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pd 1 dimer protein  (Sartorius AG)


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    Structured Review

    Sartorius AG pd 1 dimer protein
    ( A ) In silico model representing the binding of Fab fragment from clones A35795 (A35) and 135c139d6 (135c) compared to the hPD-1/hPD-L1 complex (PDB ID: 4ZQK). ( B ) Binding of A35 and 135c IgG to <t>human</t> <t>PD-1</t> (left). Competitive binding assay with pembrolizumab and biotin-labeled A35 IgG or 135c IgG (right). Symbols are means of three (left) or two (right) independent experiments. ( C ) Binding of A35 and 135c Fabs to human PD-1 (left). Biolayer interferometry of Fabs–PD-1 interactions (right). Symbols are means of three independent experiments (left). ( D ) Binding of A35 and 135c scFvs-His to human PD-1 (left). Biolayer interferometry of scFvs–PD-1 interactions (right). Symbols are the mean of three independent experiments (left). ( E ) Second-generation CAR design (left). Biotinylated PD-1 binding to anti–PD-1 CARs. Representative flow cytometry and mean of four independent experiments. PD-1, programmed cell death protein 1; PD-L1, programmed cell death ligand 1; IgG, immunoglobulin G; Fab, fragment antigen-binding region; K D , equilibrium dissociation constant; scFv, single-chain variable fragment; His, histidine; CAR, chimeric antigen receptor; EGFRt, truncated epidermal growth factor receptor; HD, hinge domain; bPD1-CAR, blocking anti–PD-1 CAR; nbPD1-CAR, nonblocking anti–PD-1 CAR; UT, untransduced cells.
    Pd 1 Dimer Protein, supplied by Sartorius AG, used in various techniques. Bioz Stars score: 99/100, based on 8481 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/product/pd+1+dimer+protein/Octet/pmc13108567-211-10-27
    Average 99 stars, based on 8481 article reviews
    pd 1 dimer protein - by Bioz Stars, 2026-09
    99/100 stars

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    1) Product Images from "Targeting PD-1 + T cells with chimeric antigen receptors to reduce the HIV reservoir"

    Article Title: Targeting PD-1 + T cells with chimeric antigen receptors to reduce the HIV reservoir

    Journal: Science Advances

    doi: 10.1126/sciadv.aeb7602

    ( A ) In silico model representing the binding of Fab fragment from clones A35795 (A35) and 135c139d6 (135c) compared to the hPD-1/hPD-L1 complex (PDB ID: 4ZQK). ( B ) Binding of A35 and 135c IgG to human PD-1 (left). Competitive binding assay with pembrolizumab and biotin-labeled A35 IgG or 135c IgG (right). Symbols are means of three (left) or two (right) independent experiments. ( C ) Binding of A35 and 135c Fabs to human PD-1 (left). Biolayer interferometry of Fabs–PD-1 interactions (right). Symbols are means of three independent experiments (left). ( D ) Binding of A35 and 135c scFvs-His to human PD-1 (left). Biolayer interferometry of scFvs–PD-1 interactions (right). Symbols are the mean of three independent experiments (left). ( E ) Second-generation CAR design (left). Biotinylated PD-1 binding to anti–PD-1 CARs. Representative flow cytometry and mean of four independent experiments. PD-1, programmed cell death protein 1; PD-L1, programmed cell death ligand 1; IgG, immunoglobulin G; Fab, fragment antigen-binding region; K D , equilibrium dissociation constant; scFv, single-chain variable fragment; His, histidine; CAR, chimeric antigen receptor; EGFRt, truncated epidermal growth factor receptor; HD, hinge domain; bPD1-CAR, blocking anti–PD-1 CAR; nbPD1-CAR, nonblocking anti–PD-1 CAR; UT, untransduced cells.
    Figure Legend Snippet: ( A ) In silico model representing the binding of Fab fragment from clones A35795 (A35) and 135c139d6 (135c) compared to the hPD-1/hPD-L1 complex (PDB ID: 4ZQK). ( B ) Binding of A35 and 135c IgG to human PD-1 (left). Competitive binding assay with pembrolizumab and biotin-labeled A35 IgG or 135c IgG (right). Symbols are means of three (left) or two (right) independent experiments. ( C ) Binding of A35 and 135c Fabs to human PD-1 (left). Biolayer interferometry of Fabs–PD-1 interactions (right). Symbols are means of three independent experiments (left). ( D ) Binding of A35 and 135c scFvs-His to human PD-1 (left). Biolayer interferometry of scFvs–PD-1 interactions (right). Symbols are the mean of three independent experiments (left). ( E ) Second-generation CAR design (left). Biotinylated PD-1 binding to anti–PD-1 CARs. Representative flow cytometry and mean of four independent experiments. PD-1, programmed cell death protein 1; PD-L1, programmed cell death ligand 1; IgG, immunoglobulin G; Fab, fragment antigen-binding region; K D , equilibrium dissociation constant; scFv, single-chain variable fragment; His, histidine; CAR, chimeric antigen receptor; EGFRt, truncated epidermal growth factor receptor; HD, hinge domain; bPD1-CAR, blocking anti–PD-1 CAR; nbPD1-CAR, nonblocking anti–PD-1 CAR; UT, untransduced cells.

    Techniques Used: In Silico, Binding Assay, Clone Assay, Competitive Binding Assay, Labeling, Flow Cytometry, Blocking Assay

