Journal: BMC Cancer

Article Title: Dickkopf-related protein 1, a new biomarker for local immune status and poor prognosis among patients with colorectal liver Oligometastases: a retrospective study

doi: 10.1186/s12885-019-6399-1

Figure Lengend Snippet: a and b The numbers of total CD3+ TILs a and CD3 + CD8+ TILs b in invasive margin (IM) with low and high serum DKK1. c Pearson correlation test of the correlation between serum DKK1 level and the number of CD3 + CD8+ T cells in IM. d Comparison of CD8+/CD3+ ratio in IM between two groups of CRCLOM. e Typical image of immunofluorescence staining of metastasis slices. Scale bars represent 100 μm. *: 0.01 ≤ P < 0.05

Article Snippet: Paraffin-embedded slides of liver metastatic lesion specimens were stained using the primary monoclonal antibodies against CD3 (1:100, Abcam, Cambridge, UK) and CD8 (1100, Abcam, Cambridge, UK) proteins.

Techniques: Immunofluorescence, Staining

Journal: PLoS ONE

Article Title: Identification of the ER-resident E3 ubiquitin ligase RNF145 as a novel LXR-regulated gene

doi: 10.1371/journal.pone.0172721

Figure Lengend Snippet: ( A ) Expression of Rnf145 was evaluated in the indicated mouse tissues by qPCR. Bars indicate mean ± SD (n = 3) ( B , C ) The indicated ( B ) human and ( C ) mouse cell lines and primary macrophages were cultured in lipoprotein-depletion medium for 16 hours and subsequently treated for 6 hours with 1μM GW3965 (GW). ( D ) THP1 cells were cultured in sterol depletion medium for 16 hrs and then treated with 1μM GW3965 (GW) and 100nM LG100268 (LG) for 6 hrs as indicated. Subsequently, ABCA1 and RNF145 expression was determined by qPCR and each bar and error represents the mean fold-change of ligand-treated cells over sterol-depleted cells ± SD (n≥3).

Article Snippet: Membranes were probed with the following antibodies: LDLR (Abcam, clone EP1553Y, 1:4000), tubulin (Sigma, clone DM1A, ascites fluid, 1:5000), ABCA1 (Novus Biologicals, NB400-105, 1:1000), FLAG (Sigma, clone M2, 1:1000), GFP (Santa Cruz sc-9996, 1:500), Myc (Santa Cruz 9E10, 1:3000), HA (Covance, clone 16B12, ascites fluid, 1:6000), HMGCR (rabbit polyserum was a kind gift from Dr. Peter Edwards, UCLA), SREBP2 (BD Biosciences, clone 1C6, 1:1000), SREBP1 (ThermoFisher, clone 2A4, 1:1000), actin (Merck Millipore, clone C4, 1:5000), V5 (Invitrogen, 46–0705, 1:3000), RNF145 (Abgent AP18281b, 1:8000), and ubiquitin (Enzo life Sciences, clone FK2, 1:1000).

Techniques: Expressing, Cell Culture

Journal: PLoS ONE

Article Title: Identification of the ER-resident E3 ubiquitin ligase RNF145 as a novel LXR-regulated gene

doi: 10.1371/journal.pone.0172721

Figure Lengend Snippet: ( A ) RAW264.7 macrophages were treated with 5μg/mL Actinomycin D (ActD) for the indicated time and expression of Abca1 and Rnf145 was determined by qPCR and plotted as mean ± SD relative to untreated cells (n = 3), ( B ) HepG2 and RAW264.7 cells were treated with 1μM GW3965 for 6 hours in the presence or absence of 5μg/ml actinomycinD for 4 hours, after which expression of the indicated genes was measured by qPCR. Bars indicate mean ± SD (n = 3) ( C , D ) THP1 macrophages were cultured in sterol-depletion medium for 16 hrs and then treated with ( C ) 1μM GW3965 (GW) for the indicated time, or ( D ) with the indicated concentration of GW3965 for 4 hrs. Subsequently, gene expression was evaluated qPCR and bars indicate mean ± SD (n = 3)

