CD160 Antibody with Great Resolution

Results Summary

CD160 is a co-inhibitory molecule which we found to be induced upon infection with PbA in mice. The molecule is sometimes difficult to detect and accordingly, a bright fluorochrome (such as PE) is advantageous. We were able to detect a strong signal using this fluorochrome/ clone combination, enabling detection of distinct CD160 positive and negative population. We found the strongest signal on CD8+ T cells, but some CD160 expression was also detected on CD4+ T cells and NK cells upon infection. I used a titer of 1:25 for the staining shown here, which was required for a signal in a large panel (41 colors). However, more economical titers of 1:50 or 1:100 also gave sufficient separation in smaller panels. The fluorochrome/ antibody withstood fixation (see below) without a problem, resulting in the observed stainings. In the following, you will find my staining protocol in short. Incubation steps were performed at 4°C in the dark. 1.Count and adjust splenocytes to 1.5E6 cells, plate in a 96-well plate. 2.If required, stain cells with viability dye. Wash. 3. Stain with extracellular master mix for 20 min. Wash 2x. 4. Fixate/ Permeabilize with buffer of choice, depending on application. I used ThermoFisher FoxP3 fixation/permeabilization kit for 20 min. Wash with perm/ Wash 2x. 5. Perform intracellular staining for 20 min. Wash 2x. 6.Resuspend in FACS buffer and acquire as soon as possible. Here, data was acquired on a 5L Cytek Aurora.