A rapid immunoassay to determine concentrations of busulfan in plasma

2007 ◽  
Vol 25 (18_suppl) ◽  
pp. 7111-7111
Author(s):  
S. J. Salamone ◽  
J. Courtney ◽  
R. Harney ◽  
S. He ◽  
G. Lundell
Keyword(s):  
Sample Preparation ◽  
Dose Adjustment ◽  
Cross Reactivity ◽  
Clinical Samples ◽  
High Dose ◽  
Standard Curve ◽  
Routine Monitoring ◽  
Low Levels ◽  
Reporter Group ◽  
Peroxidase Conjugate

7111 Background: High-dose busulfan is an important component of many bone marrow transplantation preparative regimens. High busulfan plasma levels have been shown to increase the chance of veno-occlusive disease and low levels are associated with recurrence of disease or graph rejection. Currently, busulfan levels are monitored by physical methods which are expensive and time- consuming, resulting in relatively low overall use of busulfan testing for dose adjustment. Methods: Novel, highly selective antibodies for busulfan have been generated and micro-titer plate immunoassays that are capable of quantifying busulfan levels in plasma have been developed. The assay was configured using a busulfan-horseradish peroxidase conjugate as the reporter group and busulfan monoclonal antibodies. The assay requires only 5 μL of plasma per determination with no sample preparation. Results: The immunoassay has a standard curve based on busulfan with a range of 75 to 2,000 ng/mL. The time to first result is 30 minutes with up to 240 tests generated per hour. The coefficient of variation (CV) on signal is < 5% for an entire plate and the 95% confidence interval for negative samples (n=78) is below the lowest calibrator of 75 ng/mL. Cross-reactivity with the major inactive metabolites (tetrahydrothiophene,tetramethyl sulfone and tetrahydrothiophene-3-ol-1,1-dioxide), are <0.1%. Clinical samples (n=70) correlate well to LC-MSMS or GC-MS (R > 0.97) with a slope of < 1.05. Conclusions: This immunoassay method may be suitable for determining levels of busulfan in human plasma. It offers the advantages of using a smaller sample size, does not require sample preparation and is less labor intensive than other methods. The ability to make 240 determinations per hour enables effective routine monitoring of busulfan levels in clinical practice. No significant financial relationships to disclose.

Therapeutic monitoring of cyclosporine: impact of a change in standards on 125I-monoclonal RIA performance in comparison with liquid chromatography

1990 ◽  
Vol 36 (5) ◽  
pp. 804-807 ◽  
Author(s):  
P A Keown ◽  
J Glenn ◽  
J Denegri ◽  
U Maciejewska ◽  
D Seccombe ◽  
...  
Keyword(s):  
Whole Blood ◽  
Cross Reactivity ◽  
Recent Change ◽  
Therapeutic Monitoring ◽  
Clinical Samples ◽  
Marrow Graft ◽  
Standard Curve ◽  
The Impact ◽  
Control Samples

Abstract This study examines the measurement of cyclosporine (CsA) by 125I-monoclonal RIA, and describes the impact of the recent change in the standard curve provided. CsA concentrations in serum and whole-blood control samples measured by 125I-RIA were initially 8-18% higher than those by HPLC. During the first two months of 1989, a significant and sustained deviation in the 125I-RIA produced results that exceeded the HPLC results by 21-28% (P less than 0.001). Introduction of the new standard curve in March 1989 returned the concentration of the whole-blood controls to the previous range (11-12% above HPLC, P less than 0.001). Measurement of clinical samples from heart, liver, and bone-marrow graft recipients by 125I-RIA by both old and new kit standards produced a close linear correlation (y = 0.89 x - 19.02; r = 0.99; n = 75, range = 40-850 micrograms/L), with use of the new standards yielding results 82 (SD 8)% of those with the preceding assay. However, even with the new standard curve, CsA concentrations by 125I-RIA in the clinical samples exceeded those by HPLC by a factor of 1.37 (SD 0.18) to 1.52 (SD 0.19). Segregation for transplant type did not affect the RIA/HPLC ratio. The results suggest cross-reactivity of the 125I-RIA with material present in vivo.

