Journal: Cellular and Molecular Gastroenterology and Hepatology

Article Title: Interferon-λ3 Promotes Epithelial Defense and Barrier Function Against Cryptosporidium parvum Infection

doi: 10.1016/j.jcmgh.2019.02.007

Figure Lengend Snippet: IECs up-regulate numerous ISGs in response to C parvum infection. Intestinal epithelium was harvested from ileum mucosa of neonatal piglets at the time of peak infection (days 3–5 after infection) (n = 8) and from age-matched uninfected controls (n = 4) for performance of gene expression analysis using microarrays. ( A ) Representative photomicrograph of ileum mucosa from an uninfected control and C parvum –infected piglet used for microarray analysis. H&E stain. Scale bar : 20 μm. ( B ) Villus height and crypt depth (μm) and the percentage of total villus IECs that were infected with C parvum in control (n = 4) and C parvum –infected (n = 8) piglets used for microarray analysis. Each data point represents the average of 5 measurements per piglet. Scale bars : means ± SD. ** P < .01, Student t test comparison between uninfected and C parvum –infected piglets. Heat map of significantly ( C ) up-regulated and ( D ) down-regulated genes in infected ( C parvum ) and control (Uninf) exfoliated ileum villus epithelial cells. Each control and infected biological replicate is represented. Gene IDs:fold change are listed to the right of the heat map. Known ISGs are highlighted in red. ( E ) qRT-PCR analysis of porcine ileum mucosal total cellular mRNA for the presence of ISG15 mRNA. Samples were obtained from piglets used for microarray analysis at peak C parvum infection (days 3–5, n = 5) and age-matched controls (n = 5). For each sample, the Ct value for ISG15 was normalized to expression of the housekeeping gene cyclophilin (ΔCt). Fold-change differences between each sample were compared with a representative uninfected sample using the 2 -ΔΔCt method. Scale bars : means ± SD. ** P < .01, Student t test comparison between uninfected and C parvum –infected piglets. ( F ) Fluorescence in situ hybridization showing ISG15 mRNA (red fluorescence) in villus epithelial cells of ileum mucosa from piglets at peak C parvum infection and absent in villus epithelium of control piglets. Photomicrograph representative of results of 2 independent experiments. Scale bar : 50 μm.

Article Snippet: The fragmented complementary RNA was diluted in hybridization buffer (2 N -morpholino-ethanesulfonic acid, NaCl, EDTA, Tween 20, herring sperm DNA, acetylated bovine serum albumin) containing biotin-labeled OligoB2 and Eukaryotic Hybridization Controls (Affymetrix).

Techniques: Infection, Gene Expression, Control, Microarray, Staining, Comparison, Quantitative RT-PCR, Expressing, Fluorescence, In Situ Hybridization

Journal: Oncogene

Article Title: Prediction of future metastasis and molecular characterization of head and neck squamous-cell carcinoma based on transcriptome and genome analysis by microarrays.

doi: 10.1038/onc.2008.251

Figure Lengend Snippet: Figure 2 RNA expression by seven genes relative to pathological differentiation. The box plots indicate the distribution of RNA levels measured by QRT–PCR in 134 tumour samples grouped by level of differentiation. The seven genes yielded significant P-values (Po0.01) based on Affymetrix data testing the different tumour differentiation classes. Colours correspond to the sample groups (green: well differentiated (n ¼ 40); orange: moderately differentiated (n ¼ 68); red: poorly differentiated (n ¼ 29)). The y axis gives the DCt values relative to the control genes (R18S and RPLP0). The bottom table shows the fold difference (FC) between the geometric mean values between the different classes and associated Wilcoxon P-value.

Article Snippet: Prediction analysis Initial selection was based on univariate and multivariate Cox analyses (survival R package v2.26) of Affymetrix gene chip variables using 81 samples, divided into three groups: training group S1 (20M and 20 NM samples), training group S2 (10M and 10 NM samples) and a validation group S3 (11M and 10 NM samples).

Techniques: RNA Expression, Quantitative RT-PCR, Control

Journal: Oncogene

Article Title: Prediction of future metastasis and molecular characterization of head and neck squamous-cell carcinoma based on transcriptome and genome analysis by microarrays.

doi: 10.1038/onc.2008.251

Figure Lengend Snippet: Figure 4 QRT–PCR validation of genes associated with metastasis. Transcripts (22) selected from the transcriptome array analysis (Affymetrix; Cox P-value o0.05) were quantitated by QRT–PCR analysis with 134 samples of M (n ¼ 46) and NM (n ¼ 88). Top: box plots (NM blue, M red) representing the distributions of the log2 value of the DCt values after power transformation. Bottom: the fold difference (FC) between the geometric mean values from M divided by NM samples and the p value calculated from the Cox univariate tests for each of the genes, using the entire population of 134 samples analysed by QRT–PCR.

Article Snippet: Prediction analysis Initial selection was based on univariate and multivariate Cox analyses (survival R package v2.26) of Affymetrix gene chip variables using 81 samples, divided into three groups: training group S1 (20M and 20 NM samples), training group S2 (10M and 10 NM samples) and a validation group S3 (11M and 10 NM samples).

Techniques: Quantitative RT-PCR, Biomarker Discovery, Transformation Assay

Journal: Emerging Infectious Diseases

Article Title: Identifying Influenza Viruses with Resequencing Microarrays

doi: 10.3201/eid1204.051441

Figure Lengend Snippet: Hybridization images of the respiratory pathogen microarray (RPM) version 1 prototype regions for 3 influenza virus isolates and trivalent FluMist vaccine. A) A/H1N1, B) A/H3N2, C) influenza B, and D) trivalent FluMist vaccine. In A, B, and C, only the influenza-specific tiled prototype regions of RPM version 1 are shown. Hybridization-positive identifications are shown on the right. In D, the image of the entire RPM version when hybridized with FluMist vaccine is shown. The single influenza prototype region that was hybridization negative is denoted on the right. E) Magnification of a portion of profile B showing an example of the primary sequence data generated by the hybridization of randomly amplified targets to the RPM version 1 HA3 probe set. The primary sequence generated can be read from left to right. HA, hemagglutinin; NA, neuraminidase; IQEX, internal positive hybridization control (Affymetrix); M, matrix.

Article Snippet: After the prehybridization step, 167.5 μL of hybridization cocktail master mix (3 mol/L tetramethylammonium chloride, 10 mmol/L Tris, pH 7.8, 0.01% Tween 20, 0.5 mg/mL bovine serum albumin, 0.1 mg/mL herring sperm DNA [Promega, Madison, WI, USA], 50 pmol/L Oligo B2 [Affymetrix Inc.]) and biotin-labeled DNA fragments were heated for 5 min at 95°C, equilibrated for 5 min at 45°C, and added to RPM version 1.

Techniques: Hybridization, Microarray, Virus, Sequencing, Generated, Amplification, Control