Section5:Scintillation Proximity Assays (SPA)

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Concept

SPA assays do not require a separation of free and bound radioligand and therefore are amenable to screening applications. A diagram for a standard receptor binding SPA is shown below for a 125I radioligand.

General Steps for an SPA Assay:

1. Add and incubate test compound, radioligand, receptor and SPA beads in a plate (in some cases, the SPA beads are added at a later time point).
2. Count plates in microplate scintillation counter. The appropriate settling time needs to be determined experimentally.

Advantages		Disadvantages
Non-separation method No scintillation cocktail required Reduced liquid radioactive waste Reduced handling steps (add, incubate, read) Multiple bead types (WGA, PEI-coated, etc.)		More expensive - requires license Lower counting efficiency Primarily for 3H and 125I (33P, 35S possible) Non-proximity effects Quenching by colored compounds Difficult to perform kinetic experiments Bead settling effects

Many of the advantages and disadvantages are addressed in the following sections.