Comparative evaluation of a new immunoradiometric assay for corticotropin

2006 ◽  
Vol 44 (5) ◽  
Author(s):  
Charles W. Wilkinson ◽  
Hershel Raff
Keyword(s):  
The United States ◽  
Cross Reactivity ◽  
Clinical Samples ◽  
Immunoradiometric Assay ◽  
Standard Curve ◽  
United States Food ◽  
In Vitro Diagnostic ◽  
States Food ◽  
Diagnostic Measurement

AbstractWe have characterized the performance of a commercial two-site immunoradiometric assay for manual in vitro diagnostic measurement of plasma corticotropin from Scantibodies Laboratory. We compared the results with those of a similar commonly used assay from Nichols Institute Diagnostics that has recently been withdrawn from production. The lower detection limit, range of the standard curve, cross-reactivity, and intra-assay and inter-assay imprecision of the two assays are very similar. Measurement of clinical samples and a series of samples from an experimental subject demonstrate high correlations between the two assays. These factors, together with recent clearance by the United States Food and Drug Administration for manual in vitro diagnostic measurement, make the Scantibodies corticotropin immunoradiometric assay an appropriate replacement for the Nichols assay.

scholarly journals SARS-CoV-2 within-host diversity and transmission

Science ◽  
2021 ◽  
Vol 372 (6539) ◽  
pp. eabg0821 ◽  
Author(s):  
Katrina A. Lythgoe ◽  
Matthew Hall ◽  
Luca Ferretti ◽  
Mariateresa de Cesare ◽  
George MacIntyre-Cockett ◽  
...  
Keyword(s):  
United Kingdom ◽  
Severe Acute Respiratory Syndrome ◽  
Phylogenetic Tree ◽  
Immune Escape ◽  
Clinical Samples ◽  
Diversity Patterns ◽  
Host Diversity ◽  
Viral Loads ◽  
The United Kingdom ◽  
Low Levels

Extensive global sampling and sequencing of the pandemic virus severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have enabled researchers to monitor its spread and to identify concerning new variants. Two important determinants of variant spread are how frequently they arise within individuals and how likely they are to be transmitted. To characterize within-host diversity and transmission, we deep-sequenced 1313 clinical samples from the United Kingdom. SARS-CoV-2 infections are characterized by low levels of within-host diversity when viral loads are high and by a narrow bottleneck at transmission. Most variants are either lost or occasionally fixed at the point of transmission, with minimal persistence of shared diversity, patterns that are readily observable on the phylogenetic tree. Our results suggest that transmission-enhancing and/or immune-escape SARS-CoV-2 variants are likely to arise infrequently but could spread rapidly if successfully transmitted.

scholarly journals Evaluation of mobile real-time polymerase chain reaction tests for the detection of severe acute respiratory syndrome coronavirus 2

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Chukwunonso Onyilagha ◽  
Henna Mistry ◽  
Peter Marszal ◽  
Mathieu Pinette ◽  
Darwyn Kobasa ◽  
...  
Keyword(s):  
Severe Acute Respiratory Syndrome ◽  
Real Time ◽  
Point Of Care ◽  
Middle Eastern ◽  
Turnaround Time ◽  
Cross Reactivity ◽  
Clinical Samples ◽  
Diarrhea Virus ◽  
Highly Sensitive ◽  
Transmissible Gastroenteritis

AbstractThe coronavirus disease 2019 (COVID-19) pandemic, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), calls for prompt and accurate diagnosis and rapid turnaround time for test results to limit transmission. Here, we evaluated two independent molecular assays, the Biomeme SARS-CoV-2 test, and the Precision Biomonitoring TripleLock SARS-CoV-2 test on a field-deployable point-of-care real-time PCR instrument, Franklin three9, in combination with Biomeme M1 Sample Prep Cartridge Kit for RNA 2.0 (M1) manual extraction system for rapid, specific, and sensitive detection of SARS-COV-2 in cell culture, human, and animal clinical samples. The Biomeme SARS-CoV-2 assay, which simultaneously detects two viral targets, the orf1ab and S genes, and the Precision Biomonitoring TripleLock SARS-CoV-2 assay that targets the 5′ untranslated region (5′ UTR) and the envelope (E) gene of SARS-CoV-2 were highly sensitive and detected as low as 15 SARS-CoV-2 genome copies per reaction. In addition, the two assays were specific and showed no cross-reactivity with Middle Eastern respiratory syndrome coronavirus (MERS-CoV), infectious bronchitis virus (IBV), porcine epidemic diarrhea virus (PEDV), transmissible gastroenteritis (TGE) virus, and other common human respiratory viruses and bacterial pathogens. Also, both assays were highly reproducible across different operators and instruments. When used to test animal samples, both assays equally detected SARS-CoV-2 genetic materials in the swabs from SARS-CoV-2-infected hamsters. The M1 lysis buffer completely inactivated SARS-CoV-2 within 10 min at room temperature enabling safe handling of clinical samples. Collectively, these results show that the Biomeme and Precision Biomonitoring TripleLock SARS-CoV-2 mobile testing platforms could reliably and promptly detect SARS-CoV-2 in both human and animal clinical samples in approximately an hour and can be used in remote areas or health care settings not traditionally serviced by a microbiology laboratory.