    ( A ) Experimental design for engineering human anti–PD-1 CAR-T cells. ( B ) Expansion of primary anti–PD-1 CAR-T cells compared to the anti-CD19 CAR control ( n = 7 donors, seven independent experiments). ( C ) Cumulative data of transduction efficiency for donors reported in (D) and (E). The mean ± SD values eight donors, eight independent experiments, two to three replicates transduced with different MOI is shown. Two-way ANOVA, Tukey’s multiple comparison test. ( D ) Representative flow cytometry plots showing mCherry and PD-1 expression 6 days after transduction (gated on CD3 + living cells). ( E ) Correlation between PD-1 expression and the percentage of transduction efficiency. Cherry-positive and Cherry-negative population are shown. For each experiment the percentage of PD-1 + cells were assessed in Cherry-negative and Cherry-positive cells and normalized to PD-1 expression in untransduced T cells (set as 100%). Eight donors, eight independent experiments each with two to three internal replicates (with different MOI transduction: 1.4 ± 0.6). Simple linear regression tests. ( F ) CD8/CD4 ratio in mCherry ± T cells comparing anti–PD-1 CAR and CD19 CAR-T cells ± PD-1 knockout. Data are presented as mean ± SD values of eight donors, eight independent experiments, two to three internal replicates). Two-way ANOVA, Tukey’s multiple comparisons test. ( G ) Schematic representing the killing assay (left). Cumulative percentage of annexin/Aqua + CD4 + PD-1 + GFP + target cells after 24 hours of coculture (right). Mean ± SD of three donors and three independent experiments is shown. Two-way ANOVA, Tukey’s multiple comparison test. Only statistical differences are reported as follows: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001, and **** P ≤ 0.0001. HD, hinge domain; TMD, transmembrane domain; ICD, intracellular domain; LV, lentivirus; scFv, single-chain variable fragment; CAR, chimeric antigen receptor; bPD1-CAR, blocking anti–PD-1 CAR; nbPD1-CAR, nonblocking anti–PD-1 CAR; FMO, fluorescence minus one; UT, untransduced; KO, knockout.
    Figure Legend Snippet: ( A ) Experimental design for engineering human anti–PD-1 CAR-T cells. ( B ) Expansion of primary anti–PD-1 CAR-T cells compared to the anti-CD19 CAR control ( n = 7 donors, seven independent experiments). ( C ) Cumulative data of transduction efficiency for donors reported in (D) and (E). The mean ± SD values eight donors, eight independent experiments, two to three replicates transduced with different MOI is shown. Two-way ANOVA, Tukey’s multiple comparison test. ( D ) Representative flow cytometry plots showing mCherry and PD-1 expression 6 days after transduction (gated on CD3 + living cells). ( E ) Correlation between PD-1 expression and the percentage of transduction efficiency. Cherry-positive and Cherry-negative population are shown. For each experiment the percentage of PD-1 + cells were assessed in Cherry-negative and Cherry-positive cells and normalized to PD-1 expression in untransduced T cells (set as 100%). Eight donors, eight independent experiments each with two to three internal replicates (with different MOI transduction: 1.4 ± 0.6). Simple linear regression tests. ( F ) CD8/CD4 ratio in mCherry ± T cells comparing anti–PD-1 CAR and CD19 CAR-T cells ± PD-1 knockout. Data are presented as mean ± SD values of eight donors, eight independent experiments, two to three internal replicates). Two-way ANOVA, Tukey’s multiple comparisons test. ( G ) Schematic representing the killing assay (left). Cumulative percentage of annexin/Aqua + CD4 + PD-1 + GFP + target cells after 24 hours of coculture (right). Mean ± SD of three donors and three independent experiments is shown. Two-way ANOVA, Tukey’s multiple comparison test. Only statistical differences are reported as follows: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001, and **** P ≤ 0.0001. HD, hinge domain; TMD, transmembrane domain; ICD, intracellular domain; LV, lentivirus; scFv, single-chain variable fragment; CAR, chimeric antigen receptor; bPD1-CAR, blocking anti–PD-1 CAR; nbPD1-CAR, nonblocking anti–PD-1 CAR; FMO, fluorescence minus one; UT, untransduced; KO, knockout.

    Techniques Used: Control, Transduction, Comparison, Flow Cytometry, Expressing, Knock-Out, Blocking Assay, Fluorescence

    ( A ) Generation of PD-1-transgenic luciferase + Jurkat cells with increasing PD-1 molecules number. ( B ) Luciferase-based killing assay of blocking and nonblocking CAR-T cells. Mean ± SD of three donors and three independent experiments is shown. ( C ) Phenotype of the K562 cell lines expressing either a PD-1-GFP-F fusion protein (K-PD-1-GFP-F) or PD-1 and GFP proteins with a ribosomal skipping motif in-between (K-PD-1-GFP-noF) used for the trogocytosis assay. Scale bars, 25 μm. ( D ) Representative flow cytometry 1 hour after coculture of K-PD-1-GFP-F or -PD-1-GFP-noF with anti–PD-1 CAR-T cells. ( E ) Cumulative data showing GFP expression in CAR-T cells over time. Mean ± SD values of six donors and four independent experiments is shown. Two-way ANOVA, Tukey’s multiple comparison test. ( F ) Phenotype of the K562 cell lines expressing either the wild-type PD-1 (K-PD-1 WT ) or a truncated PD-1 lacking (K-PD-1 t ) the intracellular domain used for the activation assay. ( G ) Cumulative percentage of CD25 + /CD71 + CAR-T cells after a 48 hours coculture with K-PD-1 WT or K-PD-1 t . Mean ± SD values of four to eight donors and seven independent experiments is shown. Two-way ANOVA, Tukey’s multiple comparison test. ( H ) Luciferase killing assay using PD-1 WT or PD-1 t luciferase + Jurkat cells as targets. Mean ± SD values of three donors, three independent experiments is shown. Two-way ANOVA, Tukey’s multiple comparison test was used to compare the same condition in JPD-1 versus JPD-1 t . Only statistical differences are reported as follows: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001, and **** P ≤ 0.0001. JPD-1, Jurkat-PD-1; CAR, chimeric antigen receptor; bPD1-CAR, blocking anti–PD-1 CAR; nbPD1-CAR, nonblocking anti–PD-1 CAR; E:T, effector:target; K-PD-1-GFP-F, K562 PD-1-GFP-F fusion protein; K-PD-1-GFP-noF, K562 PD-1 + GFP + ; K-PD-1, K562 PD-1 + ; K-PD-1-t, K562 PD-1 truncated; K-PD-1-KO, K562 PD-1 KO; JPD-1-t, Jurkat PD-1 truncated.
    Figure Legend Snippet: ( A ) Generation of PD-1-transgenic luciferase + Jurkat cells with increasing PD-1 molecules number. ( B ) Luciferase-based killing assay of blocking and nonblocking CAR-T cells. Mean ± SD of three donors and three independent experiments is shown. ( C ) Phenotype of the K562 cell lines expressing either a PD-1-GFP-F fusion protein (K-PD-1-GFP-F) or PD-1 and GFP proteins with a ribosomal skipping motif in-between (K-PD-1-GFP-noF) used for the trogocytosis assay. Scale bars, 25 μm. ( D ) Representative flow cytometry 1 hour after coculture of K-PD-1-GFP-F or -PD-1-GFP-noF with anti–PD-1 CAR-T cells. ( E ) Cumulative data showing GFP expression in CAR-T cells over time. Mean ± SD values of six donors and four independent experiments is shown. Two-way ANOVA, Tukey’s multiple comparison test. ( F ) Phenotype of the K562 cell lines expressing either the wild-type PD-1 (K-PD-1 WT ) or a truncated PD-1 lacking (K-PD-1 t ) the intracellular domain used for the activation assay. ( G ) Cumulative percentage of CD25 + /CD71 + CAR-T cells after a 48 hours coculture with K-PD-1 WT or K-PD-1 t . Mean ± SD values of four to eight donors and seven independent experiments is shown. Two-way ANOVA, Tukey’s multiple comparison test. ( H ) Luciferase killing assay using PD-1 WT or PD-1 t luciferase + Jurkat cells as targets. Mean ± SD values of three donors, three independent experiments is shown. Two-way ANOVA, Tukey’s multiple comparison test was used to compare the same condition in JPD-1 versus JPD-1 t . Only statistical differences are reported as follows: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001, and **** P ≤ 0.0001. JPD-1, Jurkat-PD-1; CAR, chimeric antigen receptor; bPD1-CAR, blocking anti–PD-1 CAR; nbPD1-CAR, nonblocking anti–PD-1 CAR; E:T, effector:target; K-PD-1-GFP-F, K562 PD-1-GFP-F fusion protein; K-PD-1-GFP-noF, K562 PD-1 + GFP + ; K-PD-1, K562 PD-1 + ; K-PD-1-t, K562 PD-1 truncated; K-PD-1-KO, K562 PD-1 KO; JPD-1-t, Jurkat PD-1 truncated.

