Introduction

The new method aims at progressing the study of MPS released by NK cells, allowing effectively evaluation of phenotype and function and in a differential way. The release of MPS by the two NK cell subsets CD56^dim and CD56^bright is highlighted. This aspect has to be taken into consideration when analyzing MPS present in biological fluids.

Technical features

Therefore, the subject of the present invention are microparticles released by natural killer cells characterized by expressing the NK surface markers (i.e. NKp30 and NKp46) and by contain perforins and cytokines such as IFNγ and TNFα . The new method of MPS analysis that consists of the following steps:

• Isolated NK cells;
• Stimulate NK cells in the presence or absence of target cytokines and / or cell lines and / or monoclonal antibodies specific to natural cytotoxicity receptors (NCRs) including the maximal with PMA and Ionomicina;
• Incubate the cells in plates/tubes in the appropriate culture medium 37 ° C 5% CO2 for specific times depending on the stimulation (4h, 16h, 72h);
• After the time defined according to the stimulation collect the supernatant taking care not to collect the cells and proceed with the intracytoplasmic staining without carrying out any intermediate washing in order to analyze the content and surface for the MPs of interest and acquire the flow cytometer.

Possible Applications

• MPS analysis in biological fluids  derived from healthy neoplastic donors or during infections, autoimmunity;
• Use as a natural biological drug/immunotherapy;
• Monitoring marker for acute/chronic viral infections, may be used also for diagnostic and predictive purposes;
• Tool for the control of tumor cells in conditions where standard drug therapy may be less effective/indicated.

Advantages

• Absence of any washing or dilution of the supernatant;
• Phenotypical and functional analysis of MPS NK cell derived;
• Evaluation of the MPS cytokines and perforins content.