Article Snippet: Membranes were probed with the following antibodies: LDLR (Abcam, clone EP1553Y, 1:4000), tubulin (Sigma, clone DM1A, ascites fluid, 1:5000), ABCA1 (Novus Biologicals, NB400-105, 1:1000), FLAG (Sigma, clone M2, 1:1000), GFP (Santa Cruz sc-9996, 1:500), Myc (Santa Cruz 9E10, 1:3000), HA (Covance, clone 16B12, ascites fluid, 1:6000), HMGCR (rabbit polyserum was a kind gift from Dr. Peter Edwards, UCLA), SREBP2 (BD Biosciences, clone 1C6, 1:1000), SREBP1 (ThermoFisher, clone 2A4, 1:1000), actin (Merck Millipore, clone C4, 1:5000), V5 (Invitrogen, 46–0705, 1:3000), RNF145 (Abgent AP18281b, 1:8000), and ubiquitin (Enzo life Sciences, clone FK2, 1:1000).

Techniques: Expressing, Cell Culture, Concentration Assay, Gene Expression

Journal: PLoS ONE

Article Title: Identification of the ER-resident E3 ubiquitin ligase RNF145 as a novel LXR-regulated gene

doi: 10.1371/journal.pone.0172721

Figure Lengend Snippet: ( A ) LXR ChIP-seq experiments in human THP1 cells (GSE28319) and RAW macrophage-like cells (GSE50944) were analyzed and used to identify active LXREs within the Rnf145/Rnf145 loci, as graphically illustrated. ( B , C ) Genomic location of the identified LXREs. In bold, nucleotides that were mutated to disrupt LXR binding ( C ) A 1kb genomic region upstream of the transcriptional start site of hRNF145 was cloned into a pGL3basic. The putative LXRE was also mutated as indicated above. The empty, RNF145 WT , RNF145 MUT , and ABCA1 reporter plasmids were co-transfected with or without RXRα and LXRα expression plasmids in HEK 293T cells. 24 hours post-transfection the cells were treated with 1μM GW3965 (LXR) and 100nM LG100268 (RXR) for 24 hours and measured for luciferase signal (n≥3). ( D ) Cells were transfected with an empty or a tandem LXRE-containing pGL2 as in C . In all luciferase experiments the transfection efficiency was normalized to co-transfected Renilla luciferase. Bars report normalized chemiluminescence relative to untreated control ± SD (n = 3).

Article Snippet: Membranes were probed with the following antibodies: LDLR (Abcam, clone EP1553Y, 1:4000), tubulin (Sigma, clone DM1A, ascites fluid, 1:5000), ABCA1 (Novus Biologicals, NB400-105, 1:1000), FLAG (Sigma, clone M2, 1:1000), GFP (Santa Cruz sc-9996, 1:500), Myc (Santa Cruz 9E10, 1:3000), HA (Covance, clone 16B12, ascites fluid, 1:6000), HMGCR (rabbit polyserum was a kind gift from Dr. Peter Edwards, UCLA), SREBP2 (BD Biosciences, clone 1C6, 1:1000), SREBP1 (ThermoFisher, clone 2A4, 1:1000), actin (Merck Millipore, clone C4, 1:5000), V5 (Invitrogen, 46–0705, 1:3000), RNF145 (Abgent AP18281b, 1:8000), and ubiquitin (Enzo life Sciences, clone FK2, 1:1000).

