Journal: bioRxiv

Article Title: Evolutionary fingerprint in rodent PD1 confers weakened activity and enhanced tumor immunity compared to human PD1

doi: 10.1101/2024.09.21.614250

Figure Lengend Snippet: Left, FACS histograms showing hu PD1 expressions on anti-CD3ε/anti-CD28 stimulated human primary CD4 + T cells and on stable Jurkat cells transduced with hu PD1 driven by either an dSV40 (weak) or an SFFV (strong) promoter. Right, PD1 densities on human primary CD4 + T cells plotted against duration of stimulation by anti-CD3/anti-CD28 beads. Dashed lines indicate the densities of SFFV- and dSV40-driven PD1 expressions on Jurkat cells. PD1 densities were calculated using molecular numbers quantified with a Quantum PE MESF kit and diameters of cells (human primary CD4 + T cell: 4.8 µm, Jurkat cells: 15 µm).

Article Snippet: The absolute amount of PD1 surface expressions on human primary CD4 + T cells or on Jurkat cells was quantified using a Quantum PE MESF kit (Bangs laboratories) following the manufacturer’s instruction.

Techniques: Transduction

Journal: Immunity

Article Title: PD-L1:CD80 Cis -Heterodimer Triggers the Co-stimulatory Receptor CD28 While Repressing the Inhibitory PD-1 and CTLA-4 Pathways

doi: 10.1016/j.immuni.2019.11.003

Figure Lengend Snippet: (A) Representative flow-cytometry histograms of PD-1-huFc staining of the indicated types of Raji cells. Bound PD-1-huFc was labeled by AF647 anti-human IgG Fc, the mean fluorescence intensity (MFI) of which was plotted against PD-1-huFc concentration (means ± SEM, n = 3). In gray is the staining of Raji (CD80−PD-L1-mCherry+) by isolated huFc domain.

Article Snippet: Quantification of PD-L1 and CD80 Expression For flow cytometry based quantification in Figure S1 , PD-L1 and CD80 were stained by PE anti-PD-L1 (eBioscience, 14-5983-82) and PE anti-CD80 (Biolegend, 305208) and their expression levels were quantified using the QUANTUM R-PE MESF kit (Bangs Laboratories Inc, 827), following manufacturer’s instructions.

Techniques: Flow Cytometry, Staining, Labeling, Fluorescence, Concentration Assay, Isolation

Journal: Immunity

Article Title: PD-L1:CD80 Cis -Heterodimer Triggers the Co-stimulatory Receptor CD28 While Repressing the Inhibitory PD-1 and CTLA-4 Pathways

doi: 10.1016/j.immuni.2019.11.003

Figure Lengend Snippet: (A) A Jurkat-Raji co-culture assay analyzing how PD-L1 interferes with CTLA-4-mediated CD80 depletion. Cartoons on the left depict the co-cultured cells. On the immediate right are representative flow-cytometry histograms of CD80 expression (anti-CD80 allophycocyanin) on Raji cells before (0 h) and after co-culture (0.5 h). Further on the right are representative confocal images for the Jurkat-Raji conjugate (scale bars, 10 μm). Rightmost is a bar graph showing CD80 MFI of Raji at 0.5 h, normalized to CD80 MFI at 0 h (mean ± SEM, n = 4).

Article Snippet: Quantification of PD-L1 and CD80 Expression For flow cytometry based quantification in Figure S1 , PD-L1 and CD80 were stained by PE anti-PD-L1 (eBioscience, 14-5983-82) and PE anti-CD80 (Biolegend, 305208) and their expression levels were quantified using the QUANTUM R-PE MESF kit (Bangs Laboratories Inc, 827), following manufacturer’s instructions.

Techniques: Co-culture Assay, Cell Culture, Flow Cytometry, Expressing, Co-Culture Assay

Journal: Immunity

Article Title: PD-L1:CD80 Cis -Heterodimer Triggers the Co-stimulatory Receptor CD28 While Repressing the Inhibitory PD-1 and CTLA-4 Pathways

doi: 10.1016/j.immuni.2019.11.003

Figure Lengend Snippet: (A) Representative TIRF images of PD-L1 LUVs captured by PD-1 SLB, CD80 SLB, or CD86 SLB; each LUV is registered as a green spot. Bar graph summarizes the fluorescence intensity (FI) of the LUV channel under indicated conditions, normalized to the intensity of the condition with PD-1 SLBs. Data are means ± SEM, n = 3. Scale bars, 5 μm.

Article Snippet: Quantification of PD-L1 and CD80 Expression For flow cytometry based quantification in Figure S1 , PD-L1 and CD80 were stained by PE anti-PD-L1 (eBioscience, 14-5983-82) and PE anti-CD80 (Biolegend, 305208) and their expression levels were quantified using the QUANTUM R-PE MESF kit (Bangs Laboratories Inc, 827), following manufacturer’s instructions.

Techniques: Fluorescence

Journal: Immunity

Article Title: PD-L1:CD80 Cis -Heterodimer Triggers the Co-stimulatory Receptor CD28 While Repressing the Inhibitory PD-1 and CTLA-4 Pathways

doi: 10.1016/j.immuni.2019.11.003

Figure Lengend Snippet: (A) On the left are flow-cytometry histograms of CD80 and CD86 surface levels on DCs and macrophages (Macs) isolated from tumor tissues of 4T1 implanted BALB/C female mice treated with anti-PD-L1 (magenta traces), anti-PD-1 (cyan traces), or control IgG (black traces). On the right are bar graphs summarizing the MFI of CD80 and CD86 staining under the indicated conditions. Data are shown as mean ± SEM, n ≥ 3 mice.

Article Snippet: Quantification of PD-L1 and CD80 Expression For flow cytometry based quantification in Figure S1 , PD-L1 and CD80 were stained by PE anti-PD-L1 (eBioscience, 14-5983-82) and PE anti-CD80 (Biolegend, 305208) and their expression levels were quantified using the QUANTUM R-PE MESF kit (Bangs Laboratories Inc, 827), following manufacturer’s instructions.

Techniques: Flow Cytometry, Isolation, Staining

Journal: Immunity

Article Title: PD-L1:CD80 Cis -Heterodimer Triggers the Co-stimulatory Receptor CD28 While Repressing the Inhibitory PD-1 and CTLA-4 Pathways

doi: 10.1016/j.immuni.2019.11.003

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: Quantification of PD-L1 and CD80 Expression For flow cytometry based quantification in Figure S1 , PD-L1 and CD80 were stained by PE anti-PD-L1 (eBioscience, 14-5983-82) and PE anti-CD80 (Biolegend, 305208) and their expression levels were quantified using the QUANTUM R-PE MESF kit (Bangs Laboratories Inc, 827), following manufacturer’s instructions.

Techniques: Recombinant, Isolation, Enzyme-linked Immunosorbent Assay, Cell Isolation, Software, Imaging