Journal: The journal of applied laboratory medicine

Article Title: Plasma Protein Profiling by Proximity Extension Assay Technology Reveals Novel Biomarkers of Traumatic Brain Injury-A Pilot Study.

doi: 10.1093/jalm/jfab004

Figure Lengend Snippet: Fig. 1. Normalized protein expression (NPX) of plasma biomarkers in surviving [sTBI (S); n 5 6] and non- surviving sTBI patients [sTBI (NS); n 5 4] compared with matched healthy controls (CTR, n 5 10). Biomarkers shown are (A) ST2, (B) IL6, (C) TNNI3, (D) TNFR-2, (E) CHI3L1, and (F) EN-RAGE. Reference groups shown are ovarian cancer patients (OvCa, n 5 32) and dementia patients (mild cognitive impair- ment, MCI, n 5 18; Alzheimer disease, AD, n 5 12). Dotted lines represent group median, bars show interquartile range. Mann Whitney U test; *P < 0.05, **P < 0.01, ***P < 0.001, ns, not significant. For more discussion see text.

Article Snippet: All sTBI and matched control samples were analyzed in duplicate using the commercially available ELISA kits, DRT200 TNFR2 Quantikine kit and the DST200 ST2 Quantikine kit (R&D Systems), according to manufacturer’s instructions.

Techniques: Expressing, Clinical Proteomics, MANN-WHITNEY

Journal: The journal of applied laboratory medicine

Article Title: Plasma Protein Profiling by Proximity Extension Assay Technology Reveals Novel Biomarkers of Traumatic Brain Injury-A Pilot Study.

doi: 10.1093/jalm/jfab004

Figure Lengend Snippet: Fig. 3. Plasma concentration of ST2 and TNFR-2 in healthy controls (n 5 10) and sTBI patients (n 5 10), as measured by commercially available ELISAs. Mann Whitney U test; ****P < 0.0001. For more details see text.

Article Snippet: All sTBI and matched control samples were analyzed in duplicate using the commercially available ELISA kits, DRT200 TNFR2 Quantikine kit and the DST200 ST2 Quantikine kit (R&D Systems), according to manufacturer’s instructions.

Techniques: Clinical Proteomics, Concentration Assay, MANN-WHITNEY

Journal: The journal of applied laboratory medicine

Article Title: Plasma Protein Profiling by Proximity Extension Assay Technology Reveals Novel Biomarkers of Traumatic Brain Injury-A Pilot Study.

doi: 10.1093/jalm/jfab004

Figure Lengend Snippet: Fig. 4. Comparison between commercially available proximity extension assays (Olink Proteomics) and ELISAs. Plasma ST2 was measured in (A) healthy controls (n 5 10); Spearman’s correlation (rs) 5 0.8909, P 5 0.0011, and (B) sTBI patients (n 5 10); rs 5 0.9758, P < 0.0001. Plasma TNFR-2 was measured in (A) healthy controls (n 5 10); rs 5 0.9030, P 5 0.0008, and (B) sTBI patients (n 5 10); rs 5 0.8667, P 5 0.0022. Black line shows line of best fit.

Article Snippet: All sTBI and matched control samples were analyzed in duplicate using the commercially available ELISA kits, DRT200 TNFR2 Quantikine kit and the DST200 ST2 Quantikine kit (R&D Systems), according to manufacturer’s instructions.

Techniques: Comparison, Clinical Proteomics

Journal: The Journal of Experimental Medicine

Article Title: Dietary restriction improves repopulation but impairs lymphoid differentiation capacity of hematopoietic stem cells in early aging