    Techniques Used: Transgenic Assay, Luciferase, Blocking Assay, Expressing, Trogocytosis Assay, Flow Cytometry, Comparison, Activation Assay

    ( A ) Experimental design. HLA-A2 + human T cells were isolated and activated ± prior editing of PD-1 and/or HLA-A2. ( B ) PD-1 expression in mCherry + and mCherry − cells on days 7 and 10. Mean of three to five donors in three to five independent experiments with one to two replicates transduced at different MOI (1.4 ± 0.6) for each independent experiments. ( C ) HLA-A2 expression in mCherry + and mCherry − cells on day 7 and 10. Mean of two donors in two independent experiments with one to two internal replicates (different MOI transduction: 1.4 ± 0.6). ( D ) Expansion folds of unedited versus edited CAR-T cells on day 10. Mean ± SD values of three to five donors in three to five independent experiments with one to two internal replicates (different MOI transduction: 1.4 ± 0.6). Statistics: Two-way ANOVA, Dunnett’s multiple comparison test was performed comparing each condition with the corresponding HLA-A2 CAR condition. ( E ) Cumulative data showing CD62L/CD45RA expression of mCherry + cells in edited versus unedited condition for PD-1 or HLA-A2. The mean ± SD values of three to five donors in three to five independent experiments with one to two internal replicates (different MOI transduction 1.4 ± 0.6) are shown. Kruskal-Wallis and Dunn’s multiple comparison test was performed comparing each condition with the corresponding HLA-A2 CAR condition. ( F ) Representative flow cytometry showing TIM-3 and LAG-3 expression in the different CAR populations on day 10 (left). Cumulative percentage of LAG-3 + /TIM-3 + double-positive cells (right). Paired data of three to five donors in three to five independent experiments with one to two internal replicates (different MOI transduction: 1.4 ± 0.6) are shown. Two-way ANOVA, Dunnett’s multiple comparison test was performed comparing each condition with the corresponding HLA-A2 CAR condition. Only statistical differences are reported as follows: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001, and **** P ≤ 0.0001.
    Figure Legend Snippet: ( A ) Experimental design. HLA-A2 + human T cells were isolated and activated ± prior editing of PD-1 and/or HLA-A2. ( B ) PD-1 expression in mCherry + and mCherry − cells on days 7 and 10. Mean of three to five donors in three to five independent experiments with one to two replicates transduced at different MOI (1.4 ± 0.6) for each independent experiments. ( C ) HLA-A2 expression in mCherry + and mCherry − cells on day 7 and 10. Mean of two donors in two independent experiments with one to two internal replicates (different MOI transduction: 1.4 ± 0.6). ( D ) Expansion folds of unedited versus edited CAR-T cells on day 10. Mean ± SD values of three to five donors in three to five independent experiments with one to two internal replicates (different MOI transduction: 1.4 ± 0.6). Statistics: Two-way ANOVA, Dunnett’s multiple comparison test was performed comparing each condition with the corresponding HLA-A2 CAR condition. ( E ) Cumulative data showing CD62L/CD45RA expression of mCherry + cells in edited versus unedited condition for PD-1 or HLA-A2. The mean ± SD values of three to five donors in three to five independent experiments with one to two internal replicates (different MOI transduction 1.4 ± 0.6) are shown. Kruskal-Wallis and Dunn’s multiple comparison test was performed comparing each condition with the corresponding HLA-A2 CAR condition. ( F ) Representative flow cytometry showing TIM-3 and LAG-3 expression in the different CAR populations on day 10 (left). Cumulative percentage of LAG-3 + /TIM-3 + double-positive cells (right). Paired data of three to five donors in three to five independent experiments with one to two internal replicates (different MOI transduction: 1.4 ± 0.6) are shown. Two-way ANOVA, Dunnett’s multiple comparison test was performed comparing each condition with the corresponding HLA-A2 CAR condition. Only statistical differences are reported as follows: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001, and **** P ≤ 0.0001.

    Techniques Used: Isolation, Expressing, Transduction, Comparison, Flow Cytometry

    ( A ) Experimental design. Mice received adoptive cell transfer (ACT) of anti–PD-1 CARs versus untransduced (UT) T cells under ART that was interrupted after 2 weeks (three independent experiments). The reporter used for detecting CAR-positive cells was EGFRt. ( B ) Kaplan-Meyer curve showing the percentage of mice maintaining viral control. All mice were included in the analysis. Log-rank (Mantel-Cox) test was performed using the UT control condition as reference. ( C ) Plasma viral load overtime (UT, n = 6; bPD1-CAR, n = 9; nbPD1-CAR, n = 7). ( D ) Percentage of CAR-T cells (EGFRt + gated in CD45 + cells) in the spleen, BM, and blood at the time of euthanasia. Mean ± SD is shown. bPD1-CAR, n = 9, nbPD1-CAR, n = 7. ( E ) Percentage of CD8 + , CD4 + , and CD4 − CAR-T cells detected in the spleen, BM, and blood (bPD1-CAR, n = 3, nbPD1-CAR, n = 4). Mean ± SD is shown. ( F ) Correlation between the percentage of CAR-T cells (defined as CD45 + EGFRt + cells) and the time of viral rebound (bPD1-CAR, n = 9; nbPD1-CAR, n = 7). Simple linear regression was used. ( G ) Correlation between CD4 + PD-1 + cells (gated in huCD45 + EGFRt − cells) and the time of viral rebound (bPD1-CAR, n = 9; nbPD1-CAR, n = 7). Simple linear regression was used. ( H ) Correlation between CD4 + PD-1 + cells (gated in huCD45 + EGFRt − cells) and CAR-T cells detection (bPD1-CAR, n = 9; nbPD1-CAR, n = 7). Simple linear regression was used. ( I ) Correlation between HIV integrated DNA in CD4 + EGFRt- sorted cells from spleen and BM and time of viral rebound (bPD1-CAR, n = 3; nbPD1-CAR, n = 3). Simple linear regression was used. ( J ) Correlation between HIV-integrated DNA in CD4 + EGFRt − sorted cells from spleen and BM and CAR-T cells detection (bPD1-CAR, n = 3; nbPD1-CAR, n = 3). Simple linear regression was used. ART, antiretroviral therapy; ACT, adoptive cell transfer; VC, viral control.
    Figure Legend Snippet: ( A ) Experimental design. Mice received adoptive cell transfer (ACT) of anti–PD-1 CARs versus untransduced (UT) T cells under ART that was interrupted after 2 weeks (three independent experiments). The reporter used for detecting CAR-positive cells was EGFRt. ( B ) Kaplan-Meyer curve showing the percentage of mice maintaining viral control. All mice were included in the analysis. Log-rank (Mantel-Cox) test was performed using the UT control condition as reference. ( C ) Plasma viral load overtime (UT, n = 6; bPD1-CAR, n = 9; nbPD1-CAR, n = 7). ( D ) Percentage of CAR-T cells (EGFRt + gated in CD45 + cells) in the spleen, BM, and blood at the time of euthanasia. Mean ± SD is shown. bPD1-CAR, n = 9, nbPD1-CAR, n = 7. ( E ) Percentage of CD8 + , CD4 + , and CD4 − CAR-T cells detected in the spleen, BM, and blood (bPD1-CAR, n = 3, nbPD1-CAR, n = 4). Mean ± SD is shown. ( F ) Correlation between the percentage of CAR-T cells (defined as CD45 + EGFRt + cells) and the time of viral rebound (bPD1-CAR, n = 9; nbPD1-CAR, n = 7). Simple linear regression was used. ( G ) Correlation between CD4 + PD-1 + cells (gated in huCD45 + EGFRt − cells) and the time of viral rebound (bPD1-CAR, n = 9; nbPD1-CAR, n = 7). Simple linear regression was used. ( H ) Correlation between CD4 + PD-1 + cells (gated in huCD45 + EGFRt − cells) and CAR-T cells detection (bPD1-CAR, n = 9; nbPD1-CAR, n = 7). Simple linear regression was used. ( I ) Correlation between HIV integrated DNA in CD4 + EGFRt- sorted cells from spleen and BM and time of viral rebound (bPD1-CAR, n = 3; nbPD1-CAR, n = 3). Simple linear regression was used. ( J ) Correlation between HIV-integrated DNA in CD4 + EGFRt − sorted cells from spleen and BM and CAR-T cells detection (bPD1-CAR, n = 3; nbPD1-CAR, n = 3). Simple linear regression was used. ART, antiretroviral therapy; ACT, adoptive cell transfer; VC, viral control.

