granulysin  (R&D Systems)

Journal: bioRxiv

Article Title: Blinatumomab-driven T-cell activation in αβ and γδ T-cell subsets: Insights from in vitro assays§

doi: 10.1101/2025.11.17.688178

Figure Lengend Snippet: A: BCP-ALL cells in the presence (red) or absence (grey) of BLN for three days were co-cultured with freshly isolated γδ, CD4⁺ and CD8⁺ αβ T cells/ T- and B-cell frequencies in the coculture were assessed by staining for CD3 and CD19, as exemplary shown for CD8⁺ αβ T cells. B: The release of IFNγ, GrzB and Prf into supernatants of cocultures in the presence (red) or absence (grey) of BLN was assessed via ELISA after three days, and the release of Gnly was analyzed after seven days. Data includes 4 to 7 healthy donors and * indicates p value between 0.01-0.05, ** indicates p value between 0.01-0.001 (paired and unpaired T-test).

Article Snippet: Cell culture supernatants were analyzed for Interferon-γ (IFNγ; DIF50C), Granzyme B (GrzB; DGZB00), Perforin (QK8011), Granulysin (Gnly; DY3138), Perforin (Prf; QK8011) and soluble Fas-Ligand (sFAS-L; DFL00B) using sandwich ELISA kits from R&D Systems, Minneapolis, USA according to the manufacturer’s instructions.

Techniques: Cell Culture, Isolation, Staining, Enzyme-linked Immunosorbent Assay

Journal: Nature immunology

Article Title: γδ T cells suppress Plasmodium falciparum blood stage infection by direct killing and phagocytosis

doi: 10.1038/s41590-020-00847-4

Figure Lengend Snippet: a , Frequency of circulating lymphocytes from 8 healthy donors (HD) and 8 P. falciparum patients (Pf). CD3 + CD4 + (CD4), CD3 + CD8 + (CD8), CD3 + TCRδ2 + (γδ2), CD3 − CD56 + (NK). b , c , Circulating γδ2 T cell activation in 8 HD and 5 Pf was evaluated by CD69 and CD16 staining. d , Intracellular granulysin (GNLY), perforin (PFN) and granzyme B (GzmB) in 8 HD and 8 Pf. e , CFSE-stained iRBC were co-cultured with fresh purified γδ2 T cells from 3 HD and 3 Pf at indicated E:T ratios for 12 hr and iRBC lysis was assessed by flow cytometry analysis of the number of viable CFSE + cells relative to added counting beads. f , Plasma GNLY concentration in 16 HD and 22 Pf patients by GNLY ELISA. Statistical analysis was performed by two-tailed nonparametric unpaired t-test ( a,f ), two-way ANOVA with Sidak’s multiple comparisons test ( d ), and area under the curve, followed by two-tailed nonparametric paired t-test, followed by two-tailed nonparametric paired t-test ( e ). Mean ± s.e.m. is shown. P value: *<0.05, **<0.01, ***<0.001, ****<0.0001.

Article Snippet: GNLY in plasma from HD and Pf patients was quantified using the Human Granulysin DuoSet ELISA (R&D Systems).

Techniques: Activation Assay, Staining, Cell Culture, Purification, Lysis, Flow Cytometry, Clinical Proteomics, Concentration Assay, Enzyme-linked Immunosorbent Assay, Two Tailed Test

Journal: Nature immunology

Article Title: γδ T cells suppress Plasmodium falciparum blood stage infection by direct killing and phagocytosis

doi: 10.1038/s41590-020-00847-4

Figure Lengend Snippet: a , PBMCs were stained with the Live/Dead viability dye and antibodies to CD3, CD4, CD8, TCRδ2 and CD56. Single live cells were gated on SSC-A vs FSC-A and dead cells were excluded. Lymphocyte subpopulations were gated as: CD3 + CD4 + (CD4 + T cells), CD3 + CD8 + (CD8 + T cells), CD3 + TCRδ2 + (γδ2 T cells) and CD3 − CD56 + (NK). b, PBMCs co-stained for CD69, CD16 and cytotoxic effector protein expression (GNLY, PFN and GzmB) were gated on γδ2 T cells.

Article Snippet: GNLY in plasma from HD and Pf patients was quantified using the Human Granulysin DuoSet ELISA (R&D Systems).

