Z' Does Not Need to Be > 0.5.

Area of Science:

• Drug discovery and high-throughput screening methodologies.
• Assay development and validation in pharmaceutical research.

Background:

• The Z' factor is a critical metric for high-throughput screening (HTS) assay validation.
• A common requirement for HTS assays is a Z' factor greater than 0.5.
• This stringent threshold may hinder the adoption of potentially valuable assays.

Purpose of the Study:

• To critically evaluate the widely accepted Z' > 0.5 threshold for HTS assays.
• To determine the relationship between Z' factor values and actual assay performance.
• To propose a more flexible approach to setting assay thresholds.

Main Methods:

• Utilized power analysis to assess assay performance.
• Employed a novel numerical simulation approach to model Z' factor behavior.
• Investigated assay performance across various experimental conditions.

Main Results:

• Assays with Z' > 0.5 generally exhibit superior performance.
• However, assays with Z' < 0.5 can successfully identify active compounds with minimal false positives when appropriate thresholds are set.
• The study demonstrates that lower Z' values are not necessarily indicative of unusable assays.

Conclusions:

• The strict requirement of Z' > 0.5 is not universally supported and may limit drug discovery efforts.
• Researchers can implement a justified, lower Z' threshold (< 0.5) for assays, particularly when justified by target importance and alternative assay limitations.
• A method is provided to help researchers determine appropriate, assay-specific thresholds, optimizing the use of phenotypic and cell-based screens.