If you do this, you won't get the most accurate values for Bmax and Kd. They are especially useful when you want to show a change in Bmax or Kd.ĭon't use the slope and intercept of a linear regression line to determine values for Bmax and Kd. Scatchard plots are often shown as insets to the saturation binding curves. The human retina and visual cortex evolved to detect edges (straight lines), not rectangular hyperbolas, and so it can help to display data this way. If you create a Scatcahrd plot, use it only to display your data. the ratio of specific binding to the concentration of free radioligand. One way to do this is with a Scatchard plot, which plots specific binding vs. Create a Scatchard plotīefore nonlinear regression was available, investigators had to transform curved data into straight lines, so they could analyze with linear regression.

It is the radioligand concentration needed to achieve a half-maximum binding at equilibrium. Kd is the equilibrium dissociation constant, in the same units as X. It is the specific binding extrapolated to very high concentrations of radioligand, and so its value is almost always higher than any specific binding measured in your experiment. Modelīmax is the maximum specific binding in the same units as Y. Then use the Remove Baseline analysis to subtract column B from column A, creating a new results table with the specific binding.įrom the table of specific binding, click Analyze, choose nonlinear regression, choose the panel of Saturation Binding equations, and choose One site specific binding. If you have several experimental conditions, place the first into column A, the second into column B, etc.Īn alternative approach would be to enter total binding into column A, and nonspecific into column B. Enter radioligand concentration into X, and specific binding into Y.

Globally analyze the total and nonspecific binding at one time.

Analyze the total binding only, inferring the amount of nonspecific binding from the shape of the total binding curve.

Subtract off the nonspecific, and analyze only the specific binding.

There are three approaches to dealing with nonspecific binding. The ligand binds not only to receptors sites, but also to nonspecific sites.

The goal is to determine the Kd (ligand concentration that binds to half the receptor sites at equilibrium) and Bmax (maximum number of binding sites). In a saturation binding experiment, you vary the concentration of radioligand and measure binding.

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