title: Feasibility of Singlet Analysis for Ligand Binding Assays: a Retrospective Examination of Data Generated Using the Gyrolab Platform.
abstract: There are many sources of analytical variability in ligand binding assays (LBA). One strategy to reduce variability has been duplicate analyses. With recent advances in LBA technologies, it is conceivable that singlet analysis is possible. We retrospectively evaluated singlet analysis using Gyrolab data. Relative precision of duplicates compared to singlets was evaluated using 60 datasets from toxicokinetic (TK) or pharmacokinetic (PK) studies which contained over 23,000 replicate pairs composed of standards, quality control (QC), and animal samples measured with 23 different bioanalytical assays. The comparison was first done with standard curve and QCs followed by PK parameters (i.e., Cmax and AUC). Statistical analyses were performed on combined duplicate versus singlets using a concordance correlation coefficient (CCC), a measurement used to assess agreement. Variance component analyses were conducted on PK estimates to assess the relative analytical and biological variability. Overall, 97.5% of replicate pairs had a %CV of <11% and 50% of the results had a %CV of ≤1.38%. There was no observable bias in concentration comparing the first replicate with the second (CCC of 0.99746 and accuracy value of 1). The comparison of AUC and Cmax showed no observable difference between singlet and duplicate (CCC for AUC and Cmax >0.99999). Analysis of variance indicated an AUC inter-subject variability 35.3-fold greater than replicate variability and 8.5-fold greater for Cmax. Running replicates from the same sample will not significantly reduce variation or change PK parameters. These analyses indicated the majority of variance was inter-subject and supported the use of a singlet strategy.
authors.affiliation: Pharmacokinetics, Dynamics, and Metabolism, Pfizer Worldwide R&D, MS 8220-2463, Eastern Point Rd, Groton, Connecticut, 06340, USA. Tracey.Clark@pfizer.com.
authors.forename: Tracey H
authors.initials: TH
authors.lastname: Clark
authors.affiliation: PharmaTherapeutics Clinical Research, Pfizer Worldwide R&D, Cambridge, Massachusetts, USA.
authors.forename: Phillip D
authors.initials: PD
authors.lastname: Yates
authors.affiliation: Pharmacokinetics, Dynamics, and Metabolism, Pfizer Worldwide R&D, La Jolla, California, USA.
authors.forename: Allison Given
authors.initials: AG
authors.lastname: Chunyk
authors.affiliation: Pharmacokinetics, Dynamics, and Metabolism, Pfizer Worldwide R&D, Andover, Massachusetts, USA.
authors.forename: Alison P
authors.initials: AP
authors.lastname: Joyce
authors.affiliation: Pharmacokinetics, Dynamics, and Metabolism, Pfizer Worldwide R&D, La Jolla, California, USA.
authors.forename: Aidong
authors.initials: A
authors.lastname: Wu
authors.affiliation: DMPK Oncology iScience, AstraZeneca R&D, Waltham, Massachusetts, USA.
authors.forename: Petar
authors.initials: P
authors.lastname: Pop-Damkov
authors.affiliation: Pharmacokinetics, Dynamics, and Metabolism, Pfizer Worldwide R&D, Andover, Massachusetts, USA.
authors.forename: Yiqun
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: Protein Engineering, Kanyos Bio, Cambridge, Massachusetts, USA.
authors.forename: Elizabeth A
authors.initials: EA
authors.lastname: Dreher
authors.affiliation: Pharmacokinetics, Dynamics, and Metabolism, Pfizer Worldwide R&D, MS 8220-2463, Eastern Point Rd, Groton, Connecticut, 06340, USA.
authors.forename: Laurie A
authors.initials: LA
authors.lastname: Tylaska
authors.affiliation: Pharmacokinetics, Dynamics, and Metabolism, Pfizer Worldwide R&D, MS 8220-2463, Eastern Point Rd, Groton, Connecticut, 06340, USA.
authors.forename: Jo-Ann A
authors.initials: JA
authors.lastname: Wentland
authors.affiliation: Pharmacokinetics, Dynamics, and Metabolism, Pfizer Worldwide R&D, MS 8220-2463, Eastern Point Rd, Groton, Connecticut, 06340, USA.
authors.forename: Kathleen B
authors.initials: KB
authors.lastname: Pelletier
authors.affiliation: Pharmacokinetics, Dynamics, and Metabolism, Pfizer Worldwide R&D, MS 8220-2463, Eastern Point Rd, Groton, Connecticut, 06340, USA.
authors.forename: Lindsay E
authors.initials: LE
authors.lastname: King
authors.affiliation: Pharmacokinetics, Dynamics, and Metabolism, Pfizer Worldwide R&D, La Jolla, California, USA.
authors.forename: Chad A
authors.initials: CA
authors.lastname: Ray
mesh_terms: D000818:Animals; D016208:Databases, Factual; D005240:Feasibility Studies; D000882:Haplorhini; D008024:Ligands; D051379:Mice; D004364:Pharmaceutical Preparations / Q000032:analysis / Q000378:metabolism*; D011485:Protein Binding / Q000502:physiology; D051381:Rats; D012189:Retrospective Studies; D013223:Statistics as Topic / Q000379:methods*
chemical_list: D008024:Ligands; D004364:Pharmaceutical Preparations
keywords: Gyrolab; ligand binding assay; pharmacokinetics; singlet; toxicokinetics