Functional study of multidrug resistance with fluorescent dyes. Limits of the assay for low levels of resistance and application in clinical samples

1996 ◽  
Vol 106 (1) ◽  
pp. 59-68 ◽  
Author(s):  
Yvan Canitrot ◽  
Sylvie Lahmy ◽  
Jean-Jacques Buquen ◽  
Damien Canitrot ◽  
Dominique Lautier
Keyword(s):  
Multidrug Resistance ◽  
Fluorescent Dyes ◽  
Functional Study ◽  
Clinical Samples ◽  
Low Levels

Manipulation of Clinical Materials Using AC Electrokinetics

2010 ◽  
Author(s):  
Mandy L. Y. Sin ◽  
Pak Kin Wong
Keyword(s):  
Experimental Data ◽  
Sample Preparation ◽  
Theoretical Prediction ◽  
Fluid Motion ◽  
Lab On A Chip ◽  
Clinical Samples ◽  
Electrochemical Reactions ◽  
Ac Electrokinetics ◽  
Wide Range ◽  
Electrothermal Flow

AC electrokinetics is a promising approach for sample preparation and reaction enhancement in lab-on-a-chip devices. However, relative little has been done on the electrokinetic manipulation of physiological fluids and buffers with similar properties, such as conductivity. Herein, electrokinetic manipulation of fluids with a wide range of conductivities has been studied as a function of voltage and frequency. AC electrothermal flow is determined to dominate the fluid motion when the applied frequency of the AC potential is above 100 kHz. Interestingly, experimental data deviate from theoretical prediction for fluids with high conductivities (> 1 Sm−1). The deviation can be understood by voltage modulated electrochemical reactions and should be accounted for when manipulating clinical materials with high conductivities. The study will provide useful in sights in designing lab-on-a-chip devices for manipulating clinical samples in the future.

scholarly journals Differential Effects of IL-2 and IL-6 on the Development of Three Distinct Precursor T-Cell Populations in the Thymus

1990 ◽  
Vol 1 (2) ◽  
pp. 77-84 ◽  
Author(s):  
Naoko Nakano ◽  
Hitoshi Kikutani ◽  
Tadamitsu Kishimoto
Keyword(s):  
Bone Marrow ◽  
T Cell ◽  
Bone Marrow Cells ◽  
Cell Populations ◽  
High Dose ◽  
Transfer System ◽  
Cell Sorter ◽  
Lineage Markers ◽  
Low Levels ◽  
Normal Differentiation

Three distinct T-cell precursors: bone marrow cells that express low levels of the Thy-1 antigen but no lineage markers (Thy-1-lo/BM); CD4-, CD8-, and CD3-thymocytes that express low levels of the Thy-1 antigen (Thy-1-lo/Thym); and CD4-, CD8-, and CD3-thymocytes that express high levels of the Thy-1 antigen and the IL-2 Rαchain (Thy-1+/ IL2R+) were isolated by fluorescence-activated cell sorter (FACS). These three populations expanded with different kinetics in the thymus of irradiated recipient mice after intrathymic transfer. When a high dose of human recombinant IL-2 (r-IL-2) or human recombinant IL-6 (r-IL-6) was administered, r-IL-6 accelerated donor Thy-1+/IL2R+to differentiate, whereas r-IL-2 blocked normal differentiation and expansion of donor Thy-1-lo/Thym, but did not show any significant effect on donor Thy-1+/IL2R+. Neither r-IL-2 nor r-IL-6 worked directly on donor Thy-1-lo/BM in this transfer system.

