Immune Activity Screening Package for novel and established compounds

The aim of FOCUS Immunology’s “Immune Screening” testing package is to find out whether novel or known compounds affect basic immune functions; this may alert early on in development to potential side effects in clinical studies and on the other hand it may pave the way towards definition of new indications.

To this end FOCUS Immunology provides a selection of assays comprised in the “Immune Screening Package”. This package is purposefully set to address the “major” aspects of immune cell activation but this package may at any time be complemented by more refined assays from the the “Specific Immune System Package”, the “Innate Immune System Package”, or the “Biologicals (of higher risk) Package”.

Specifically, the following assays are offered to screen for possible immuno-modulatory effects of known or to be developed compounds:

Cell proliferation: Upon stimulation, cell proliferation is determined either in bulk cultures by metabolic assays or by measuring DNA synthesis (BrdU incorporation). Alternatively, cell proliferation can be determined on the single cell level by using a fluorescence-based assay (CFSE assay).

Cytokine release: Upon stimulation cytokine production can be measured using either of three assays: (a) ELISA, to determine the total amount of cytokine(s) secreted into the cell culture medium (link to List), (b) EliSpot, to determine the frequency of cells secreting (a) particular cytokine(s) and (c) Intracellular Cytokine Staining (ICS), to measure multiple cytokines and correlate their expression on the single cell level with lineage markers.

Expression of activation markers: Immune cells are not only characterized by cell-type-specific markers such as CD3 for T cells, CD19 or CD20 for B cells or CD14 for monocytes, but also by a continuously growing panel of CD markers to distinguish subpopulations. Examples are CD4 and CD8 for helper and cytotoxic T cells and CD4 + CD25 for regulatory T cells. Additionally, cells of the lymphoid lineage acquire specific activation markers upon stimulation, which are not expressed on resting cells. Examples for activation markers of T cells are CD69 and CD86.

The combination of lineage and activation markers in one assay (“multi-color flow cytometry”) allows analyzing the functional states of immune cells on the single cell level. This provides refined information on the effects test-compounds on the functionality of immune cell subpopulations.

FOCUS Immunology operates a dual-laser FACSCalibur from BD Biosciences to analyze up to four different markers in parallel allowing a detailed analysis of cell surface and/or intracellular markers.