|
| Status |
Public on Jul 07, 2017 |
| Title |
subject_55-Terminal Ileum-biopsy_29 |
| Sample type |
RNA |
| |
|
| Source name |
Terminal Ileum
|
| Organism |
Homo sapiens |
| Characteristics |
diagnosis: inflammatory bowel disease
tissue: Terminal Ileum
age (yr): 36
Sex: F
inflammation: Non-involved area
|
| Treatment protocol |
All blood samples were stored at −80 °C until RNA isolation was performed. Blood samples were hybridized by microarray for genome-wide mRNA expression profiling. Ileal, rectal, and colonic biopsies were collected at the screening visits from a sub-group of subjects who consented to this procedure. Biopsies were obtained from selected colonic sections, including the ascending, transverse, and descending colonic regions, and from both inflamed and non-involved areas. The inflamed regions were defined as the colonic segments with the most severe disease activity. Additional biopsies were collected from the terminal ileum and from the rectum (i.e., the distal 10 cm of the colon).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA including micro ribonucleic acids (miRNA) was isolated with PAXgene Blood RNA MDx Kit plus customized reagent BM3 (Cat#762431, lot 136237127) according to the manufacturer’s instructions (Qiagen Inc., Valencia, CA). Briefly, PAXgene Blood RNA Tubes were incubated at room temperature around 2 hrs before extraction. After centrifugation for 10 min at 3000–5000 x g, the pellet was re-suspended in 290 μl Buffer BR1 with 35 μl proteinase K. The remaining procedures were performed on a BioRobot Universal System using a customized protocol, PAXgene Blood RNA UNIV rcV73_miRNA Centocor.pro (Qiagen).
|
| Label |
NuGEN Encore Biotin Module
|
| Label protocol |
RNA samples were amplified by NuGEN Ovation RNA Amplification System V2 Whole Blood Solution (NuGEN, San Carlos, CA) and purified using Agencourt RNAClean magnetic beads. Labeling was done using the NuGEN Encore Biotin Module (NuGEN). (Agencourt, Beverly, MA) on a Caliper SciClone robot.
|
| |
|
| Hybridization protocol |
Gene expression were carried out using Affymetrix GeneChip HTHGU133+ (Affymetrix, Santa Clara, CA, USA Cat# 901262) according to the manufacturer’s protocol with the exception that dimethyl sulphoxide replaced the Tetramethylammonium Chloride Solution in the hybridization buffer.
|
| Scan protocol |
Arrays were washed and stained on the Affymetrix GeneChip Array Station then scanned on an HTAPS scanner.
|
| Data processing |
Array Studio software version 6.0 (OmicSoft Corp., St. Morrisville, NC) was used for data analysis. The microarray data were pre-processed and normalized using RMA. Log2 transformed data was used for statistical analysis.
|
| |
|
| Submission date |
Jul 05, 2017 |
| Last update date |
Jul 09, 2018 |
| Contact name |
Eric Schadt |
| E-mail(s) |
eric.schadt@mssm.edu
|
| Organization name |
Icahn School of Medicine at Mount Sinai
|
| Street address |
1425 Madison Avenue
|
| City |
New York |
| ZIP/Postal code |
10029 |
| Country |
USA |
| |
|
| Platform ID |
GPL13158 |
| Series (1) |
| GSE100833 |
A functional genomics predictive network model identifies regulators of inflammatory bowel disease: Microarray Analysis of Human Blood and Intestinal Biopsy Samples from a Phase 2b, Double-blind, Placebo-controlled Study of Ustekinumab in Crohn's Disease |
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