doi: 10.1084/jem.20151100

Figure Lengend Snippet: IGF1 injection reverts the increases in HSC quiescence but not the suppression of lymphopoiesis in mice exposed to DR. (A) ELISA-determined serum level of IGF1 in mice treated with DR or AL for 4 d or refed with an AL diet for 3 d after 4 d of DR ( n = 3–4 mice per group; n = 2 independent experiments). (B) ELISA-determined serum level of IGF1 in 2-mo-old p53 −/− mice and p53 +/+ mice (littermates) that were treated with DR or AL for 2 wk ( n = 3–4 mice per group; n = 2 independent experiments). (C–G) Mice were treated with a DR or AL diet for 8 d. In parallel, recombinant human IGF1 protein or vehicle control was subcutaneously injected twice per day at a dose of 500 µg/kg ( n = 4 mice per group; n = 2 independent experiments). FACS analysis was performed 24 h after the last injection on freshly isolated BM cells: percentage of HSCs in the indicated cell cycle stages (C), total numbers of CLPs (D), pro–B cells (E), MEPs (F), and proEry’s (G) per million total BM cells are show. AL con, control-injected AL mice; AL IGF1, IGF1-injected AL mice; DR con, control-injected DR mice; DR IGF1, IGF1-injected DR mice. Data are displayed as mean ± SEM. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001 by one-way ANOVA. ns, not significant.

Article Snippet: ELISA for detecting IGF1, IL-6, IL-7, and total corticosterone was performed using the IGF-I mouse/rat ELISA kit (Mediagnost), the Mouse IL-6 Quantikine ELISA kit (R&D Systems), the Mouse IL-7 Quantikine ELISA kit (R&D Systems), and Corticosterone EIA kit (ARBOR ASSAYS), respectively, according to the manufacturer’s instructions.

Techniques: Injection, Enzyme-linked Immunosorbent Assay, Recombinant, Control, Isolation

Journal: The Journal of Experimental Medicine

Article Title: Dietary restriction improves repopulation but impairs lymphoid differentiation capacity of hematopoietic stem cells in early aging

doi: 10.1084/jem.20151100

Figure Lengend Snippet: IL-7 or IL-6 injection reverts suppression of lymphopoiesis in mice exposed to DR. (A) ELISA-determined serum level of IL-7 in mice treated with DR or AL for 7 d or refed with AL diet for 3 d after 7 d of DR ( n = 3–4 mice per group; n = 2 independent experiments). One-way ANOVA. (B–G) 2-mo-old mice were treated with DR or AL diet for 9 d. In parallel, mouse IL-7 protein or vehicle control was subcutaneously injected for 9 d at a dose of 50 µg/kg once per day ( n = 5 mice per group; n = 2 independent experiments). Mice were sacrificed for analysis 24 h after the last injection. (B–D) FACS analysis was performed on freshly isolated BM cells to determine the total number of CLPs (B) and MEPs (C), and the percentage of CLPs (D) of DR mice in the indicated phases of the cell cycle (FACS analysis by Ki67 and DAPI staining). (E and F) the ratio of CLPs versus lymphoid-biased HSCs (E) and of pro–B cells versus CLPs (F). (G) The mRNA expression levels of genes that are associated with lymphoid cell differentiation and erythroid cell differentiation were determined by qRT-PCR in freshly isolated lymphoid-biased HSCs under the indicated treatment conditions. One-way ANOVA (B, C, and E–G) or unpaired two-tailed Student’s t test (D) was used. (H) ELISA-determined serum level of IL-6 in mice treated with DR or AL for 7 d or refed with AL diet for 3 d after 7 d of DR ( n = 3–4 mice per group; n = 2 independent experiments). One-way ANOVA was used. (I) 2-mo-old mice were treated with a DR or AL diet for 5 d. In parallel, mouse IL-6 protein or vehicle control was subcutaneously injected into mice daily at a dose of 50 µg/kg ( n = 4 mice per group; n = 2 independent experiments). The total number of CLPs was determined by FACS analysis, 24 h after the last injection. One-way ANOVA analysis. (J–P) ADX or sham surgery was performed on 2-mo-old mice. 10 d after operation, mice were treated with DR or AL for 9 d ( n = 4–5 mice per group; n = 2 independent experiments). Mice of the indicated cohorts were analyzed as follows: (J, O, and P) ELISA-determined serum levels of total corticosterone (J), IL-7 (O), and IL-6 (P); (K–M) FACS analysis of the total number of CLPs (K) and MEPs (M) per million BM cells and of the percentage of CLPs (L) of DR mice in different stages of the cell cycle (by Ki67 and DAPI staining); (N) expression of lymphoid- and erythroid-specific genes in lymphoid-biased HSCs of mice of the indicated cohorts. One-way ANOVA (J, K, and M–P) or unpaired two-tailed Student’s t test (L) was used. AL con, control-injected AL mice or sham-operated AL mice; AL IL-7, IL-7–injected AL mice; DR con, control-injected DR mice or sham-operated DR mice; DR IL-7, IL-7–injected DR mice; AL IL-6, IL-6–injected AL mice; DR IL-6, IL-6–injected DR mice; AL ADX, AL mice with ADX; DR ADX, DR mice with ADX. Data are displayed as mean ± SEM. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001. ns, not significant.