    Techniques Used: Control, Clinical Proteomics

    ( A ) Immunofluorescence multicolor confocal images of spleens of four control and seven treated hu-mice with detectable CAR-T cells (untransduced treated, n = 4; nbPD1-CAR-T treated, n = 4; bPD1-CAR-T treated, n = 3) 8 to 10 weeks after ACT. The following stainings were performed: DAPI (blue), CD4 (green), CD20 (red), PD-1 (magenta), CD8 (yellow), Grzb (cyan), and CD57 (gray). Scale bars, 100 μm. ( B ) Histo-cytometry analysis showing CD4 + PD-1 + , and CD8 + PD-1 + cell counts/mm 2 in the CD20-enriched areas and CD20 + cell counts/mm 2 in the CD4-enriched zones. Each dot represents a CD20- or CD4-enriched region ( n = 154 and n = 158). Median is shown. Kruskal-Wallis and Dunn’s multiple comparison test. All control mice ( n = 6) and treated mice with detectable CARs ( n = 7) that survived until the end of the experiment were included. ( C ) Mean distance between each B cell and its five nearest neighbors in the CD20-enriched regions in the 13 spleens based on IF staining data. Median is shown. Kruskal-Wallis and Dunn’s multiple comparison test was performed. ( D ) Combined IF and RNAscope for CD8 (green), DAPI (blue), HIV RNA (red), and CAR RNA (magenta) performed on the spleen of one representative control mouse (S2), one HIV suppressed responder mouse (S11), and one nonresponder mouse with viral rebound (S5) 8 weeks after ACT. Scale bars, 50 μm. ( E ) Histo-cytometry analysis showing absolute counts of CAR RNA events (gated on CD8 + cells) and HIV RNA events for each mouse tissue. Two-way ANOVA, Tukey’s multiple comparison test. Only statistical differences are reported as follows: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001, and **** P ≤ 0.0001. S, spleen; bCAR, blocking anti–PD-1 CAR; nbCAR, nonblocking anti–PD-1 CAR; ACT, adoptive cell transfer; UT, untransduced T cells.
    Figure Legend Snippet: ( A ) Immunofluorescence multicolor confocal images of spleens of four control and seven treated hu-mice with detectable CAR-T cells (untransduced treated, n = 4; nbPD1-CAR-T treated, n = 4; bPD1-CAR-T treated, n = 3) 8 to 10 weeks after ACT. The following stainings were performed: DAPI (blue), CD4 (green), CD20 (red), PD-1 (magenta), CD8 (yellow), Grzb (cyan), and CD57 (gray). Scale bars, 100 μm. ( B ) Histo-cytometry analysis showing CD4 + PD-1 + , and CD8 + PD-1 + cell counts/mm 2 in the CD20-enriched areas and CD20 + cell counts/mm 2 in the CD4-enriched zones. Each dot represents a CD20- or CD4-enriched region ( n = 154 and n = 158). Median is shown. Kruskal-Wallis and Dunn’s multiple comparison test. All control mice ( n = 6) and treated mice with detectable CARs ( n = 7) that survived until the end of the experiment were included. ( C ) Mean distance between each B cell and its five nearest neighbors in the CD20-enriched regions in the 13 spleens based on IF staining data. Median is shown. Kruskal-Wallis and Dunn’s multiple comparison test was performed. ( D ) Combined IF and RNAscope for CD8 (green), DAPI (blue), HIV RNA (red), and CAR RNA (magenta) performed on the spleen of one representative control mouse (S2), one HIV suppressed responder mouse (S11), and one nonresponder mouse with viral rebound (S5) 8 weeks after ACT. Scale bars, 50 μm. ( E ) Histo-cytometry analysis showing absolute counts of CAR RNA events (gated on CD8 + cells) and HIV RNA events for each mouse tissue. Two-way ANOVA, Tukey’s multiple comparison test. Only statistical differences are reported as follows: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001, and **** P ≤ 0.0001. S, spleen; bCAR, blocking anti–PD-1 CAR; nbCAR, nonblocking anti–PD-1 CAR; ACT, adoptive cell transfer; UT, untransduced T cells.

    Techniques Used: Immunofluorescence, Control, Cytometry, Comparison, Staining, RNAscope, Blocking Assay

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    Binding Assay:

    Article Title: Targeting PD-1 + T cells with chimeric antigen receptors to reduce the HIV reservoir
    Article Snippet: .. Kinetic analysis of A35/135C IgG, Fab, and scFv binding to PD-1 dimer protein was performed by biolayer interferometry immobilizing PD-1 biotinylated dimer (3 μg/ml) on streptavidin biosensors (Sartorius, Göttingen, Germany, catalog no. 18-5019) dipped into a solution of the Fab/scFv at different concentrations diluted in PBS (ranging from 40 to 240 nM), and the nm shift was recorded on the Octet. ..



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    pd 1 dimer protein  (Sartorius AG)
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    Sartorius AG pd 1 dimer protein
    ( A ) In silico model representing the binding of Fab fragment from clones A35795 (A35) and 135c139d6 (135c) compared to the hPD-1/hPD-L1 complex (PDB ID: 4ZQK). ( B ) Binding of A35 and 135c IgG to <t>human</t> <t>PD-1</t> (left). Competitive binding assay with pembrolizumab and biotin-labeled A35 IgG or 135c IgG (right). Symbols are means of three (left) or two (right) independent experiments. ( C ) Binding of A35 and 135c Fabs to human PD-1 (left). Biolayer interferometry of Fabs–PD-1 interactions (right). Symbols are means of three independent experiments (left). ( D ) Binding of A35 and 135c scFvs-His to human PD-1 (left). Biolayer interferometry of scFvs–PD-1 interactions (right). Symbols are the mean of three independent experiments (left). ( E ) Second-generation CAR design (left). Biotinylated PD-1 binding to anti–PD-1 CARs. Representative flow cytometry and mean of four independent experiments. PD-1, programmed cell death protein 1; PD-L1, programmed cell death ligand 1; IgG, immunoglobulin G; Fab, fragment antigen-binding region; K D , equilibrium dissociation constant; scFv, single-chain variable fragment; His, histidine; CAR, chimeric antigen receptor; EGFRt, truncated epidermal growth factor receptor; HD, hinge domain; bPD1-CAR, blocking anti–PD-1 CAR; nbPD1-CAR, nonblocking anti–PD-1 CAR; UT, untransduced cells.
    Pd 1 Dimer Protein, supplied by Sartorius AG, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/product/pd+1+dimer+protein/Octet/pmc13108567-211-10-27
    Average 99 stars, based on 1 article reviews
    pd 1 dimer protein - by Bioz Stars, 2026-09
    99/100 stars
    		  Buy from Supplier

