Critical reagents in flow cytometry, instrumentation and application in drug discovery development

Bioanalysis ◽  
2021 ◽  
Author(s):  
Vellalore N Kakkanaiah ◽  
Katie Matys ◽  
Patrick Bennett
Keyword(s):  
Flow Cytometry ◽  
Drug Discovery ◽  
Drug Development ◽  
Ligand Binding ◽  
Specific Interaction ◽  
Flow Cytometer ◽  
Analysis Tool ◽  
Binding Assays ◽  
Cellular Analysis ◽  
Main Components

Flow cytometer is a powerful cellular analysis tool consists of three main components; fluidics, optics and electronics. Flow cytometry methods have been used in all stages of drug development as like ligand binding assays (LBA). Both LBA and flow cytometry methods require specific interaction between the critical reagents and the analytes. Antibodies and their conjugates, viable dyes and permeabilizing buffer are the main critical reagents in flow cytometry methods. Similarly, antibodies, engineered proteins and their conjugates are the main critical reagents in LBA. The main difference between the two methods is the lack of true reference standards for flow cytometry cellular analysis.

Laboratory automation of high-quality and efficient ligand-binding assays for biotherapeutic drug development

Bioanalysis ◽  
2013 ◽  
Vol 5 (13) ◽  
pp. 1635-1648 ◽  
Author(s):  
Jin Wang ◽  
Vimal Patel ◽  
Daniel Burns ◽  
John Laycock ◽  
Kinnari Pandya ◽  
...  
Keyword(s):  
Drug Development ◽  
Ligand Binding ◽  
Laboratory Automation ◽  
Binding Assays ◽  
High Quality

Ligand Binding Assays in Drug Development

2013 ◽  
pp. 647-657
Author(s):  
Jeffrey M. Sailstad ◽  
Ronald R. Bowsher ◽  
Omar F. Laterza ◽  
William Nowatzke
Keyword(s):  
Drug Development ◽  
Ligand Binding ◽  
Binding Assays

HTPS Flow Cytometry: A Novel Platform for Automated High Throughput Drug Discovery and Characterization

2001 ◽  
Vol 6 (2) ◽  
pp. 83-90 ◽  
Author(s):  
Bruce S. Edwards ◽  
Frederick W. Kuckuck ◽  
Erick R. Prossnitz ◽  
John T. Ransom ◽  
Larry A. Sklar
Keyword(s):  
Flow Cytometry ◽  
Drug Discovery ◽  
Intracellular Calcium ◽  
High Throughput ◽  
Combinatorial Libraries ◽  
Flow Cytometer ◽  
Automated System ◽  
Primary Screening ◽  
Formyl Peptide ◽  
Multifactorial Approach

The flow cytometer is unique among biomedical analysis instruments because it makes simultaneous and multiple optical measurements on individual cells or particles at high rates. High throughput flow cytometry represents a potentially important multifactorial approach for screening large combinatorial libraries of compounds. Limiting this approach has been the availability of instrumentation and methods in flow cytometry for automated sample handling on the scale required for drug discovery applications. Here, we describe an automated system in which a novel patented fluidics-based pharmacology platform, the HTPS (High Throughput Pharmacological System), is coupled to a flow cytometer using a recently described plug flow-coupling valve technology. Individual samples are aspirated sequentially from microplate wells and delivered to a flow cytometer for rapid multiparametric analysis. For primary screening to detect and quantify cell fluorescence in endpoint assays, a high-speed no-wash protocol enabled processing of 9-10 cell samples/min from 96-well microplates. In an alternate primary screening format, soluble receptor ligands were sampled from microplate wells at rates of 3-4 samples/minute and successfully assessed for the ability to elicit intracellular calcium responses. Experiments with fluorescent beads validated the accurate automated production by the HTPS of exponential and linear gradients of soluble compounds. This feature enabled rapid (2- to 3-min) characterization of the intracellular calcium dose response of myeloid cells to formyl peptide as well as the quantitative relationship between formyl peptide receptor occupancy and cell response. HTPS flow cytometry thus represents a powerful high throughput multifactorial approach to increase the efficiency with which novel bioresponse-modifying drugs may be identified and characterized.

Data quality in drug discovery: the role of analytical performance in ligand binding assays

2015 ◽  
Vol 29 (9) ◽  
pp. 847-865 ◽  
Author(s):  
Hermann Wätzig ◽  
Imke Oltmann-Norden ◽  
Franziska Steinicke ◽  
Hassan A. Alhazmi ◽  
Markus Nachbar ◽  
...  
Keyword(s):  
Drug Discovery ◽  
Ligand Binding ◽  
Data Quality ◽  
Analytical Performance ◽  
Binding Assays

scholarly journals Recommendations for Use and Fit-for-Purpose Validation of Biomarker Multiplex Ligand Binding Assays in Drug Development

2015 ◽  
Vol 18 (1) ◽  
pp. 1-14 ◽  
Author(s):  
Darshana Jani ◽  
John Allinson ◽  
Flora Berisha ◽  
Kyra J. Cowan ◽  
Viswanath Devanarayan ◽  
...  
Keyword(s):  
Drug Development ◽  
Ligand Binding ◽  
Binding Assays ◽  
Fit For Purpose

Development of Ligand-Binding Assays for Drug Development Support

2009 ◽  
pp. 39-79 ◽  
Author(s):  
Masood N. Khan ◽  
Proveen D. Dass ◽  
John H. Leete ◽  
Richard F. Schuman ◽  
Michele Gunsior ◽  
...  
Keyword(s):  
Drug Development ◽  
Ligand Binding ◽  
Binding Assays ◽  
Development Support

Flow cytometry and pharmacokinetics

Bioanalysis ◽  
2021 ◽  
Author(s):  
Kevin Lang ◽  
Katie Matys ◽  
Patrick Bennett ◽  
Vellalore N Kakkanaiah
Keyword(s):  
Flow Cytometry ◽  
Method Development ◽  
Positive Control ◽  
Analysis Tool ◽  
Cell Therapies ◽  
Multiparametric Flow Cytometry ◽  
Cellular Kinetics ◽  
Cellular Analysis ◽  
Sample Stability ◽  
Car T

Multiparametric flow cytometry is a powerful cellular analysis tool used in various stages of drug development. In adoptive cell therapies, the flow cytometry methods are used for the evaluation of advanced cellular products during manufacturing and to monitor cellular kinetics after infusion. In this report, we discussed the bioanalytical method development challenges to monitor cellular kinetics in CAR-T cell therapies. These method development challenges include procuring positive control samples for the development of the method, flow cytometry panel design, LLOQ, prestain sample stability, staining reagents and data analysis.

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