Assay / ELISA / Binding: poor standard curve

1. Ranked causes

   1. Degraded or wrongly reconstituted standard stock (lot / storage)Fresh standard lot recovers the curve; samples unchanged.
   2. Standard dilution series pipetting errorPoints zigzag; remade dilutions restore a smooth sigmoid/linear segment.
   3. Incubation time/temperature drifted during standard platingEarly vs late standards diverge; timed incubations fix shape.
   4. Reader wavelength or blanking mis-set for the substrateCorrect λ restores expected OD span on the same plate.

2. Recovery steps

   1. Remake standards with fresh serial dilutions and new tips each stepPipetting error is the most common poor-curve cause.
   2. Reconstitute a new standard vial per kit instructions; avoid extra freeze-thawDegraded stock cannot be rescued by dilution technique alone.
   3. Verify reader wavelength and blank against kit substrate instructionsWrong λ compresses or warps the apparent curve.

3. Controls first

   • Do positive-control / kit-standard wells also fail to form a usable standard curve?If yes: Standards, incubation, or reader setup — not sample unknowns alone.

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