Techniques: ChIP-sequencing, Binding Assay, Clone Assay, Transfection, Expressing, Luciferase, Control

Journal: Molecules and Cells

Article Title: Regulation of Hepatic Gluconeogenesis by Nuclear Receptor Coactivator 6

doi: 10.14348/molcells.2022.2222

Figure Lengend Snippet: (A) Schematic representation of the liver-specific knockout strategy used for the Ncoa6 gene, based on the Cre-LoxP system. Blue arrows denote the specific primer binding sites for genotyping. (B-D) The liver specific knockout of Ncoa6 was confirmed at the level of genomic DNA (gDNA), mRNA and protein. gDNA and mRNA were isolated from the liver, muscle, kidney and white adipose tissue (WAT) of Ncoa6 fl/fl /Cre +/- (LKO) and Ncoa6 fl/fl /Cre -/- (F/F) mice. The Ncoa6 gDNA levels were determined by conventional PCR (B). The Ncoa6 mRNA and NCOA6 protein levels were determined by qRT-PCR (C; n = 4) and western blotting (D; n = 3), respectively. mRNA data are presented as the mean ± SEM; *** P < 0.001 by t -test comparisons of the genotypes.

Article Snippet: After membrane blocking with 5% skim milk, the membrane was probed with primary antibodies against NCOA6 (Cat. No. NB200-335; Novus Biologicals, USA), PCK1 (Cat. No. sc-32879; Santa Cruz Biotechnology, USA), CREB (Cat. No. 9197; Cell Signaling Technology, USA), p-CREB (Cat. No. 9198S; Cell Signaling Technology), or α-tubulin (Cat. No. T9026; Sigma-Aldrich).

Techniques: Knock-Out, Binding Assay, Isolation, Quantitative RT-PCR, Western Blot

Journal: Molecules and Cells

Article Title: Regulation of Hepatic Gluconeogenesis by Nuclear Receptor Coactivator 6

doi: 10.14348/molcells.2022.2222

Figure Lengend Snippet: (A) The blood glucose levels were measured in Ncoa6 LKO (n = 8) and WT (F/F, n = 9) mice after 16 h fasting. (B) Improved glucose tolerance in the Ncoa6 LKO mice. Intraperitoneal glucose tolerance tests were performed in 10-week-old mice (n = 9 for F/F, n = 8 for LKO) after a 16 h fast. (C) The serum insulin levels were measured in Ncoa6 LKO (n = 6 for fed, n = 7 for fasted) and WT mice (n = 7 for both fed and fasted) before and after a 16 h fast. (D) Enhanced insulin sensitivity in Ncoa6 LKO mice. Intraperitoneal insulin tolerance tests were performed in 10-week-old Ncoa6 LKO (n = 13) and WT mice (n = 10) after a 6 h fast. (E) Pyruvate tolerance was determined in 10-week-old Ncoa6 LKO (n = 11) and WT (n = 10) mice via an intraperitoneal injection with pyruvate (2 g/kg body weight) after overnight fasting for 16 h. AUC values for GTT, ITT, and PTT were calculated and are presented on the right of each graph. Data are presented as the mean ± SEM; * P < 0.05, ** P < 0.01 by t -test comparisons of the genotypes. WT, wild type; GTT, glucose tolerance test; ITT, insulin tolerance test; PTT, pyruvate tolerance test.

Article Snippet: After membrane blocking with 5% skim milk, the membrane was probed with primary antibodies against NCOA6 (Cat. No. NB200-335; Novus Biologicals, USA), PCK1 (Cat. No. sc-32879; Santa Cruz Biotechnology, USA), CREB (Cat. No. 9197; Cell Signaling Technology, USA), p-CREB (Cat. No. 9198S; Cell Signaling Technology), or α-tubulin (Cat. No. T9026; Sigma-Aldrich).

Techniques: Injection

Journal: Molecules and Cells

Article Title: Regulation of Hepatic Gluconeogenesis by Nuclear Receptor Coactivator 6

