dataset: Validation age: 83.1 Sex: female therapy: 5-FU + RT surgery type: Abdominoperineal excision (APE) clinical tumor category (0,i,ii,iii,iv - according to uicc tnm classification): 2 clinical lymphnode status (0,1 - according to uicc tnm classification): 1 clinical tumor stage (0,i,ii,iii,iv - according to uicc tnm classification): III A pathological tumor category after neoadjuvant treatment and surgery (0,i,ii,iii,iv - according to uicc tnm classification): 1 pathological lymphnode status after neoadjuvant treatment and surgery (0,1,2 - according to uicc tnm classification): 1 pathological tumor stage after neoadjuvant treatment and surgery (0,i,ii,iii, iv - according to uicc tnm classification): III A tumor regression grading (trg) after neoadjuvant treatment and surgery (0=no regression - iv=maximal regression): 3 disease free survival (dfs) in months from surgery date: NA local recurrence or distance metastasis event (1) or no event (0): NA cancer specific survival (css) in months from surgery date: NA tumor related death (1) or data censoring without event (0): NA rin: 8.7
Treatment protocol
No treatment prior to expression profiling analysis was performed.
Extracted molecule
total RNA
Extraction protocol
Biospies from patients not cell lines were used for analyses. Biopsies were immediately stored in RNAlater (Qiagen, Hilden, Germany). Subsequently, RNA and DNA was isolated using TRIzol (Invitrogen, Carlsbad, CA) according to the manufacturer´s instructions. Nucleic acid quantity, quality and purity were determined using a spectrophotometer (Nanodrop, Rockland, DE) and a 2100 Bioanalyzer (Agilent Technologies, Palo Alto, CA).
Label
Cy3
Label protocol
1 µg of total RNA was labeled with Cy3 using the Low RNA Input Fluorescent Linear Amplification Kit according to the manufacturer's recommendations (Agilent Technologies, Santa Clara, CA). Quantity and efficiency of the labeled amplified cRNA were determined using the NanoDrop ND-1000 UV-VIS Spectrophotometer version 3.2.1.
Hybridization protocol
1.5 mg of Cy3-labeled cRNA was hybridized to an oligonucleotide-based Whole Human Genome Microarray (4x44K, Agilent Technologies) and incubated at 65˚C for 17 hours.
Scan protocol
Slides were washed and scanned using an Agilent G2565BA scanner.
Description
Human pretherapeutic rectal cancer pat244
Data processing
Raw data were extracted using the Feature Extraction software version 9.1 (Agilent Technologies). Median signal intensities from the Feature Extraction Software were converted to log2 and quantile normalized using the R package “limma”.
