title: Stepping up ELISpot: Multi-Level Analysis in FluoroSpot Assays.
abstract: ELISpot is one of the most commonly used immune monitoring assays, which allows the functional assessment of the immune system at the single cell level. With its outstanding sensitivity and ease of performance, the assay has recently advanced from the mere single function cell analysis to multifunctional analysis by implementing detection reagents that are labeled with fluorophores (FluoroSpot), allowing the detection of secretion patterns of two or more analytes in a single well. However, the automated evaluation of such assays presents various challenges for image analysis. Here we dissect the technical and methodological requirements for a reliable analysis of FluoroSpot assays, introduce important quality control measures and provide advice for proper interpretation of results obtained by automated imaging systems.
authors.affiliation: ZellNet Consulting, Inc., 555 North Avenue, Suite 25-S, Fort Lee, NJ 07024, USA. sylvia@zellnet.com.
authors.forename: Sylvia
authors.initials: S
authors.lastname: Janetzki
authors.affiliation: Autoimmun Diagnostika GmbH, Ebinger Strasse 4, D-72479 Strassberg, Germany. rueger@aid-diagnostika.com.
authors.forename: Markus
authors.initials: M
authors.lastname: Rueger
authors.affiliation: Mabtech AB, Box 1233, SE-131 28 Nacka Strand, Sweden. Tomas.Dillenbeck@mabtech.com.
authors.forename: Tomas
authors.initials: T
authors.lastname: Dillenbeck