scholarly journals Multiple-Cycle Polymeric Extracellular Vesicle Precipitation and Its Evaluation by Targeted Mass Spectrometry

2021 ◽  
Vol 22 (9) ◽  
pp. 4311
Author(s):  
Jisook Park ◽  
Eun-Bi Go ◽  
Ji Sun Oh ◽  
Jong Kyun Lee ◽  
Soo-Youn Lee
Keyword(s):  
Multiple Reaction Monitoring ◽  
Shotgun Proteomics ◽  
Extracellular Vesicle ◽  
Sample Volume ◽  
Small Sample ◽  
Modified Method ◽  
Galectin 3 ◽  
Multiple Cycle ◽  
Time Requirements ◽  
Blood Specimens

The multiple roles of extracellular vesicles (EVs) in pathogenesis have received much attention, as they are valuable as diagnostic and prognostic biomarkers. We employed polymeric EV precipitation to isolate EVs from clinical specimens to overcome the limitations of ultracentrifugation (UC), such as low protein yields, a large volume of specimens used, and time requirements. Multiple-cycle polymeric EV precipitation was applied to enhance the purity of the EV fractions with a small sample volume. Then, the purity was assessed using a multiple reaction monitoring (MRM) panel consisting of alpha-2-macroglobulin (A2M), thrombospondin 1 (THBS 1), galectin 3 binding protein (LGALS3BP), and serum albumin (ALB). For purity evaluation, the EV fractions isolated from blood specimens were subjected to shotgun proteomics and MRM-based targeted proteomics analyses. We demonstrate that the modified method is an easy and convenient method compared with UC. In the shotgun proteomics analysis, the proteome profile of EV fraction contains 89% EV-related proteins by matching with the EVpedia database. In conclusion, we suggest that multiple-cycle polymeric EV precipitation is not only a more effective method for EV isolation for further proteomics-based experiments, but may also be useful for further clinical applications due to the higher EV yield and low sample requirements.

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.

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.

Mediastinal Nodular Lymphocyte Predominant Hodgkin Lymphoma Achieved by Endoscopic Transesophageal Cryobiopsy

Respiration ◽  
2021 ◽  
pp. 1-5
Author(s):  
Zan-Sheng Huang ◽  
Dong Zhou ◽  
Jing Zhang ◽  
Wan-Lei Fu ◽  
Jing Wang ◽  
...  
Keyword(s):  
Hodgkin Lymphoma ◽  
Diagnostic Yield ◽  
Cancer Staging ◽  
Needle Aspiration ◽  
Sample Volume ◽  
Small Sample ◽  
Endobronchial Ultrasound ◽  
Ultrasound Guided ◽  
Transbronchial Needle Aspiration ◽  
Initial Sampling

Guidelines have recommended endobronchial ultrasound-guided transbronchial needle aspiration (EBUS-TBNA) and endoscopic ultrasound-guided fine-needle aspiration biopsy as initial sampling approaches of mediastinal lymph nodes for lung cancer staging. However, the small sample volume might restrict the diagnostic utility of needle aspiration in certain mediastinal diseases. We have recently shown that transbronchial mediastinal cryobiopsy, which is capable of providing larger amounts of intact tissue, improves diagnostic yield in rare tumors and benign diseases compared to EBUS-TBNA. Here, we present a case of mediastinal nodular lymphocyte predominant Hodgkin lymphoma successfully diagnosed by endoscopic transesophageal cryobiopsy.

scholarly journals Perennial Ryegrass Contains Gluten-Like Proteins That Could Contaminate Cereal Crops

2021 ◽  
Vol 8 ◽  
Author(s):  
Sophia Escobar-Correas ◽  
James A. Broadbent ◽  
Alicja Andraszek ◽  
Sally Stockwell ◽  
Crispin A. Howitt ◽  
...  
Keyword(s):  
Multiple Reaction Monitoring ◽  
Wheat Gluten ◽  
Shotgun Proteomics ◽  
Cereal Crops ◽  
Wheat Cultivars ◽  
Liquid Chromatography Mass Spectrometry ◽  
Free Products ◽  
Poaceae Family ◽  
Gliadin Peptides ◽  
First Time

Background: To ensure safe consumption of gluten-free products, there is a need to understand all sources of unintentional contamination with gluten in the food chain. In this study, ryegrass (Lolium perenne), a common weed infesting cereal crop, is analysed as a potential source of gluten-like peptide contamination.Materials and Methods: Ten ryegrass cultivars were analysed using shotgun proteomics for the presence of proteins from the prolamin superfamily. A relative quantitative assay was developed to detect ryegrass gluten-like peptides in comparison with those found in 10 common wheat cultivars.Results: A total of 19 protein accessions were found across 10 cultivars of ryegrass for the protein families of PF00234-Tryp_alpha_amyl, PF13016-Gliadin, and PF03157-Glutenin_HMW. Protein and peptide homology searches revealed that gliadin-like peptides were similar to avenin and gamma-gliadin peptides. A total of 20 peptides, characteristic of prolamin superfamily proteins, were selected for liquid chromatography mass spectrometry (LC-MS) with multiple reaction monitoring (MRM). Only two of the monitored peptides were detected with high abundance in wheat, and all others were detected in ryegrass. Glutenin and alpha-amylase/trypsin inhibitor peptides were reported for the first time in ryegrass and were noted to be conserved across the Poaceae family.Conclusion: A suite of gluten-like peptides were identified using proteomics that showed consistent abundance across ryegrass cultivars but were not detected in wheat cultivars. These peptides will be useful for differentiating wheat gluten contamination from ryegrass gluten contamination.

