X-Nico

Strictly standardized mean difference

In an HTS assay, one primary goal is to select compounds with a desired size of inhibition or activation effect.

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can display both average fold change and SSMD for all test compounds in an assay and help to integrate both of them to select hits in HTS experiments

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In a confirmatory or primary screen with replicates, for the i-th test compound with $n$ replicates, we calculate the paired difference between the measured value (usually on the log scale) of the compound and the median value of a negative control in a plate, then obtain the mean $\backslash bar\{d\}\; i$ and variance $s\; i^2$ of the paired difference across replicates.

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In a primary screen without replicates, assuming the measured value (usually on the log scale) in a well for a tested compound is $X\; i$ and the negative reference in that plate has sample size $n\; N$, sample mean $\backslash bar\{X\}\; N$, median $\backslash tilde\{X\}\; N$, standard deviation $s\; N$ and median absolute deviation $\backslash tilde\{s\}\; N$, the SSMD for this compound is estimated as

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For hit selection, the size of effects of a compound (i.e., a small molecule or an siRNA) is represented by the magnitude of difference between the compound and a negative reference.

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The size of the compound effect is represented by the magnitude of difference between a test compound and a negative reference group with no specific inhibition/activation effects.

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The other strategy is to test whether a compound has effects strong enough to reach a pre-set level.

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A compound with a desired size of effects in an HTS screen is called a hit.

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An important QC characteristic in an HTS assay is how much the positive controls, test compounds, and negative controls differ from one another in the assay.

see also