doi: 10.14348/molcells.2022.2222

Figure Lengend Snippet: (A) Decreased induction of gluconeogenic gene transcripts in the Ncoa6 LKO liver following fasting. Liver RNAs were isolated from Ncoa6 LKO and WT mice after a 24 h fast (n = 6 for each group). The mRNA levels of Pc , Pck1 , and G6pc were determined by real-time qRT-PCR. (B) Attenuated induction of PCK protein in the Ncoa6 LKO liver after fasting. Protein expression was analyzed in the liver of ad libitum fed mice and 16 h fasted mice (n = 7 for each group) by western blot analysis. Intensities of the protein bands were measured using the ImageJ program. (C) Glucagon tolerance of Ncoa6 LKO mice (n = 10 for F/F, n = 14 for LKO). Ten-week-old mice were injected intraperitoneally with glucagon (15 µg/kg) after 6 h of fasting. Blood samples were prepared at 0, 30, 60, 90, and 120 min after the glucagon injection. AUC values are shown on the right. (D) Effects of NCOA6 on the glucose production level in primary hepatocytes. Glucose production was compared between Ncoa6 LKO and WT mouse primary hepatocytes after a 4 h forskolin treatment using glucose free media supplemented with lactate and sodium pyruvate (n = 4 independent experiments). (E) Alteration of Pck1 and G6pc transcript induction in Ncoa6 LKO primary hepatocytes following a 4 h treatment of these cells with forskolin (n = 5 independent experiments). Transcript levels were analyzed by real-time qRT-PCR. Data are presented as the mean ± SEM; ** P < 0.01, *** P < 0.001, determined using one-way ANOVA. Fsk, forskolin.

Article Snippet: After membrane blocking with 5% skim milk, the membrane was probed with primary antibodies against NCOA6 (Cat. No. NB200-335; Novus Biologicals, USA), PCK1 (Cat. No. sc-32879; Santa Cruz Biotechnology, USA), CREB (Cat. No. 9197; Cell Signaling Technology, USA), p-CREB (Cat. No. 9198S; Cell Signaling Technology), or α-tubulin (Cat. No. T9026; Sigma-Aldrich).

Techniques: Isolation, Quantitative RT-PCR, Expressing, Western Blot, Injection

Journal: Molecules and Cells

Article Title: Regulation of Hepatic Gluconeogenesis by Nuclear Receptor Coactivator 6

doi: 10.14348/molcells.2022.2222

Figure Lengend Snippet: (A) Serum glucagon levels in the Ncoa6 LKO (n = 6 for fed, n = 7 for fasted) and WT mice (n = 7 for both fed and fasted) before and after 16 h of fasting. (B) mRNA levels of gluconeogenic transcriptional factors in the liver of Ncoa6 LKO and WT mice in a fed or 24 h fasted state (n = 6 for each group). Hepatic mRNAs were analyzed by real-time qRT-PCR. (C) Transcript levels of the CREB target gene Nr4a1 and FOXO1 target gene Igfbp1 in the primary hepatocytes of Ncoa6 LKO (n = 4) and WT mice (n = 3). (D) Induced phosphorylation of CREB by 16 h of fasting in the livers of Ncoa6 LKO and WT mice (n = 3 for each group). (E) Forskolin-induced phosphorylation of CREB in primary hepatocytes of Ncoa6 LKO and WT mice. Protein levels were determined by western blot analyses using anti-CREB, anti-pCREB (Ser133) or anti-tubulin antibodies (n = 5 independent experiments). The intensities of the protein bands were measured using the ImageJ program. Data are presented as the mean ± SEM; *** P < 0.001, determined using one-way ANOVA. Fsk, forskolin.

Article Snippet: After membrane blocking with 5% skim milk, the membrane was probed with primary antibodies against NCOA6 (Cat. No. NB200-335; Novus Biologicals, USA), PCK1 (Cat. No. sc-32879; Santa Cruz Biotechnology, USA), CREB (Cat. No. 9197; Cell Signaling Technology, USA), p-CREB (Cat. No. 9198S; Cell Signaling Technology), or α-tubulin (Cat. No. T9026; Sigma-Aldrich).