Design and Performance of a Miniaturized High-Speed Continuous-Flow Analyzer

1974 ◽  
Vol 20 (4) ◽  
pp. 424-427 ◽  
Author(s):  
William E Neeley ◽  
Stephen C Wardlaw ◽  
Helen C Sing
Keyword(s):  
Sample Size ◽  
Continuous Flow ◽  
High Speed ◽  
Clinical Laboratory ◽  
Small Animal ◽  
Sample Volume ◽  
Small Sample ◽  
Reagent Consumption ◽  
And Performance ◽  
Made In

Abstract Design features and performance of a miniaturized high-speed continuous-flow analyzer are described. Special emphasis is made in the design towards a system that is free from the operational and mechanical complexities found in most of today’s advanced systems. Depending on the particular analyses, sample size varies from 3 to 25 µl and reagent consumption is less than 180 µl per sample. Analyses are performed under steady-state conditions at sampling rates of 150 samples per hour with a 2:1 or 3:1 sample-to-wash ratio. The marked reduction in sample size makes the system ideal for microanalyses, especially in the pediatric clinical laboratory, in small animal research, and in any other cases where small sample volume is especially important.

scholarly journals Portable Real-Time Detection of Pb(II) Using a CMOS MEMS-Based Nanomechanical Sensing Array Modified with PEDOT:PSS

Nanomaterials ◽  
2020 ◽  
Vol 10 (12) ◽  
pp. 2454
Author(s):  
Yi-Kuang Yen ◽  
Chao-Yu Lai
Keyword(s):  
Limit Of Detection ◽  
Quality Measurement ◽  
Complementary Metal Oxide Semiconductor ◽  
Cost Effective ◽  
Sample Volume ◽  
Small Sample ◽  
Oxide Semiconductor ◽  
Resistance Change ◽  
Site Water ◽  
Cmos Mems

Detecting the concentration of Pb2+ ions is important for monitoring the quality of water due to it can become a health threat as being in certain level. In this study, we report a nanomechanical Pb2+ sensor by employing the complementary metal-oxide-semiconductor microelectromechanical system (CMOS MEMS)-based piezoresistive microcantilevers coated with PEDOT:PSS sensing layers. Upon reaction with Pb2+, the PEDOT:PSS layer was oxidized which induced the surface stress change resulted in a subsequent bending of the microcantilever with the signal response of relative resistance change. This sensing platform has the advantages of being mass-produced, miniaturized, and portable. The sensor exhibited its sensitivity to Pb2+ concentrations in a linear range of 0.01–1000 ppm, and the limit of detection was 5 ppb. Moreover, the sensor showed the specificity to Pb2+, required a small sample volume and was easy to operate. Therefore, the proposed analytical method described here may be a sensitive, cost-effective and portable sensing tool for on-site water quality measurement and pollution detection.

Specific 3H radioimmunoassay with a monoclonal antibody for monitoring cyclosporine in blood.

1988 ◽  
Vol 34 (2) ◽  
pp. 257-260 ◽  
Author(s):  
P E Ball ◽  
H Munzer ◽  
H P Keller ◽  
E Abisch ◽  
J Rosenthaler
Keyword(s):  
Monoclonal Antibody ◽  
Bone Marrow ◽  
High Performance Liquid Chromatography ◽  
High Performance ◽  
Limit Of Detection ◽  
Parent Drug ◽  
Sample Volume ◽  
Specific Radioimmunoassay ◽  
The Mean ◽  
Blood Specimens

Abstract A specific radioimmunoassay involving a mouse monoclonal antibody to cyclosporine has been developed for monitoring the parent drug in blood. Pretreatment with methanol removes cyclosporine from the erythrocytes. The limit of detection is about 12 micrograms/L, sample volume is 50 microL of blood, and within- and between-assay CVs are less than 7%. Assay results correlated well with those obtained by "high-performance" liquid chromatography (HPLC) for liver (n = 42), for heart (n = 64), for bone-marrow (n = 36), and for kidney (n = 140). For blood specimens obtained from patients treated with cyclosporine postoperatively for as long as 65 months, the mean RIA/HPLC ratio in all with transplant indications was close to 1. Therefore, the specific radioimmunoassay apparently can be used instead of HPLC to measure the parent drug in blood.

scholarly journals Large-scale, high-throughput screening for coagulation and hematologic phenotypes in mice*

2002 ◽  
Vol 11 (3) ◽  
pp. 185-193 ◽  
Author(s):  
Luanne L. Peters ◽  
Eleanor M. Cheever ◽  
Heather R. Ellis ◽  
Phyllis A. Magnani ◽  
Karen L. Svenson ◽  
...  
Keyword(s):  
High Throughput ◽  
High Throughput Screening ◽  
Large Scale ◽  
Inbred Mouse ◽  
Inbred Strains ◽  
Sample Volume ◽  
Small Sample ◽  
Mouse Strains ◽  
Phenotypic Data ◽  
Multiple Tests

The Mouse Phenome Project is an international effort to systematically gather phenotypic data for a defined set of inbred mouse strains. For such large-scale projects the development of high-throughput screening protocols that allow multiple tests to be performed on a single mouse is essential. Here we report hematologic and coagulation data for more than 30 inbred strains. Complete blood counts were performed using an Advia 120 analyzer. For coagulation testing, we successfully adapted the Dade Behring BCS automated coagulation analyzer for use in mice by lowering sample and reagent volume requirements. Seven automated assay procedures were developed. Small sample volume requirements make it possible to perform multiple tests on a single animal without euthanasia, while reductions in reagent volume requirements reduce costs. The data show that considerable variation in many basic hematological and coagulation parameters exists among the inbred strains. These data, freely available on the World Wide Web, allow investigators to knowledgeably select the most appropriate strain(s) to meet their individual study designs and goals.

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