optically with sub-nanoscale precision. Using a low electric field local actuation of fluids we generate quasi-monochromatic propagating capillary waves and employ a pair of single-lens based compact interferometers to measure attenuation of capillary waves in real-time. Our setup allows rapid measurement of viscosity of a wide variety of polar, non-polar, transparent, opaque, thin or thick fluids having viscosity values varying over four orders of magnitude from $$10^{0}{-}10^{4}~\text{mPa} \, \text{s}$$ 10 0 - 10 4 mPa s . Furthermore, we discuss two additional damping mechanisms for nanomechanical capillary waves caused by bottom friction and top nano-layer appearing in micro-litre droplets. Such self-stabilized droplets when coupled with precision interferometers form interesting microscopic platform for picomechanical optofluidics for fundamental, industrial and medical applications.

scholarly journals Demonstration of a Label-Free and Low-Cost Optical Cavity-Based Biosensor Using Streptavidin and C-Reactive Protein

Biosensors ◽  
2020 ◽  
Vol 11 (1) ◽  
pp. 4
Author(s):  
Donggee Rho ◽  
Seunghyun Kim
Keyword(s):  
Limit Of Detection ◽  
Point Of Care ◽  
Low Cost ◽  
Optical Cavity ◽  
Sample Volume ◽  
Small Sample ◽  
Label Free ◽  
C Reactive Protein ◽  
Reactive Protein ◽  
Cavity Structure

An optical cavity-based biosensor (OCB) has been developed for point-of-care (POC) applications. This label-free biosensor employs low-cost components and simple fabrication processes to lower the overall cost while achieving high sensitivity using a differential detection method. To experimentally demonstrate its limit of detection (LOD), we conducted biosensing experiments with streptavidin and C-reactive protein (CRP). The optical cavity structure was optimized further for better sensitivity and easier fluid control. We utilized the polymer swelling property to fine-tune the optical cavity width, which significantly improved the success rate to produce measurable samples. Four different concentrations of streptavidin were tested in triplicate, and the LOD of the OCB was determined to be 1.35 nM. The OCB also successfully detected three different concentrations of human CRP using biotinylated CRP antibody. The LOD for CRP detection was 377 pM. All measurements were done using a small sample volume of 15 µL within 30 min. By reducing the sensing area, improving the functionalization and passivation processes, and increasing the sample volume, the LOD of the OCB are estimated to be reduced further to the femto-molar range. Overall, the demonstrated capability of the OCB in the present work shows great potential to be used as a promising POC biosensor.

scholarly journals A tandem giant magnetoresistance assay for one-shot quantification of clinically relevant concentrations of N-terminal pro-B-type natriuretic peptide in human blood

2021 ◽  
Author(s):  
Fanda Meng ◽  
Weisong Huo ◽  
Jie Lian ◽  
Lei Zhang ◽  
Xizeng Shi ◽  
...  
Keyword(s):  
Detection Limit ◽  
Giant Magnetoresistance ◽  
Dynamic Range ◽  
Point Of Care ◽  
Sample Volume ◽  
Small Sample ◽  
Broad Dynamic Range ◽  
Gmr Sensors ◽  
Gmr Sensor ◽  
Sample Volume Requirement

AbstractWe report a microfluidic sandwich immunoassay constructed around a dual-giant magnetoresistance (GMR) sensor array to quantify the heart failure biomarker NT-proBNP in human plasma at the clinically relevant concentration levels between 15 pg/mL and 40 ng/mL. The broad dynamic range was achieved by differential coating of two identical GMR sensors operated in tandem, and combining two standard curves. The detection limit was determined as 5 pg/mL. The assay, involving 53 plasma samples from patients with different cardiovascular diseases, was validated against the Roche Cobas e411 analyzer. The salient features of this system are its wide concentration range, low detection limit, small sample volume requirement (50 μL), and the need for a short measurement time of 15 min, making it a prospective candidate for practical use in point of care analysis.

scholarly journals Sample Preparation and Extraction in Small Sample Volumes Suitable for Pediatric Clinical Studies: Challenges, Advances, and Experiences of a Bioanalytical HPLC-MS/MS Method Validation Using Enalapril and Enalaprilat

2015 ◽  
Vol 2015 ◽  
pp. 1-11 ◽  
Author(s):  
Bjoern B. Burckhardt ◽  
Stephanie Laeer
Keyword(s):  
Sample Preparation ◽  
Solid Phase Extraction ◽  
Method Development ◽  
Matrix Effects ◽  
Solid Phase ◽  
Scale Up ◽  
Small Sample ◽  
Positive Pressure ◽  
Phase Extraction ◽  
Bioanalytical Method

In USA and Europe, medicines agencies force the development of child-appropriate medications and intend to increase the availability of information on the pediatric use. This asks for bioanalytical methods which are able to deal with small sample volumes as the trial-related blood lost is very restricted in children. Broadly used HPLC-MS/MS, being able to cope with small volumes, is susceptible to matrix effects. The latter restrains the precise drug quantification through, for example, causing signal suppression. Sophisticated sample preparation and purification utilizing solid-phase extraction was applied to reduce and control matrix effects. A scale-up from vacuum manifold to positive pressure manifold was conducted to meet the demands of high-throughput within a clinical setting. Faced challenges, advances, and experiences in solid-phase extraction are exemplarily presented on the basis of the bioanalytical method development and validation of low-volume samples (50 μL serum). Enalapril, enalaprilat, and benazepril served as sample drugs. The applied sample preparation and extraction successfully reduced the absolute and relative matrix effect to comply with international guidelines. Recoveries ranged from 77 to 104% for enalapril and from 93 to 118% for enalaprilat. The bioanalytical method comprising sample extraction by solid-phase extraction was fully validated according to FDA and EMA bioanalytical guidelines and was used in a Phase I study in 24 volunteers.

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