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User Versus Software-Dependent Variability of ELISPOT Counts

In each human donor’s peripheral blood mononuclear cell (PBMC), there is a defined number of T-cells specific for any given antigen. A major goal of immune monitoring with enzyme-linked immunospot (ELISPOT) is to measure this number accurately and reproducibly between different laboratories.

In ELISPOT assays, cytokine spots produced by antigen-specific T-cells show a broad spectrum of sizes and densities over a variable background. Therefore, even experienced investigators are likely to come up with different spot counts when subjectively judging the minimal spot size / density to be counted and the maximal spot size for the cut off between single cell-derived spots versus those created by cell clusters.

This study aims to find out whether statistics-based automated gating can harmonise spot counts obtained in different laboratories.

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