Techniques: Quantitative RT-PCR, Phospho-proteomics, Western Blot

Journal: Molecules and Cells

Article Title: Regulation of Hepatic Gluconeogenesis by Nuclear Receptor Coactivator 6

doi: 10.14348/molcells.2022.2222

Figure Lengend Snippet: (A) Interactions between CREB and NCOA6. 293T cells were transfected with HA-CREB and FLAG-NCOA6 and lysates were immunoprecipitated from these cells using anti-HA or anti-FLAG antibody. The coimmunoprecipitates were then analyzed by western blotting with anti-HA or anti-FLAG antibody. (B) Sequence alignment of the CRE-containing promoter regions of the Pck1 or G6pc gene among human, mouse, and rat. (C) Schematic diagram of the r Pck1 or h G6PC promoter-Luc reporter containing WT or MT CRE. The red bars indicate the loci of the ChIP primers used in and . (D and E) Effects of NCOA6 on the CREB-mediated transcriptional activation of the r Pck1 (D) and h G6PC (E) promoter (n = 5 independent experiments). WT or MT CRE-containing Pck1 or G6PC promoter-Luc reporters were cotransfected into HepG2 cells with or without HA-CREB, FLAG-NCOA6 and actin-β-galactosidase. Luciferase activities were then measured with a luminometer and normalized using β-galactosidase activities. Data are presented as the mean ± SEM; *** P < 0.001, determined using one-way ANOVA. IP, immunoprecipitation; WT, wild type; MT, mutant; Fsk, forskolin.

Article Snippet: After membrane blocking with 5% skim milk, the membrane was probed with primary antibodies against NCOA6 (Cat. No. NB200-335; Novus Biologicals, USA), PCK1 (Cat. No. sc-32879; Santa Cruz Biotechnology, USA), CREB (Cat. No. 9197; Cell Signaling Technology, USA), p-CREB (Cat. No. 9198S; Cell Signaling Technology), or α-tubulin (Cat. No. T9026; Sigma-Aldrich).

Techniques: Transfection, Immunoprecipitation, Western Blot, Sequencing, Activation Assay, Luciferase, Mutagenesis

Journal: Molecules and Cells

Article Title: Regulation of Hepatic Gluconeogenesis by Nuclear Receptor Coactivator 6

doi: 10.14348/molcells.2022.2222

Figure Lengend Snippet: (A and B) 293T cells were cotransfected with a Pck1 (A) or G6PC (B) promoter-Luc reporter, HA-CREB and FLAG-NCOA6. Modified ChIP assays were then performed using anti-HA, anti-FLAG or IgG antibodies, followed by analysis using conventional PCR (n = 4 independent experiments). The intensities of the PCR bands were measured using the ImageJ program. Data are presented as the mean ± SEM; * P < 0.05, ** P < 0.01, determined by t -test. WT, wild type; MT, mutant; IP, immunoprecipitation. (C) Hypothetical model for the gluconeogenic transcription of the Pck1 and G6pc genes mediated via NCOA6.

Article Snippet: After membrane blocking with 5% skim milk, the membrane was probed with primary antibodies against NCOA6 (Cat. No. NB200-335; Novus Biologicals, USA), PCK1 (Cat. No. sc-32879; Santa Cruz Biotechnology, USA), CREB (Cat. No. 9197; Cell Signaling Technology, USA), p-CREB (Cat. No. 9198S; Cell Signaling Technology), or α-tubulin (Cat. No. T9026; Sigma-Aldrich).

Techniques: Modification, Mutagenesis, Immunoprecipitation

Journal: Clinical and Vaccine Immunology

Article Title: Immune Modulator Adamantylamide Dipeptide Stimulates Efficient Major Histocompatibility Complex Class I-Restricted Responses in Mice

doi: 10.1128/cvi.00316-06

Figure Lengend Snippet: FIG. 1. Kinetic analysis of -Gal-specific serum antibodies in vaccinated animals. BALB/c mice were immunized with PBS, -Gal (50 g) alone, or -Gal coadministered with AdDP (200 g) as the adjuvant. Immunizations are indicated by arrows (days 1, 8, and 15). Antibody titers were determined by endpoint dilution ELISA. Each point represents the group mean titer of total anti--Gal IgM (left panel) and IgG (right panel). The SEMs are indicated by vertical lines. Differences were statistically significant at P 0.05 (*) and P 0.001 (**).