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    ( A ) In silico model representing the binding of Fab fragment from clones A35795 (A35) and 135c139d6 (135c) compared to the hPD-1/hPD-L1 complex (PDB ID: 4ZQK). ( B ) Binding of A35 and 135c IgG to human PD-1 (left). Competitive binding assay with pembrolizumab and biotin-labeled A35 IgG or 135c IgG (right). Symbols are means of three (left) or two (right) independent experiments. ( C ) Binding of A35 and 135c Fabs to human PD-1 (left). Biolayer interferometry of Fabs–PD-1 interactions (right). Symbols are means of three independent experiments (left). ( D ) Binding of A35 and 135c scFvs-His to human PD-1 (left). Biolayer interferometry of scFvs–PD-1 interactions (right). Symbols are the mean of three independent experiments (left). ( E ) Second-generation CAR design (left). Biotinylated PD-1 binding to anti–PD-1 CARs. Representative flow cytometry and mean of four independent experiments. PD-1, programmed cell death protein 1; PD-L1, programmed cell death ligand 1; IgG, immunoglobulin G; Fab, fragment antigen-binding region; K D , equilibrium dissociation constant; scFv, single-chain variable fragment; His, histidine; CAR, chimeric antigen receptor; EGFRt, truncated epidermal growth factor receptor; HD, hinge domain; bPD1-CAR, blocking anti–PD-1 CAR; nbPD1-CAR, nonblocking anti–PD-1 CAR; UT, untransduced cells.

    Journal: Science Advances

    Article Title: Targeting PD-1 + T cells with chimeric antigen receptors to reduce the HIV reservoir

    doi: 10.1126/sciadv.aeb7602

    Figure Lengend Snippet: ( A ) In silico model representing the binding of Fab fragment from clones A35795 (A35) and 135c139d6 (135c) compared to the hPD-1/hPD-L1 complex (PDB ID: 4ZQK). ( B ) Binding of A35 and 135c IgG to human PD-1 (left). Competitive binding assay with pembrolizumab and biotin-labeled A35 IgG or 135c IgG (right). Symbols are means of three (left) or two (right) independent experiments. ( C ) Binding of A35 and 135c Fabs to human PD-1 (left). Biolayer interferometry of Fabs–PD-1 interactions (right). Symbols are means of three independent experiments (left). ( D ) Binding of A35 and 135c scFvs-His to human PD-1 (left). Biolayer interferometry of scFvs–PD-1 interactions (right). Symbols are the mean of three independent experiments (left). ( E ) Second-generation CAR design (left). Biotinylated PD-1 binding to anti–PD-1 CARs. Representative flow cytometry and mean of four independent experiments. PD-1, programmed cell death protein 1; PD-L1, programmed cell death ligand 1; IgG, immunoglobulin G; Fab, fragment antigen-binding region; K D , equilibrium dissociation constant; scFv, single-chain variable fragment; His, histidine; CAR, chimeric antigen receptor; EGFRt, truncated epidermal growth factor receptor; HD, hinge domain; bPD1-CAR, blocking anti–PD-1 CAR; nbPD1-CAR, nonblocking anti–PD-1 CAR; UT, untransduced cells.

    Article Snippet: Kinetic analysis of A35/135C IgG, Fab, and scFv binding to PD-1 dimer protein was performed by biolayer interferometry immobilizing PD-1 biotinylated dimer (3 μg/ml) on streptavidin biosensors (Sartorius, Göttingen, Germany, catalog no. 18-5019) dipped into a solution of the Fab/scFv at different concentrations diluted in PBS (ranging from 40 to 240 nM), and the nm shift was recorded on the Octet.

    Techniques: In Silico, Binding Assay, Clone Assay, Competitive Binding Assay, Labeling, Flow Cytometry, Blocking Assay

    ( A ) Experimental design for engineering human anti–PD-1 CAR-T cells. ( B ) Expansion of primary anti–PD-1 CAR-T cells compared to the anti-CD19 CAR control ( n = 7 donors, seven independent experiments). ( C ) Cumulative data of transduction efficiency for donors reported in (D) and (E). The mean ± SD values eight donors, eight independent experiments, two to three replicates transduced with different MOI is shown. Two-way ANOVA, Tukey’s multiple comparison test. ( D ) Representative flow cytometry plots showing mCherry and PD-1 expression 6 days after transduction (gated on CD3 + living cells). ( E ) Correlation between PD-1 expression and the percentage of transduction efficiency. Cherry-positive and Cherry-negative population are shown. For each experiment the percentage of PD-1 + cells were assessed in Cherry-negative and Cherry-positive cells and normalized to PD-1 expression in untransduced T cells (set as 100%). Eight donors, eight independent experiments each with two to three internal replicates (with different MOI transduction: 1.4 ± 0.6). Simple linear regression tests. ( F ) CD8/CD4 ratio in mCherry ± T cells comparing anti–PD-1 CAR and CD19 CAR-T cells ± PD-1 knockout. Data are presented as mean ± SD values of eight donors, eight independent experiments, two to three internal replicates). Two-way ANOVA, Tukey’s multiple comparisons test. ( G ) Schematic representing the killing assay (left). Cumulative percentage of annexin/Aqua + CD4 + PD-1 + GFP + target cells after 24 hours of coculture (right). Mean ± SD of three donors and three independent experiments is shown. Two-way ANOVA, Tukey’s multiple comparison test. Only statistical differences are reported as follows: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001, and **** P ≤ 0.0001. HD, hinge domain; TMD, transmembrane domain; ICD, intracellular domain; LV, lentivirus; scFv, single-chain variable fragment; CAR, chimeric antigen receptor; bPD1-CAR, blocking anti–PD-1 CAR; nbPD1-CAR, nonblocking anti–PD-1 CAR; FMO, fluorescence minus one; UT, untransduced; KO, knockout.

