M2 Macrophage Suppression Assay

Macrophages - the most abundant antigen-presenting cells in the periphery - represent by themselves a wide group of functionally-different subpopulations involved in several immunological functions, including inflammatory responses and tissue homeostasis maintain & repair. Unlike M1 macrophages which promote a pro-inflammatory response, alternatively activated M2 macrophages, with diametrically opposed functions, trigger inflammation resolution and suppress T cell activation via several mechanisms including immune checkpoint engagement (e.g. PDL1), release of anti-inflammatory cytokines such as IL-10 and TGFb, and metabolic activities which promote essential amino acid depletion (e.g. Arginine). – which characteristics that they share with the so-called tumor-associated macrophages (TAMs).
 
Our newly validated M2 suppression assay based on i) the co-culture of autologous monocyte-derived M2 macrophages and activated CD4+ T cells (or PBMCs) and on ii) the quantitation of IFNg levels as surrogate of T cell activation, is specifically designed to assess new immunotherapeutics for their modulatory activity on the phenotype and function of M2 macrophages. Candidate compounds can thus be evaluated as single agents or in combinatorial treatments, for their potential to repolarize / switch M2 macrophages and to antagonize M2-mediated T cell suppression.

Download our M2 Macrophage Suppression Assay white paper         

Assay principle





 
  • HTRF-based quantification of key cytokines released by macrophages, e.g. IL-6, IL-10, IL-12 as to evaluate the potency of candidate compounds to revert M2 phenotype.
  • Human autologous M2 suppression assay, based on a co-culture of monocyte-derived M2 macrophages with activated T cells (or PBMCs) and measurement of IFNγ, as one of the most representative surrogates of T cell activation.
  • Robust functional assay, microplate format tailored for screening campaigns
  • Complementary analysis can be implemented to decipher MOA (e.g. activation markers, signaling pathways, etc)

Illustrative data

White paper - M2 Macrophage Suppression Assay
Macrophages - the most abundant antigen-presenting cells in the periphery - represent by themselves a wide group of functionally-different subpopulations involved in innate immunity, including inflammatory responses and tissue homeostasis maintain & repair. Unlike M1 macrophages (pro-inflammatory) which promote Th1 cell immunity, alternatively activated M2 macrophages, with diametrically opposed functions, trigger inflammation resolution and suppress T cell activation via several mechanisms including release of anti-inflammatory cytokines e.g. IL10, for instance – which characteristics that they share with the so-called tumor-associated macrophages (TAMs).

 

Macrophages - the most abundant antigen-presenting cells in the periphery - represent by themselves ...

Monocyte-derived macrophage polarization
While displaying an expected variation from a donor to another one, the cytokine macrophage profiles are maintained - characterizing M1 and M2 populations. Isolated peripheral blood monocytes from two donors are subjmitted to M1 or M2 macrophage polarization process. Cytokine levels released in the supernatants are measured using a specific HTRF-based specific kit. M1 macrophages display a IL6High / IL10Low profile while M2 display a IL6Low / IL10High  / IL12Low  profile.

While displaying an expected variation from a donor to another one, the cytokine macrophage ...

M2 macrophage suppression of T cell activation
M2 macrophage suppression results in a decrease of IFNγ surrogate of T cell activation. Monocyte-derived M2 macrophages are mixed with CD4+ T cells each isolated from the same donor and incubated or not with αCD3. IFNγ levels released in the supernatants are measured after 72h using a specific HTRF-based specific kit.

M2 macrophage suppression results in a decrease of IFNγ surrogate of T cell activation. ...