scholarly journals Correction to: Optimization and validation of sample preparation for metagenomic sequencing of viruses in clinical samples

Microbiome ◽  
2017 ◽  
Vol 5 (1) ◽  
Author(s):  
Dagmara W. Lewandowska ◽  
Osvaldo Zagordi ◽  
Fabienne-Desirée Geissberger ◽  
Verena Kufner ◽  
Stefan Schmutz ◽  
...  
Keyword(s):  
Sample Preparation ◽  
Clinical Samples ◽  
Metagenomic Sequencing

scholarly journals Routine Determination of Ice Thickness for Cryo-EM Grids

2018 ◽  
Author(s):  
William J. Rice ◽  
Anchi Cheng ◽  
Alex J. Noble ◽  
Edward T. Eng ◽  
Laura Y. Kim ◽  
...  
Keyword(s):  
Data Collection ◽  
Sample Preparation ◽  
Quality Data ◽  
Ice Thickness ◽  
Limiting Factor ◽  
Test Cases ◽  
Slowing Down ◽  
Routine Monitoring ◽  
The Many

AbstractRecent advances in instrumentation and automation have made cryo-EM a popular method for producing near-atomic resolution structures of a variety of proteins and complexes. Sample preparation is still a limiting factor in collecting high quality data. Thickness of the vitreous ice in which the particles are embedded is one of the many variables that need to be optimized for collection of the highest quality data. Here we present two methods, using either an energy filter or scattering outside the objective aperture, to measure ice thickness for potentially every image collected. Unlike geometrical or tomographic methods, these can be implemented directly in the single particle collection workflow without interrupting or significantly slowing down data collection. We describe the methods as implemented into the Leginon/Appion data collection workflow, along with some examples from test cases. Routine monitoring of ice thickness should prove helpful for optimizing sample preparation, data collection, and data processing.

scholarly journals Multiplexed Detection of Sepsis Markers in Whole Blood using Nanocomposite Coated Electrochemical Sensors

2020 ◽  
Author(s):  
Uroš Zupančič ◽  
Pawan Jolly ◽  
Pedro Estrela ◽  
Despina Moschou ◽  
Donald E. Ingber
Keyword(s):  
Electrochemical Detection ◽  
Whole Blood ◽  
Electrochemical Sensors ◽  
Point Of Care ◽  
Sample Volume ◽  
Nanocomposite Coating ◽  
Cross Reactivity ◽  
Detection Methods ◽  
Clinical Samples ◽  
Undiluted Serum

ABSTRACTSepsis is a leading cause of mortality worldwide that is difficult to diagnose and manage because this requires simultaneous analysis of multiple biomarkers. Electrochemical detection methods could potentially provide a way to accurately quantify multiple sepsis biomarkers in a multiplexed manner as they have very low limits of detection and require minimal sensor instrumentation; however, affinity-based electrochemical sensors are usually hampered by biological fouling. Here we describe development of an electrochemical detection platform that enables detection of multiple sepsis biomarkers simultaneously by incorporating a recently developed nanocomposite coating composed of crosslinked bovine serum albumin containing a network of reduced graphene oxide nanoparticles that prevents biofouling. Using nanocomposite coated planar gold electrodes, we constructed a procalcitonin sensor and demonstrated sensitive PCT detection in undiluted serum and clinical samples, as well as excellent correlation with a conventional ELISA (adjusted r2 = 0.95). Sensors for two additional sepsis biomarkers — C-reactive protein and pathogen-associated molecular patterns — were developed on the same multiplexed platform and tested in whole blood. Due to the excellent antifouling properties of the nanocomposite coating, all three sensors exhibited specific responses within the clinically significant range without any cross-reactivity in the same channel with low sample volume. This platform enables sensitive simultaneous electrochemical detection of multiple analytes in human whole blood, which can be expanded further to any target analyte with an appropriate antibody pair or capturing probe, and thus, may offer a potentially valuable tool for development of clinical point-of-care diagnostics.GRAPHICAL ABSTRACT

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