7 - Drug binding: binding assays

What are some common radiolabels?

3H (tritium) / 14C / 125I

Why do we need to detect binding?

To quantify how much of a drug has bound in a tissue sample

What is the normal 3 steps in radioligand binding assay?

Incubation / separation / quantification

What happens during incubation in radioligand binding assay?

Binding takes place

What happens during separation in radioligand binding assay?

Removal of any excess unbound radioligand

How do you quantify with beta emitting nuclides? (state the method)

Use liquid scintillation counting (LSC)

Which of the common radiolabel is beta emitting?

3H

What is the basic principle of LSC?

Add solution of compounds that emit light when struck by radiation (scintillant) / then measure amount of light produced by sample

What are scintillant?

Compounds that emit light when struck by radiation.

What does LSC stand for?

Liquid scintillation counting

How would you quantify gamma emitters?

Using gamma counter

Which of the common radiolabel that is a gamma emitter?

125I

How does gamma counter work?

Tube with sample is placed next to a reusable solid scintillant / gamma ray from sample pass through and strike the solid / light emits

What does NSB stand for?

Non-specific binding

What does NSB represent?

A v.large unsaturable pool of low affinity binding sites

What is the equation for specific binding?

Specific binding = total binding - NSB

What is saturation binding assay generally used for?

Establish affinity (KD) and receptor density (Bmax) for radioligand in tissue of interest