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Curves and Constants: Kd, Ki, IC50 and EC50

Now we make it quantitative. Dose–response curves turn binding and effect into numbers, and four constants — Kd, Ki, IC50 and EC50 — let you compare drugs honestly. Here's what each one really means and how not to mix them up.

The dose–response curve

Give more drug, get more effect — until you can't. Plot the response against drug concentration and you get the dose–response curve. Plotted against the logarithm of concentration it takes a tidy S-shape: a flat low-dose foot, a steep rise in the middle, and a plateau at the top where the system is maxed out. Two features of this curve carry almost all the information: how far left it sits (small doses already work = potent) and how high it plateaus (the size of the biggest possible effect).

Kd and Ki: the binding constants

The dissociation constant Kd is the clean definition of affinity: it's the concentration of drug at which half the target copies are occupied. So a Kd of 10 nM means 10 nM of drug fills half the pockets. The logic is simple — if it takes very little drug to fill half, the drug must grab tightly, so a small Kd means high affinity. Kd is measured directly in a binding experiment, with no biological response required.

The inhibition constant Ki is the same idea for a blocker: the affinity of an inhibitor for its target, measured from how well it competes a known ligand off. Like Kd, smaller Ki = tighter binding. The beauty of Ki is that, unlike a raw IC50, it's corrected for assay conditions, so Ki values can be compared across different experiments — which is why medicinal chemists love to report it.

IC50 and EC50: the working numbers

Two more numbers come straight off the dose–response curve. The EC50 is the concentration of an activator that produces half of its maximum effect — it's the midpoint of an agonist's curve, and a direct read of potency. The IC50 is the mirror image for an inhibitor: the concentration that knocks the signal down to half. Both are simply "the dose that gets you halfway," one going up, one going down.

ONE-LINE DEFINITIONS  (smaller value = more potent / higher affinity)

  Kd    half the TARGETS are occupied         (binding only, no effect)
  Ki    inhibitor's true affinity             (assay-corrected, portable)
  EC50  half the maximum EFFECT (activator)    (functional potency, up)
  IC50  half the signal remains (inhibitor)    (functional potency, down)

Rule of thumb: Kd/Ki are about BINDING.
               EC50/IC50 are about the EFFECT you see in the assay.
The four constants at a glance — binding constants vs functional (effect-based) constants.

The key warning: an IC50 is not a fixed property of the drug. Because most blockers act by competition, the measured IC50 slides depending on how much substrate or agonist is present in that particular assay — more competitor, higher IC50. That's exactly why we convert to Ki when we want a number we can trust across experiments, and why a careful Schild analysis is used to pin down a competitive antagonist's true affinity. Treat EC50/IC50 as honest readings of *this assay*, and Kd/Ki as the deeper, portable truth about *binding*.