Article Snippet: The immunoglobulin G (IgG) isotypes present in the serum samples were determined by an ELISA, as described previously (2), by using as secondary antibodies biotin-conjugated rat anti-mouse IgG1, IgG2a, IgG2b, or IgG3 (Southern Biotechnology Associates, Birmingham, AL).

Techniques: Adjuvant, Enzyme-linked Immunosorbent Assay

Journal: Clinical and Vaccine Immunology

Article Title: Immune Modulator Adamantylamide Dipeptide Stimulates Efficient Major Histocompatibility Complex Class I-Restricted Responses in Mice

doi: 10.1128/cvi.00316-06

Figure Lengend Snippet: FIG. 2. Analysis of OVA-specific serum antibodies in C57BL/6 mice immunized by the intranasal (i.n.) route with AdDP as the adju- vant. OVA-specific IgG titers from serum samples were determined by endpoint dilution ELISA. Each bar represents the group mean end- point titer. The SEMs are indicated by vertical lines. Differences were statistically significant at P 0.05 (*).

Article Snippet: The immunoglobulin G (IgG) isotypes present in the serum samples were determined by an ELISA, as described previously (2), by using as secondary antibodies biotin-conjugated rat anti-mouse IgG1, IgG2a, IgG2b, or IgG3 (Southern Biotechnology Associates, Birmingham, AL).

Techniques: Enzyme-linked Immunosorbent Assay

Journal: Clinical and Vaccine Immunology

Article Title: Immune Modulator Adamantylamide Dipeptide Stimulates Efficient Major Histocompatibility Complex Class I-Restricted Responses in Mice

doi: 10.1128/cvi.00316-06

Figure Lengend Snippet: FIG. 3. Determination of the number of antibody-secreting cells present in immunized animals. Spleen cells from BALB/c mice vacci- nated with -Gal were cultured for 6 h, and the number of IgG- secreting cells was evaluated by the ELISPOT assay. The results are presented as the total number of IgG spot-forming units (SFU)/106

Article Snippet: The immunoglobulin G (IgG) isotypes present in the serum samples were determined by an ELISA, as described previously (2), by using as secondary antibodies biotin-conjugated rat anti-mouse IgG1, IgG2a, IgG2b, or IgG3 (Southern Biotechnology Associates, Birmingham, AL).

Techniques: Cell Culture, Enzyme-linked Immunospot

Journal: EBioMedicine

Article Title: Adjuvant-enhanced CD4 T Cell Responses are Critical to Durable Vaccine Immunity

doi: 10.1016/j.ebiom.2015.11.041

Figure Lengend Snippet: Adjuvants impact the durability of protection conferred by eVLP vaccination. (a) C57BL/6 mice were vaccinated IM two times with 10 μg of eVLP and challenged four (short-term) weeks or twenty-two (long-term) weeks after the vaccine boost. Data in A are pooled from 8 individual studies with 6–10 animals/group. Fisher's exact test: survival in the short-term group was significantly higher than in the long-term group (p < 0.0001). (b) C57BL/6 mice were vaccinated IM two times with 10 μg of eVLP and challenged at the indicated days after the second vaccination. n = 9 or 10/group. Cochran-Armitage test: percentage surviving declined as time to challenge increased (p = 0.0046). There was a significant difference (p = 0.03) between survival on Day 77 and Day 175. (c) Serum samples collected from animals in (B) one week prior to challenge were subjected to an ELISA for the evaluation of anti-GP IgG, IgG1, and IgG2c antibody titers. Red symbols indicate titers of animals that succumbed to challenge while black indicate titers of survivors; red symbols with black outlines indicate that one of these two animals succumbed to challenge, but animal tags were indeterminate after challenge. Median and IQR shown. (d) C57BL/6 mice were vaccinated two times (IM) with VLP, with or without the indicated adjuvants. Animals were challenged twenty-two weeks after the vaccine boost. Data in D are pooled from at least 4 separate studies with a total of at least 35 animals per group. P-values comparing VLP alone to vaccination with VLP and adjuvant are shown, calculated using Fisher's exact tests with stepdown Bonferroni correction. V = VLP, VP = VLP + PolyICLC, VC = VLP + CpG, VM = VLP + MPLA, and VA = VLP + alhydrogel.

