Is the measured analyte active?Should the method quantify the parent antiviral, an active metabolite, a phosphorylated intracellular form, or a prodrug-derived product?
Is intracellular exposure central?Do cell lysates, intracellular extracts, or phosphorylated metabolite measurements better represent antiviral activity than plasma parent-drug levels?
Is prodrug conversion involved?Should the workflow separate the administered compound, conversion intermediates, active form, and inactive transformation products?
Is this a combination regimen?Does the study require simultaneous quantification of multiple antivirals, boosters, or parent–metabolite pairs?
Are DDI or transporter effects relevant?Could CYP metabolism, transporter-associated disposition, or pharmacokinetic boosting alter antiviral exposure?
Which response markers matter?Should exposure be paired with viral load, replication markers, IFN signals, cytokines, immune activation, or cell-population readouts?