    Journal: Science Advances

    Article Title: Targeting PD-1 + T cells with chimeric antigen receptors to reduce the HIV reservoir

    doi: 10.1126/sciadv.aeb7602

    Figure Lengend Snippet: ( A ) Experimental design for engineering human anti–PD-1 CAR-T cells. ( B ) Expansion of primary anti–PD-1 CAR-T cells compared to the anti-CD19 CAR control ( n = 7 donors, seven independent experiments). ( C ) Cumulative data of transduction efficiency for donors reported in (D) and (E). The mean ± SD values eight donors, eight independent experiments, two to three replicates transduced with different MOI is shown. Two-way ANOVA, Tukey’s multiple comparison test. ( D ) Representative flow cytometry plots showing mCherry and PD-1 expression 6 days after transduction (gated on CD3 + living cells). ( E ) Correlation between PD-1 expression and the percentage of transduction efficiency. Cherry-positive and Cherry-negative population are shown. For each experiment the percentage of PD-1 + cells were assessed in Cherry-negative and Cherry-positive cells and normalized to PD-1 expression in untransduced T cells (set as 100%). Eight donors, eight independent experiments each with two to three internal replicates (with different MOI transduction: 1.4 ± 0.6). Simple linear regression tests. ( F ) CD8/CD4 ratio in mCherry ± T cells comparing anti–PD-1 CAR and CD19 CAR-T cells ± PD-1 knockout. Data are presented as mean ± SD values of eight donors, eight independent experiments, two to three internal replicates). Two-way ANOVA, Tukey’s multiple comparisons test. ( G ) Schematic representing the killing assay (left). Cumulative percentage of annexin/Aqua + CD4 + PD-1 + GFP + target cells after 24 hours of coculture (right). Mean ± SD of three donors and three independent experiments is shown. Two-way ANOVA, Tukey’s multiple comparison test. Only statistical differences are reported as follows: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001, and **** P ≤ 0.0001. HD, hinge domain; TMD, transmembrane domain; ICD, intracellular domain; LV, lentivirus; scFv, single-chain variable fragment; CAR, chimeric antigen receptor; bPD1-CAR, blocking anti–PD-1 CAR; nbPD1-CAR, nonblocking anti–PD-1 CAR; FMO, fluorescence minus one; UT, untransduced; KO, knockout.

    Article Snippet: Kinetic analysis of A35/135C IgG, Fab, and scFv binding to PD-1 dimer protein was performed by biolayer interferometry immobilizing PD-1 biotinylated dimer (3 μg/ml) on streptavidin biosensors (Sartorius, Göttingen, Germany, catalog no. 18-5019) dipped into a solution of the Fab/scFv at different concentrations diluted in PBS (ranging from 40 to 240 nM), and the nm shift was recorded on the Octet.

    Techniques: Control, Transduction, Comparison, Flow Cytometry, Expressing, Knock-Out, Blocking Assay, Fluorescence

    ( A ) Generation of PD-1-transgenic luciferase + Jurkat cells with increasing PD-1 molecules number. ( B ) Luciferase-based killing assay of blocking and nonblocking CAR-T cells. Mean ± SD of three donors and three independent experiments is shown. ( C ) Phenotype of the K562 cell lines expressing either a PD-1-GFP-F fusion protein (K-PD-1-GFP-F) or PD-1 and GFP proteins with a ribosomal skipping motif in-between (K-PD-1-GFP-noF) used for the trogocytosis assay. Scale bars, 25 μm. ( D ) Representative flow cytometry 1 hour after coculture of K-PD-1-GFP-F or -PD-1-GFP-noF with anti–PD-1 CAR-T cells. ( E ) Cumulative data showing GFP expression in CAR-T cells over time. Mean ± SD values of six donors and four independent experiments is shown. Two-way ANOVA, Tukey’s multiple comparison test. ( F ) Phenotype of the K562 cell lines expressing either the wild-type PD-1 (K-PD-1 WT ) or a truncated PD-1 lacking (K-PD-1 t ) the intracellular domain used for the activation assay. ( G ) Cumulative percentage of CD25 + /CD71 + CAR-T cells after a 48 hours coculture with K-PD-1 WT or K-PD-1 t . Mean ± SD values of four to eight donors and seven independent experiments is shown. Two-way ANOVA, Tukey’s multiple comparison test. ( H ) Luciferase killing assay using PD-1 WT or PD-1 t luciferase + Jurkat cells as targets. Mean ± SD values of three donors, three independent experiments is shown. Two-way ANOVA, Tukey’s multiple comparison test was used to compare the same condition in JPD-1 versus JPD-1 t . Only statistical differences are reported as follows: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001, and **** P ≤ 0.0001. JPD-1, Jurkat-PD-1; CAR, chimeric antigen receptor; bPD1-CAR, blocking anti–PD-1 CAR; nbPD1-CAR, nonblocking anti–PD-1 CAR; E:T, effector:target; K-PD-1-GFP-F, K562 PD-1-GFP-F fusion protein; K-PD-1-GFP-noF, K562 PD-1 + GFP + ; K-PD-1, K562 PD-1 + ; K-PD-1-t, K562 PD-1 truncated; K-PD-1-KO, K562 PD-1 KO; JPD-1-t, Jurkat PD-1 truncated.

    Journal: Science Advances

    Article Title: Targeting PD-1 + T cells with chimeric antigen receptors to reduce the HIV reservoir

    doi: 10.1126/sciadv.aeb7602

    Figure Lengend Snippet: ( A ) Generation of PD-1-transgenic luciferase + Jurkat cells with increasing PD-1 molecules number. ( B ) Luciferase-based killing assay of blocking and nonblocking CAR-T cells. Mean ± SD of three donors and three independent experiments is shown. ( C ) Phenotype of the K562 cell lines expressing either a PD-1-GFP-F fusion protein (K-PD-1-GFP-F) or PD-1 and GFP proteins with a ribosomal skipping motif in-between (K-PD-1-GFP-noF) used for the trogocytosis assay. Scale bars, 25 μm. ( D ) Representative flow cytometry 1 hour after coculture of K-PD-1-GFP-F or -PD-1-GFP-noF with anti–PD-1 CAR-T cells. ( E ) Cumulative data showing GFP expression in CAR-T cells over time. Mean ± SD values of six donors and four independent experiments is shown. Two-way ANOVA, Tukey’s multiple comparison test. ( F ) Phenotype of the K562 cell lines expressing either the wild-type PD-1 (K-PD-1 WT ) or a truncated PD-1 lacking (K-PD-1 t ) the intracellular domain used for the activation assay. ( G ) Cumulative percentage of CD25 + /CD71 + CAR-T cells after a 48 hours coculture with K-PD-1 WT or K-PD-1 t . Mean ± SD values of four to eight donors and seven independent experiments is shown. Two-way ANOVA, Tukey’s multiple comparison test. ( H ) Luciferase killing assay using PD-1 WT or PD-1 t luciferase + Jurkat cells as targets. Mean ± SD values of three donors, three independent experiments is shown. Two-way ANOVA, Tukey’s multiple comparison test was used to compare the same condition in JPD-1 versus JPD-1 t . Only statistical differences are reported as follows: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001, and **** P ≤ 0.0001. JPD-1, Jurkat-PD-1; CAR, chimeric antigen receptor; bPD1-CAR, blocking anti–PD-1 CAR; nbPD1-CAR, nonblocking anti–PD-1 CAR; E:T, effector:target; K-PD-1-GFP-F, K562 PD-1-GFP-F fusion protein; K-PD-1-GFP-noF, K562 PD-1 + GFP + ; K-PD-1, K562 PD-1 + ; K-PD-1-t, K562 PD-1 truncated; K-PD-1-KO, K562 PD-1 KO; JPD-1-t, Jurkat PD-1 truncated.

    Article Snippet: Kinetic analysis of A35/135C IgG, Fab, and scFv binding to PD-1 dimer protein was performed by biolayer interferometry immobilizing PD-1 biotinylated dimer (3 μg/ml) on streptavidin biosensors (Sartorius, Göttingen, Germany, catalog no. 18-5019) dipped into a solution of the Fab/scFv at different concentrations diluted in PBS (ranging from 40 to 240 nM), and the nm shift was recorded on the Octet.