Article Snippet: ELISA for detecting IGF1, IL-6, IL-7, and total corticosterone was performed using the IGF-I mouse/rat ELISA kit (Mediagnost), the Mouse IL-6 Quantikine ELISA kit (R&D Systems), the Mouse IL-7 Quantikine ELISA kit (R&D Systems), and Corticosterone EIA kit (ARBOR ASSAYS), respectively, according to the manufacturer’s instructions.

Techniques: Injection, Enzyme-linked Immunosorbent Assay, Control, Isolation, Staining, Expressing, Cell Differentiation, Quantitative RT-PCR, Two Tailed Test

Journal: Cancer immunology research

Article Title: Antitumor effects of CAR T cells redirected to the EDB splice variant of fibronectin

doi: 10.1158/2326-6066.CIR-20-0280

Figure Lengend Snippet: (A) Scheme of retroviral vectors encoding the EDB-CAR, a 2A sequence, and truncated CD19 (tCD19); SH: short hinge; TM: transmembrane domain. (B-C) EDB-CAR expression was determined on T cells 7 days post-transduction by flow cytometric analysis. (B) Representative histogram (Black-filled line: NT T cells; Red-filled line: EDB-CAR transduced T cells) and (C) summary plot (n=7; Student’s t-test; ****p<0.0001). (D) NT and EDB-CAR T cells were incubated for 24 hours with increasing concentrations of rhFN-EDB-coated wells. IFNγ production in supernatants was determined by ELISA (n=4; two-way ANOVA; ***p<0.001; ****p<0.0001). (E) EDB expression of LM7, A673, A549, and U87 tumor cells, and two primary fibroblast cell lines (Fib 1, Fib 2) determined by RT-qPCR; dotted line represents the ΔCt score above which samples are considered positive. (F) NT and EDB-CAR T cells were incubated for 48 hours with EDB-positive tumor cells. IFNγ and IL2 production in supernatants was determined by ELISA (two-way ANOVA; ****p<0.0001). (G) Cytolytic activity of NT or EDB-CAR T cells at a E:T ratio of 4:1 against EDB-positive tumor cells (MTS assay; n=3; two-way ANOVA; *p<0.05; **p<0.01; ***p<0.001). (H) Cytolytic activity of NT or EDB-CAR T cells against primary fibroblasts (data of Fib 1 and Fib 2 was combined) at the indicated E:T ratios, with A549 serving as controls (n=3; two-way ANOVA; ****p<0.0001). (I) NT, EDB-CAR, or mutEDB-CAR T cells were incubated for 48 hours with U87 or U87FN−/− cells. IFNγ production in supernatants was determined by ELISA (n=3; two-way ANOVA; ****p<0.0001). (J) Cytolytic activity of NT, EDB-CAR, or mutEDB-CAR T cells against U87 or U87FN−/− cells (n=3; two-way ANOVA; ****p<0.0001). Mean+SEM is shown in panels.

Article Snippet: Cytokines were measured using human IFNγ and IL2 ELISA kits (R&D Systems).

Techniques: Retroviral, Sequencing, Expressing, Transduction, Incubation, Enzyme-linked Immunosorbent Assay, Quantitative RT-PCR, Activity Assay, MTS Assay