Anti nkp46 pe mab pch101

Assessment of NK and Treg cells was done using both flow cytometry (Cytomics FC500 flow cytometer) and ELISA. Monoclonal antibodies (mab) were purchased from (eBioscience, San Diego, CA), anti-CD3-FITC, anti-CD4-PE and anti-CD25-FITC, or anti-CD56-PE. The frequency of each cell subset was calculated based on the percentage of positive cells in the total lymphocyte gate. For intracellular staining, cells were first labelled with relative marker, fixed, and permeabilized using a fix/perm kit according to the manufacturer’s instructions and then labelled with anti-FOXP3-PE or anti-NKp46-PE mab (PCH101; eBioscience).
NKp46 and FOXP3 were measured also in cell lysate by sandwich ELISA for in vitro quantitative measurement (mybiosource.com/prods/ELISA-Kit/mouse/forkhead-box-P3, NKp46). Following cell culture of mouse spleen cells, repeated freezing and thawing cycles were done to damage the cells and release the inside components of the cell, followed by centrifugation at 2000–3000 RPM for approximately 20 min. The supernatants were collected carefully and centrifuged again when sedimentation occurred during storage.

Assessment of NK and Treg cells was done using both flow cytometry (Cytomics FC500 flow cytometer) and ELISA. Monoclonal antibodies (mab) were purchased from (eBioscience, San Diego, CA), anti-CD3-FITC, anti-CD4-PE and anti-CD25-FITC or anti-CD56-PE. The frequency of each cell subset was calculated based on the percentage of positive cells in the total lymphocyte gate. For intracellular staining, cells were first labeled with relative marker, fixed, and permeabilized using a fix/perm kit according to the manufacturer’s instructions and then labeled with anti-FOXP3-PE or anti-NKp46-PE mab (PCH101; eBioscience).
NKp46 and Foxp3 were measured also in cell lysate by sandwich ELISA for in vitro quantitative measurement (mybiosource.com/prods/ELISA-Kit/mouse/forkhead-box-P3, NKp46). Following cell culture of mouse spleen Cells, repeated freezing and thawing cycles were done to damage cells and let out the inside components of the cell then Centrifugation at 2000-3000 RPM for approximately 20 minutes was done. The supernatants were collected carefully and centrifuged again when sediments occurred during storage.