    Techniques: Transgenic Assay, Luciferase, Blocking Assay, Expressing, Trogocytosis Assay, Flow Cytometry, Comparison, Activation Assay

    ( A ) Experimental design. HLA-A2 + human T cells were isolated and activated ± prior editing of PD-1 and/or HLA-A2. ( B ) PD-1 expression in mCherry + and mCherry − cells on days 7 and 10. Mean of three to five donors in three to five independent experiments with one to two replicates transduced at different MOI (1.4 ± 0.6) for each independent experiments. ( C ) HLA-A2 expression in mCherry + and mCherry − cells on day 7 and 10. Mean of two donors in two independent experiments with one to two internal replicates (different MOI transduction: 1.4 ± 0.6). ( D ) Expansion folds of unedited versus edited CAR-T cells on day 10. Mean ± SD values of three to five donors in three to five independent experiments with one to two internal replicates (different MOI transduction: 1.4 ± 0.6). Statistics: Two-way ANOVA, Dunnett’s multiple comparison test was performed comparing each condition with the corresponding HLA-A2 CAR condition. ( E ) Cumulative data showing CD62L/CD45RA expression of mCherry + cells in edited versus unedited condition for PD-1 or HLA-A2. The mean ± SD values of three to five donors in three to five independent experiments with one to two internal replicates (different MOI transduction 1.4 ± 0.6) are shown. Kruskal-Wallis and Dunn’s multiple comparison test was performed comparing each condition with the corresponding HLA-A2 CAR condition. ( F ) Representative flow cytometry showing TIM-3 and LAG-3 expression in the different CAR populations on day 10 (left). Cumulative percentage of LAG-3 + /TIM-3 + double-positive cells (right). Paired data of three to five donors in three to five independent experiments with one to two internal replicates (different MOI transduction: 1.4 ± 0.6) are shown. Two-way ANOVA, Dunnett’s multiple comparison test was performed comparing each condition with the corresponding HLA-A2 CAR condition. Only statistical differences are reported as follows: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001, and **** P ≤ 0.0001.

    Journal: Science Advances

    Article Title: Targeting PD-1 + T cells with chimeric antigen receptors to reduce the HIV reservoir

    doi: 10.1126/sciadv.aeb7602

    Figure Lengend Snippet: ( A ) Experimental design. HLA-A2 + human T cells were isolated and activated ± prior editing of PD-1 and/or HLA-A2. ( B ) PD-1 expression in mCherry + and mCherry − cells on days 7 and 10. Mean of three to five donors in three to five independent experiments with one to two replicates transduced at different MOI (1.4 ± 0.6) for each independent experiments. ( C ) HLA-A2 expression in mCherry + and mCherry − cells on day 7 and 10. Mean of two donors in two independent experiments with one to two internal replicates (different MOI transduction: 1.4 ± 0.6). ( D ) Expansion folds of unedited versus edited CAR-T cells on day 10. Mean ± SD values of three to five donors in three to five independent experiments with one to two internal replicates (different MOI transduction: 1.4 ± 0.6). Statistics: Two-way ANOVA, Dunnett’s multiple comparison test was performed comparing each condition with the corresponding HLA-A2 CAR condition. ( E ) Cumulative data showing CD62L/CD45RA expression of mCherry + cells in edited versus unedited condition for PD-1 or HLA-A2. The mean ± SD values of three to five donors in three to five independent experiments with one to two internal replicates (different MOI transduction 1.4 ± 0.6) are shown. Kruskal-Wallis and Dunn’s multiple comparison test was performed comparing each condition with the corresponding HLA-A2 CAR condition. ( F ) Representative flow cytometry showing TIM-3 and LAG-3 expression in the different CAR populations on day 10 (left). Cumulative percentage of LAG-3 + /TIM-3 + double-positive cells (right). Paired data of three to five donors in three to five independent experiments with one to two internal replicates (different MOI transduction: 1.4 ± 0.6) are shown. Two-way ANOVA, Dunnett’s multiple comparison test was performed comparing each condition with the corresponding HLA-A2 CAR condition. Only statistical differences are reported as follows: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001, and **** P ≤ 0.0001.

    Article Snippet: Kinetic analysis of A35/135C IgG, Fab, and scFv binding to PD-1 dimer protein was performed by biolayer interferometry immobilizing PD-1 biotinylated dimer (3 μg/ml) on streptavidin biosensors (Sartorius, Göttingen, Germany, catalog no. 18-5019) dipped into a solution of the Fab/scFv at different concentrations diluted in PBS (ranging from 40 to 240 nM), and the nm shift was recorded on the Octet.

    Techniques: Isolation, Expressing, Transduction, Comparison, Flow Cytometry

    ( A ) Experimental design. Mice received adoptive cell transfer (ACT) of anti–PD-1 CARs versus untransduced (UT) T cells under ART that was interrupted after 2 weeks (three independent experiments). The reporter used for detecting CAR-positive cells was EGFRt. ( B ) Kaplan-Meyer curve showing the percentage of mice maintaining viral control. All mice were included in the analysis. Log-rank (Mantel-Cox) test was performed using the UT control condition as reference. ( C ) Plasma viral load overtime (UT, n = 6; bPD1-CAR, n = 9; nbPD1-CAR, n = 7). ( D ) Percentage of CAR-T cells (EGFRt + gated in CD45 + cells) in the spleen, BM, and blood at the time of euthanasia. Mean ± SD is shown. bPD1-CAR, n = 9, nbPD1-CAR, n = 7. ( E ) Percentage of CD8 + , CD4 + , and CD4 − CAR-T cells detected in the spleen, BM, and blood (bPD1-CAR, n = 3, nbPD1-CAR, n = 4). Mean ± SD is shown. ( F ) Correlation between the percentage of CAR-T cells (defined as CD45 + EGFRt + cells) and the time of viral rebound (bPD1-CAR, n = 9; nbPD1-CAR, n = 7). Simple linear regression was used. ( G ) Correlation between CD4 + PD-1 + cells (gated in huCD45 + EGFRt − cells) and the time of viral rebound (bPD1-CAR, n = 9; nbPD1-CAR, n = 7). Simple linear regression was used. ( H ) Correlation between CD4 + PD-1 + cells (gated in huCD45 + EGFRt − cells) and CAR-T cells detection (bPD1-CAR, n = 9; nbPD1-CAR, n = 7). Simple linear regression was used. ( I ) Correlation between HIV integrated DNA in CD4 + EGFRt- sorted cells from spleen and BM and time of viral rebound (bPD1-CAR, n = 3; nbPD1-CAR, n = 3). Simple linear regression was used. ( J ) Correlation between HIV-integrated DNA in CD4 + EGFRt − sorted cells from spleen and BM and CAR-T cells detection (bPD1-CAR, n = 3; nbPD1-CAR, n = 3). Simple linear regression was used. ART, antiretroviral therapy; ACT, adoptive cell transfer; VC, viral control.

