ELISA
Overview
Enzyme-linked immunosorbent assay — the workhorse for quantifying a soluble analyte (a cytokine in a culture supernatant, an antibody in serum) against a standard curve. It is the method behind most absolute concentration figures in this wiki that are not flow cytometry percentages.
This page currently documents ELISA as used for cytokine quantification in culture supernatants. It will grow as sources using serological ELISA are ingested; note that several already-ingested sources use ELISA for dengue serology without it having been documented here — see Limitations.
Background context (not sourced to an ingested paper): sandwich ELISA captures analyte on an immobilised antibody, detects with a second enzyme-conjugated antibody against a different epitope, and converts substrate turnover to concentration by interpolation from a standard curve of known concentrations. Accuracy depends on the sample falling within the curve, which is why dilution series are run.
Key Points from Literature
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Cytokine panel from a single supernatant. Human IFN-α (Bender MedSystems module set), IFN-β (BioSource International) and IL-8 (R&D Systems Duoset) were measured from the same cell-free culture supernatants, allowing two distinct TLR7 signalling arms to be read from one experiment (see Wang2006 - Flavivirus Activation of pDCs and TLR7 Signaling, in vitro).
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Basic quality practice worth recording: samples assayed in duplicate, and at dilutions chosen so that readings fall within the range of the standard curve (see Wang2006 - Flavivirus Activation of pDCs and TLR7 Signaling). A value read off the flat top or bottom of a sigmoid curve is not a measurement.
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Supernatant handling. Culture supernatants collected at a defined timepoint (20 h for primary Plasmacytoid Dendritic Cell cultures, 40 h for the TLR Reporter Cell Assay), centrifuged cell-free, aliquoted and stored at −20 °C before assay (see Wang2006 - Flavivirus Activation of pDCs and TLR7 Signaling).
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The dynamic range is wide and matters for interpretation. Reported IFN-α values in one experiment spanned 948 pg/ml (D2V, MOI 2.5) to 64,407 pg/ml (56 °C-inactivated influenza) — nearly two orders of magnitude, across which no saturation is apparent, which is part of why the source could rank agonist potency at all (see Wang2006 - Flavivirus Activation of pDCs and TLR7 Signaling).
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⚠ Units differ between analytes and are not interchangeable. In the same figure, IFN-α is reported in pg/ml and IFN-β in IU/ml (see Wang2006 - Flavivirus Activation of pDCs and TLR7 Signaling, Fig 5) — mass units versus bioactivity units, with no fixed conversion between them.
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[2026-08-29] Serum autoantibody quantification alongside a cohort-scale B cell panel. 9G4⁺ (VH4.34-idiotype) IgG, IgM and IgA by in-house ELISA; anti-dsDNA and anti-chromatin IgG by commercial kit (QUANTA Lite, INOVA Diagnostics); anti-RNA IgG by ELISA — run on the same subjects as an 8-marker B cell panel, which is what allowed composition and serology to be associated patient-by-patient (see Jenks2021 - B Cell Subset Composition in Cutaneous Lupus, n=207 patients + 39 additional SLE + 69 healthy controls, cross-sectional). ⚠ The autoantibody findings are out of scope for this wiki under the standing [2026-05-02] 9G4 decision and are recorded on the source page as background only; the method is noted here because the paired design — serology and immunophenotype on one blood draw — is the template the dengue study would need to link cells to antibody output.
Contradictions & Debates
Percentage-of-control reporting hides absolute magnitude. Inhibitor experiments are commonly plotted as ”% of stimulus alone,” which is the right normalisation for a blocking effect but discards the information that the stimuli differ 30-fold in absolute output. Wang2006 - Flavivirus Activation of pDCs and TLR7 Signaling handles this well by stating the absolute mean pg/ml for every stimulus in the figure legend; where a source does not, the wiki should not infer relative potency from percentage plots.
Coverage gap in this wiki. Several previously ingested dengue sources used ELISA for serology (IgM/IgG capture assays for primary-versus-secondary infection assignment, isotype quantification) without this page existing to record it. Those pages were not retro-fitted during this ingest. Treat the sources: count here as an undercount of actual ELISA use across the wiki until a lint pass reconciles it.
Related Pages
ELISpot, TLR Reporter Cell Assay, Plasmacytoid Dendritic Cell, Type I Interferon, Serum Proteomics, IgG, IgM