Article Snippet: Secondary antibodies included goat anti-mouse IgG-HRP (Southern Biotech 1030–05), IgG1-HRP (Southern Biotech 1070–05), IgG2c-HRP (Southern Biotech 1079–05), and IgG3-HRP (Southern Biotech 1100–05).

Techniques: Enzyme-linked Immunosorbent Assay, Adjuvant

Journal: EBioMedicine

Article Title: Adjuvant-enhanced CD4 T Cell Responses are Critical to Durable Vaccine Immunity

doi: 10.1016/j.ebiom.2015.11.041

Figure Lengend Snippet: Adjuvants have variable impact on IgG subclasses and antibody neutralization. (a) Serum was collected 14 days and 147 days after the vaccine boost (days 35 and 168, respectively) and evaluated for anti-GP IgG, IgG1, IgG2c, and IgG3 levels using an ELISA. Data shown are pooled from at least two separate experiments per group. (b) Pairwise comparison using DSCF multiple pairwise comparison was used and p values greater than 0.05 are shown as “ns”. (c) Summary of results shown in A, B and D. (d) Neutralizing antibody titers were evaluated using the PsVNA, with titers giving 80% neutralization shown; median and IQR shown. Samples within each group were selected randomly from three separate studies for evaluation in the assay. Pairwise comparison using post-hoc Tukey's studentized range test procedure was used to evaluate differences between groups at both days 35 and day 168, where “*” indicates 0.01 < p < 0.05, “**” indicates 0.001 < p < 0.01, “***” indicates 0.0001 < p < 0.001, and “****” indicates p < 0.0001.

Article Snippet: Secondary antibodies included goat anti-mouse IgG-HRP (Southern Biotech 1030–05), IgG1-HRP (Southern Biotech 1070–05), IgG2c-HRP (Southern Biotech 1079–05), and IgG3-HRP (Southern Biotech 1100–05).

Techniques: Neutralization, Enzyme-linked Immunosorbent Assay, Comparison

Journal: EBioMedicine

Article Title: Adjuvant-enhanced CD4 T Cell Responses are Critical to Durable Vaccine Immunity

doi: 10.1016/j.ebiom.2015.11.041

Figure Lengend Snippet: CD8 T cell deficiency does not impact short term or long term survival of vaccinated C57BL/6J mice. (a) Mice were treated IM twice with saline, VLP, VLP and polyICLC, or VLP and CpG. Four weeks after the second vaccination, mice were challenged. Closed symbols represent wild type C57BL/6J mice and open symbols indicate CD8-deficient mice. (b) Mice were vaccinated on the same schedule as in A, but challenge occurred 22 weeks after the second vaccination. (c) Two weeks after the second vaccination and 1 week prior to challenge, blood was collected from vaccinated animals and evaluated for anti-GP IgG antibody titers. (D) A subset of vaccinated animals was euthanized 4 days after the vaccine boost to evaluate CD4 + T cell responses. Median response of C57BL/6J mice and CD8-deficient mice is shown. N = 8–10 per treated group for A–C and D presents data pooled from two separate evaluations of 4–8 mice per group each. “*” indicates 0.005 < p < 0.05.

Article Snippet: Secondary antibodies included goat anti-mouse IgG-HRP (Southern Biotech 1030–05), IgG1-HRP (Southern Biotech 1070–05), IgG2c-HRP (Southern Biotech 1079–05), and IgG3-HRP (Southern Biotech 1100–05).

Techniques: Saline