    Journal: Science Advances

    Article Title: Targeting PD-1 + T cells with chimeric antigen receptors to reduce the HIV reservoir

    doi: 10.1126/sciadv.aeb7602

    Figure Lengend Snippet: ( A ) Experimental design. Mice received adoptive cell transfer (ACT) of anti–PD-1 CARs versus untransduced (UT) T cells under ART that was interrupted after 2 weeks (three independent experiments). The reporter used for detecting CAR-positive cells was EGFRt. ( B ) Kaplan-Meyer curve showing the percentage of mice maintaining viral control. All mice were included in the analysis. Log-rank (Mantel-Cox) test was performed using the UT control condition as reference. ( C ) Plasma viral load overtime (UT, n = 6; bPD1-CAR, n = 9; nbPD1-CAR, n = 7). ( D ) Percentage of CAR-T cells (EGFRt + gated in CD45 + cells) in the spleen, BM, and blood at the time of euthanasia. Mean ± SD is shown. bPD1-CAR, n = 9, nbPD1-CAR, n = 7. ( E ) Percentage of CD8 + , CD4 + , and CD4 − CAR-T cells detected in the spleen, BM, and blood (bPD1-CAR, n = 3, nbPD1-CAR, n = 4). Mean ± SD is shown. ( F ) Correlation between the percentage of CAR-T cells (defined as CD45 + EGFRt + cells) and the time of viral rebound (bPD1-CAR, n = 9; nbPD1-CAR, n = 7). Simple linear regression was used. ( G ) Correlation between CD4 + PD-1 + cells (gated in huCD45 + EGFRt − cells) and the time of viral rebound (bPD1-CAR, n = 9; nbPD1-CAR, n = 7). Simple linear regression was used. ( H ) Correlation between CD4 + PD-1 + cells (gated in huCD45 + EGFRt − cells) and CAR-T cells detection (bPD1-CAR, n = 9; nbPD1-CAR, n = 7). Simple linear regression was used. ( I ) Correlation between HIV integrated DNA in CD4 + EGFRt- sorted cells from spleen and BM and time of viral rebound (bPD1-CAR, n = 3; nbPD1-CAR, n = 3). Simple linear regression was used. ( J ) Correlation between HIV-integrated DNA in CD4 + EGFRt − sorted cells from spleen and BM and CAR-T cells detection (bPD1-CAR, n = 3; nbPD1-CAR, n = 3). Simple linear regression was used. ART, antiretroviral therapy; ACT, adoptive cell transfer; VC, viral control.

    Article Snippet: Kinetic analysis of A35/135C IgG, Fab, and scFv binding to PD-1 dimer protein was performed by biolayer interferometry immobilizing PD-1 biotinylated dimer (3 μg/ml) on streptavidin biosensors (Sartorius, Göttingen, Germany, catalog no. 18-5019) dipped into a solution of the Fab/scFv at different concentrations diluted in PBS (ranging from 40 to 240 nM), and the nm shift was recorded on the Octet.

    Techniques: Control, Clinical Proteomics

    ( A ) Immunofluorescence multicolor confocal images of spleens of four control and seven treated hu-mice with detectable CAR-T cells (untransduced treated, n = 4; nbPD1-CAR-T treated, n = 4; bPD1-CAR-T treated, n = 3) 8 to 10 weeks after ACT. The following stainings were performed: DAPI (blue), CD4 (green), CD20 (red), PD-1 (magenta), CD8 (yellow), Grzb (cyan), and CD57 (gray). Scale bars, 100 μm. ( B ) Histo-cytometry analysis showing CD4 + PD-1 + , and CD8 + PD-1 + cell counts/mm 2 in the CD20-enriched areas and CD20 + cell counts/mm 2 in the CD4-enriched zones. Each dot represents a CD20- or CD4-enriched region ( n = 154 and n = 158). Median is shown. Kruskal-Wallis and Dunn’s multiple comparison test. All control mice ( n = 6) and treated mice with detectable CARs ( n = 7) that survived until the end of the experiment were included. ( C ) Mean distance between each B cell and its five nearest neighbors in the CD20-enriched regions in the 13 spleens based on IF staining data. Median is shown. Kruskal-Wallis and Dunn’s multiple comparison test was performed. ( D ) Combined IF and RNAscope for CD8 (green), DAPI (blue), HIV RNA (red), and CAR RNA (magenta) performed on the spleen of one representative control mouse (S2), one HIV suppressed responder mouse (S11), and one nonresponder mouse with viral rebound (S5) 8 weeks after ACT. Scale bars, 50 μm. ( E ) Histo-cytometry analysis showing absolute counts of CAR RNA events (gated on CD8 + cells) and HIV RNA events for each mouse tissue. Two-way ANOVA, Tukey’s multiple comparison test. Only statistical differences are reported as follows: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001, and **** P ≤ 0.0001. S, spleen; bCAR, blocking anti–PD-1 CAR; nbCAR, nonblocking anti–PD-1 CAR; ACT, adoptive cell transfer; UT, untransduced T cells.

    Journal: Science Advances

    Article Title: Targeting PD-1 + T cells with chimeric antigen receptors to reduce the HIV reservoir

    doi: 10.1126/sciadv.aeb7602

    Figure Lengend Snippet: ( A ) Immunofluorescence multicolor confocal images of spleens of four control and seven treated hu-mice with detectable CAR-T cells (untransduced treated, n = 4; nbPD1-CAR-T treated, n = 4; bPD1-CAR-T treated, n = 3) 8 to 10 weeks after ACT. The following stainings were performed: DAPI (blue), CD4 (green), CD20 (red), PD-1 (magenta), CD8 (yellow), Grzb (cyan), and CD57 (gray). Scale bars, 100 μm. ( B ) Histo-cytometry analysis showing CD4 + PD-1 + , and CD8 + PD-1 + cell counts/mm 2 in the CD20-enriched areas and CD20 + cell counts/mm 2 in the CD4-enriched zones. Each dot represents a CD20- or CD4-enriched region ( n = 154 and n = 158). Median is shown. Kruskal-Wallis and Dunn’s multiple comparison test. All control mice ( n = 6) and treated mice with detectable CARs ( n = 7) that survived until the end of the experiment were included. ( C ) Mean distance between each B cell and its five nearest neighbors in the CD20-enriched regions in the 13 spleens based on IF staining data. Median is shown. Kruskal-Wallis and Dunn’s multiple comparison test was performed. ( D ) Combined IF and RNAscope for CD8 (green), DAPI (blue), HIV RNA (red), and CAR RNA (magenta) performed on the spleen of one representative control mouse (S2), one HIV suppressed responder mouse (S11), and one nonresponder mouse with viral rebound (S5) 8 weeks after ACT. Scale bars, 50 μm. ( E ) Histo-cytometry analysis showing absolute counts of CAR RNA events (gated on CD8 + cells) and HIV RNA events for each mouse tissue. Two-way ANOVA, Tukey’s multiple comparison test. Only statistical differences are reported as follows: * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001, and **** P ≤ 0.0001. S, spleen; bCAR, blocking anti–PD-1 CAR; nbCAR, nonblocking anti–PD-1 CAR; ACT, adoptive cell transfer; UT, untransduced T cells.

    Article Snippet: Kinetic analysis of A35/135C IgG, Fab, and scFv binding to PD-1 dimer protein was performed by biolayer interferometry immobilizing PD-1 biotinylated dimer (3 μg/ml) on streptavidin biosensors (Sartorius, Göttingen, Germany, catalog no. 18-5019) dipped into a solution of the Fab/scFv at different concentrations diluted in PBS (ranging from 40 to 240 nM), and the nm shift was recorded on the Octet.

    Techniques: Immunofluorescence, Control, Cytometry, Comparison, Staining, RNAscope, Blocking Assay