Techniques: Staining, Expressing

Journal: Nature immunology

Article Title: γδ T cells suppress Plasmodium falciparum blood stage infection by direct killing and phagocytosis

doi: 10.1038/s41590-020-00847-4

Figure Lengend Snippet: a-c , PBMC from HD were evaluated ex vivo (day 0) and after co-culture with uRBCs or iRBCs for 7 days. Shown are representative flow cytometry histograms (left) and percent positive cells for 4 HD samples (right) after staining for cell surface CD3 + CD4 + (CD4), CD3 + CD8 + (CD8), CD3 + TCRδ2 + (γδ2), CD3 − CD56 + (NK) and intracellular GNLY ( a ), PFN ( b ) or GzmB ( c ). The vertical gating lines were drawn based on the unstained sample. d , e , γδ2 T cell activation before and during co-culture with uRBCs or iRBCs was evaluated by CD69 and CD16 staining, n=3 independent donor samples. Statistical analysis was performed by two-way ANOVA with Sidak’s multiple comparisons test ( a-c ) and two-way ANOVA with Tukey’s multiple comparisons test ( d-e ). Mean ± s.e.m. is shown. P value: *<0.05, **<0.01,***<0.001, ****<0.0001. Data shown are representative of three independent experiments.

Article Snippet: GNLY in plasma from HD and Pf patients was quantified using the Human Granulysin DuoSet ELISA (R&D Systems).

Techniques: Ex Vivo, Co-Culture Assay, Flow Cytometry, Staining, Activation Assay

Journal: Nature immunology

Article Title: γδ T cells suppress Plasmodium falciparum blood stage infection by direct killing and phagocytosis

doi: 10.1038/s41590-020-00847-4

Figure Lengend Snippet: a-c , RBCs, incubated with indicated combinations of cytotoxic effector proteins (n=3), were assayed for RBC lysis by LDH release ( a ), parasite reinvasion into fresh RBCs by flow cytometry ( b ), and parasite morphology by electron microscopy ( c,d ). ( c ) quantifies prominent morphological alterations in EM images - parasitophorous vacuole (PV) detachment (black arrows) and increased parasite vacuolization (red arrows) in 15 images from 3 sections. Parasite nucleus (N), food vacuole (FV), hemozoin (Hz) are labeled in untreated iRBCs. e, uRBCs or unsynchronized iRBCs, stained with 25-NBD or filipin as cholesterol probes and Hoechst for parasite DNA, were analyzed by imaging flow cytometry using DNA staining to gate for different parasite stages (R ring, T trophozoite, S schizont). BF, bright field, H, Hoechst dye. f , Representative histograms (top) and quantification of multiple samples (bottom) of AF647-GNLY binding to uRBCs and iRBCs (trophozoite stage) by flow cytometry (n=5). g,h , GzmB-AF488 internalization in the absence or presence of GNLY or PFN assessed by imaging flow cytometry (n=3). ( g ) shows GzmB-AF488 (Gzm) internalization in representative images and ( h ) shows the proportion of RBCs at different parasite stages that stained for GzmB in multiple experiments, n=3. i, Representative images (left) and quantification (right) of imaging flow cytometry showing RBC, uninfected or infected at indicated parasite stage, incubated with GzmB-AF488, GNLY-AF647 and unlabeled GNLY, n=3. M, merozoite; R, ring; T, trophozoite; S, schizont; H, Hoechst dye; BF, bright field; n, biological independent samples. Scale bars: 500 nm ( d ), 7 μm ( e,g,i ). Statistical analysis was by one-way ANOVA with Tukey’s multiple comparisons test ( a,b,c,h,i ) or two-tailed nonparametric unpaired t-test ( f ). Mean ± s.e.m. is shown. P value: *<0.05, **<0.01, ***<0.001, ****<0.0001. Data shown are representative of three independent experiments.

Article Snippet: GNLY in plasma from HD and Pf patients was quantified using the Human Granulysin DuoSet ELISA (R&D Systems).

Techniques: Incubation, Red Blood Cell Lysis, Flow Cytometry, Electron Microscopy, Labeling, Staining, Imaging, Binding Assay, Infection, Two Tailed Test

Journal: Nature immunology

Article Title: γδ T cells suppress Plasmodium falciparum blood stage infection by direct killing and phagocytosis

doi: 10.1038/s41590-020-00847-4

Figure Lengend Snippet: a , Electron microscopy of a ring stage iRBC treated or not with GzmB, PFN and GNLY showing disruption of morphology after treatment (right). In ( a ), parasitophorous vacuole (PV) detachment is indicated by a black arrow and increased parasite vacuolization by a red arrow. Parasite nucleus (N), hemoglobin vacuole (Hb), hemozoin (H). b , Imaging flow cytometry gating strategy for parasite developmental stages based on DNA content by Hoechst staining. M merozoites, R rings, T trophozoites, S schizonts. c, GzmB-AF488 uptake in the presence or absence of GNLY. Scale bar: 500 nm ( a ). n, biological independent samples. Data shown are representative of at least three independent experiments.

Article Snippet: GNLY in plasma from HD and Pf patients was quantified using the Human Granulysin DuoSet ELISA (R&D Systems).

Techniques: Electron Microscopy, Disruption, Imaging, Flow Cytometry, Staining