|
| Status |
Public on Dec 31, 2014 |
| Title |
Recurrence_patient-62_SCC_str_bio-rep2 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
Recurrence case, patient-62, SCC, stromum
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: squamous cell carcinoma stroma
patient: 62
recurrence: Y
months_to_recurrence: 40
histology: SCC
age_at_surgery: 63
gender: F
race: White
smoking_history: former
pathologic_stage: 1A
months_to_death_or_censorship: 84
tumor_component: S
|
| Treatment protocol |
Laser capture microdissection was performed to isolate epithelial or stromal compartments of tumors using hematoxylin-eosin-stained tissue sections. Refer to Patnaik et al., BMC Research Notes, 2012, 5:40 (PMID 22260539) for details on tissue section preparation and microdissection.
|
| Growth protocol |
Formalin-fixed and paraffin-embedded, resected, stage I, human lung cancer tumor tissue samples were provided by the Department of Pathology of Roswell Park Cancer Institute, Buffalo, NY, USA.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted from microdissectates using the FFPE RNA Purification kit (catalog# 25300, Norgen-Biotek®, Thorold, ON, Canada) following the kit manufacturer's guidelines. The kit uses silicon carbide spin-columns. The extracted RNA preparations were not treated with DNase. The low-range Quant-iT™ Ribogreen assay (Life Technologies®, Carlsbad, CA) was used to estimate RNA concentration using yeast tRNA as the standard. Refer to Patnaik et al., BMC Research Notes, 2012, 5:40 (PMID 22260539) for details on RNA isolation and quantitation. The reference RNA, a human 'universal reference total RNA,' was obtained by Exiqon® from Ambion® (catalog# AM6000; Austin, TX).
|
| Label |
Hy3
|
| Label protocol |
The miRCURY™ LNA microRNA Hi-Power Labeling kit from Exiqon® was used to label 350 ng of test RNA with the Cy3-like Hy3™ dye. The reference RNA (350 ng) was similarly labeled with the Cy5-like Hy5™ dye. Artificial small RNAs were spiked in to test and reference RNA samples before labeling. This work was performed as a commercial service by Exiqon®.
|
| |
|
| Channel 2 |
| Source name |
Reference RNA
|
| Organism |
Homo sapiens |
| Characteristics |
sample type: reference
|
| Treatment protocol |
Laser capture microdissection was performed to isolate epithelial or stromal compartments of tumors using hematoxylin-eosin-stained tissue sections. Refer to Patnaik et al., BMC Research Notes, 2012, 5:40 (PMID 22260539) for details on tissue section preparation and microdissection.
|
| Growth protocol |
Formalin-fixed and paraffin-embedded, resected, stage I, human lung cancer tumor tissue samples were provided by the Department of Pathology of Roswell Park Cancer Institute, Buffalo, NY, USA.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted from microdissectates using the FFPE RNA Purification kit (catalog# 25300, Norgen-Biotek®, Thorold, ON, Canada) following the kit manufacturer's guidelines. The kit uses silicon carbide spin-columns. The extracted RNA preparations were not treated with DNase. The low-range Quant-iT™ Ribogreen assay (Life Technologies®, Carlsbad, CA) was used to estimate RNA concentration using yeast tRNA as the standard. Refer to Patnaik et al., BMC Research Notes, 2012, 5:40 (PMID 22260539) for details on RNA isolation and quantitation. The reference RNA, a human 'universal reference total RNA,' was obtained by Exiqon® from Ambion® (catalog# AM6000; Austin, TX).
|
| Label |
Hy5
|
| Label protocol |
The miRCURY™ LNA microRNA Hi-Power Labeling kit from Exiqon® was used to label 350 ng of test RNA with the Cy3-like Hy3™ dye. The reference RNA (350 ng) was similarly labeled with the Cy5-like Hy5™ dye. Artificial small RNAs were spiked in to test and reference RNA samples before labeling. This work was performed as a commercial service by Exiqon®.
|
| |
|
| |
| Hybridization protocol |
Hybridizations and washes were performed according to the Exiqon® miRCURY™ LNA microRNA Array Instruction manual using an HS4800 hybridization station (Tecan®, Grödig, Austria). This work was performed as a commercial service by Exiqon®.
|
| Scan protocol |
Arrays were scanned using the G2565BA Microarray Scanner System (Agilent®), and image analysis was carried out using the ImaGene™ software (version 9; BioDiscovery®, Hawthorne, CA). This work was performed as a commercial service by Exiqon®.
|
| Description |
Sample 144.
Biological replicate for sample 128.
|
| Data processing |
Raw Hy3™ and Hy5™ signal data for all 136 arrays were processed together in R (version 2.15.1) using the limma Bioconductor package (version 3.12.3) on Mac OS X 10.6.8. Briefly, background signals were adjusted for using the 'normexp' method with 'offset' value of 10. Array data was then subjected to within-array global loess normalization with a 'span' value of 1/3. This was followed by between-array 'quantile' normalization. Probe-set summarization for Hy3™/Hy5™ ratio values (test RNA/reference RNA) was then performed, using the mean value from replicate probe-spots when the maximum was <1.5x minimum value, using the median value otherwise.
Raw data files:
1_Exiqon*: Hy3
0_Exiqon*: Hy5
|
| |
|
| Submission date |
Nov 20, 2012 |
| Last update date |
Dec 31, 2014 |
| Contact name |
Santosh Kumar Patnaik |
| Phone |
716-8458364
|
| Organization name |
Roswell Park Comprehensive Cancer Center
|
| Department |
Thoracic Surgery
|
| Street address |
Elm and Carlton Streets
|
| City |
Buffalo |
| State/province |
NY |
| ZIP/Postal code |
14263 |
| Country |
USA |
| |
|
| Platform ID |
GPL16294 |
| Series (1) |
| GSE42425 |
MicroRNA expression in epithelial and stromal compartments of stage I human non-small cell lung cancer tumors |
|
