Supplementary MaterialsSupplementary Number 1: Binding of KIR2DL3-Fc to GKL, GAL, YF, HIV-1 and HCV peptide pulsed 221-TAPko-HLA-C*03:04 cells. derived from YFV-vaccinated individuals, or HIV-1- or HCV-infected individuals revealed that the YFV/HLA-C*03:04-NS2A4?13-tetramer bound to a larger proportion of KIR2DL2/3+ NK cells compared to HIV-1/HLA-C*03:04-Gag296?304- or HCV/HLA-C*03:04-Core136?144-tetramers. The YFV/HLA-C*03:04-NS2A4?13-tetramer also exhibited a stronger avidity to KIR2DL2/3 compared to the other tested tetramers. The proportional frequencies of KIR2DL2/3+ NK cells binding to the three tested HLA-C*03:04 tetramers were identical between YFV-vaccinated individuals or HIV-1- or HCV-infected individuals, and remained stable following YFV vaccination. These data show constant hierarchies in the rate of recurrence of major KIR2DL2/3+ NK cells binding HLA-C*03:04/peptide complexes which were dependant on the HLA-C-presented peptide rather than modulated from the root viral disease or vaccination. stain major human being NK cells of YFV-vaccinated (28 times post vaccination), HIV-1-contaminated or HCV-infected people (Desk ?(Desk1).1). Stainings had been performed using isolated PBMCs for healthful settings newly, YFV vaccine recipients and HCV-infected people, or freezing PBMCs produced from HIV-1-contaminated people (Shape ?(Figure1).1). While frequencies of tetramer+ KIR2DL2/3+ (clone REA147+) NK cells assorted between your different study organizations, the comparative hierarchy from the particular tetramer+ NK cells didn’t differ between ABT-737 inhibitor HIV-1-contaminated, HCV-infected, YFV-vaccinated or control people. YFV/HLA-C*03:04-NS2A4?13-tetramers bound to the best percentage of KIR2DL2/3+ NK cells consistently, whereas HCV/HLA-C*03:04-Primary136?hIV/HLA-C*03:04-Gag296 and 144-tetramers?304-tetramers bound to a significantly decrease percentage of KIR2DL2/3+ NK cells (Numbers 1B,C). Binding of KIR2DL3-Fc create to 221-TAPko-HLA-C*03:04 pulsed with YFV/NSA2A4?13, HIV/Gag296?304, HCV/Primary136?144 peptide, respectively, demonstrated similar hierarchies (Supplementary Shape 1). The percentage of YFV/HLA-C*03:04-NS2A4?13-tetramer-binding KIR2DL2/3+ NK cells did furthermore not differ between all those encoding for HLA-C*03 and HLA-C*03-adverse all those (in the 10 people from the YFV vaccine and healthful cohorts that Rabbit Polyclonal to ELOVL1 HLA class We typing was obtainable, median of 74.2 vs. 78.8%, 0.9, Supplementary Shape 2). Taken collectively, these data display that KIR2DL2/3+ NK cells adhere to a consistent peptide-dependent hierarchy in their binding to HLA-C*03:04 tetramers, which is not influenced by whether a study subject encodes for HLA-C*03 and is furthermore independent of the underlying viral setting, suggesting a lack of antigen-dependent expansion of these NK cell populations. HLA-C group 1 tetramers, such as the HLA-C*03:04 tetramers used here, can therefore serve as a reagent to monitor the frequencies of KIR2DL2+ or KIR2DL3+ NK cells. Stable frequencies of YFV-specific tetramer+ KIR2DL2/3+ NK cells in YFV-vaccinated individuals over time To assess whether KIR2DL2/3+ NK cells expanded following antigen challenge, we performed YFV/HLA-C*03:04-NS2A4?13-tetramer staining of primary PBMCs in 5 YFV-vaccinated individuals at 0, 1, 3, and day 28 of vaccination with YFV-17D. Stainings with HIV/HLA-C*03:04-Gag296?304- and HCV/HLA-C*03:04-Core136?144-tetramers were performed at the same times as controls. To control for a possible influence of HCMV infection on NK cell frequencies, vaccine recipients were ABT-737 inhibitor tested for HCMV infection (3 individuals were positive and 2 negative for HCMV IgG or IgM, data not shown). No changes in the average frequency of YFV/HLA-C*03:04-NS2A4?13-tetramer+ KIR2DL2/3+ NK cells were observed following YFV-17D vaccination (Figure ?(Figure2).2). Already before YFV vaccination, YFV/HLA-C*03:04-NS2A4?13-tetramers bound to the majority of KIR2DL2/3+ NK cells (mean 74%, range 57C90%), and did not significantly change following vaccination. The percentage ABT-737 inhibitor of KIR2DL2/3+ NK cells binding either the HIV/HLA-C*03:04-Gag296?304- or the HCV/HLA-C*03:04-Core136?144-tetramer also.