 |
 |
GEO help: Mouse over screen elements for information. |
|
| Status |
Public on Sep 08, 2018 |
| Title |
Extraction-Free Whole Transcriptome Profiling from Specific Subareas of Unstained or H&E Stained FFPE Tissues |
| Organisms |
Homo sapiens; Mus musculus; Rattus norvegicus |
| Experiment type |
Expression profiling by high throughput sequencing
|
| Summary |
We describe the use of a ligation-based targeted whole transcriptome expression profiling assay, TempO-Seq™, to profile formalin-fixed paraffin-embedded (FFPE) tissue, including H&E stained FFPE tissue, by directly lysing tissue scraped from slides without extracting RNA or converting the RNA to cDNA. The correlation of measured gene expression changes in unfixed and fixed samples using blocks prepared from a pellet of a single cell type was R2 = 0.97, demonstrating that no significant artifacts were introduced by fixation. Fixed and fresh samples prepared in an equivalent manner produced comparable sequencing depth results (+/- 20%), with similar %CV (11.5 and 12.7%, respectively), indicating no significant loss of measurable RNA due to fixation. The sensitivity of the TempO-Seq assay was the same whether the tissue section was fixed or not. The assay performance was equivalent for human, mouse, or rat whole transcriptome. The results from 10 mm2 and 2 mm2 areas of tissue obtained from 5 μm thick sections were equivalent, thus demonstrating high sensitivity and ability to profile focal areas of histology within a section. Replicate reproducibility of separate areas of tissue ranged from R2= 0.83 (lung) to 0.96 (liver) depending on the tissue type, with an average correlation of R2 = 0.90 across nine tissue types. The average %CV’s were 16.8% for genes expressed at greater than 200 counts, and 20.3% for genes greater than 50 counts. Tissue specific differences in gene expression were identified and agreed with the literature. There was negligible impact on assay performance using FFPE tissues that had been archived for up to 30 years. Similarly, there was negligible impact of H&E staining, facilitating accurate visualization for scraping and assay of small focal areas of specific histology within a section.
|
| |
|
| Overall design |
FFPE tissues and cell pellets from human, mouse, and rat were used to assess performance of a targeted whole transcriptome expression profiling method. A subset of human FFPE tissue was stained with hemotoxylin and eosin. Another subset of human FFPE tissue was pulled from archives and were greater than 24 years old at the time of processing. A subset of rat tissue was fixed for variable amounts of time before embedding. Various amounts of FFPE tissue was used as input into the assay. For all FFPE samples, FFPE tissue was used as direct input into the assay without RNA extraction. Twelve samples are FFPE formed by fixing, embedding, and sectioning pellets from cultured MCF7 and MDA-MB-231 cells. Six samples (marked "fresh") are lysates of unfixed cells processed through the same assay for comparison.
|
| |
|
| Contributor(s) |
Trejo CL, Babić M |
| Citation(s) |
30794557 |
| |
| Submission date |
Sep 07, 2018 |
| Last update date |
Mar 26, 2019 |
| Contact name |
Milos Babic |
| E-mail(s) |
milosbabic@biospyder.com
|
| Phone |
5204406930
|
| Organization name |
BioSpyder Technologies Inc.
|
| Street address |
5922 Farnsworth Court, Suite 102
|
| City |
Carlsbad |
| State/province |
CA |
| ZIP/Postal code |
92008 |
| Country |
USA |
| |
|
| Platforms (3) |
| GPL18573 |
Illumina NextSeq 500 (Homo sapiens) |
| GPL19057 |
Illumina NextSeq 500 (Mus musculus) |
| GPL20084 |
Illumina NextSeq 500 (Rattus norvegicus) |
|
| Samples (155)
|
| GSM3379604 |
Fig 2 colon replicate 1 |
| GSM3379605 |
Fig 2 colon replicate 2 |
| GSM3379606 |
Fig 2 colon replicate 3 |
| GSM3379607 |
Fig 2 prostate replicate 1 |
| GSM3379608 |
Fig 2 prostate replicate 2 |
| GSM3379609 |
Fig 2 prostate replicate 3 |
| GSM3379610 |
Fig 2 breast replicate 1 |
| GSM3379611 |
Fig 2 breast replicate 2 |
| GSM3379612 |
Fig 2 breast replicate 3 |
| GSM3379613 |
Fig 2 lung replicate 1 |
| GSM3379614 |
Fig 2 lung replicate 2 |
| GSM3379615 |
Fig 2 lung replicate 3 |
| GSM3379616 |
Fig 2 muscle replicate 1 |
| GSM3379617 |
Fig 2 muscle replicate 2 |
| GSM3379618 |
Fig 2 muscle replicate 3 |
| GSM3379619 |
Fig 2 brain replicate 1 |
| GSM3379620 |
Fig 2 brain replicate 2 |
| GSM3379621 |
Fig 2 brain replicate 3 |
| GSM3379622 |
Fig 2 kidney replicate 1 |
| GSM3379623 |
Fig 2 kidney replicate 2 |
| GSM3379624 |
Fig 2 kidney replicate 3 |
| GSM3379625 |
Fig 2 liver replicate 1 |
| GSM3379626 |
Fig 2 liver replicate 2 |
| GSM3379627 |
Fig 2 liver replicate 3 |
| GSM3379628 |
Fig 7 deparaffinized 1 |
| GSM3379629 |
Fig 7 deparaffinized 2 |
| GSM3379630 |
Fig 7 paraffinized 1 |
| GSM3379631 |
Fig 7 paraffinized 2 |
| GSM3379632 |
Fig 7 paraffinized 3 |
| GSM3379633 |
Fig 7 paraffinized 4 |
| GSM3379634 |
Fig 7 stained 1 |
| GSM3379635 |
Fig 7 stained 2 |
| GSM3379636 |
Fig 5 - 384 hrs -1 |
| GSM3379637 |
Fig 5 - 384 hrs -2 |
| GSM3379638 |
Fig 5 - 384 hrs -3 |
| GSM3379639 |
Fig 5 - 24 hrs -1 |
| GSM3379640 |
Fig 5 - 24 hrs -2 |
| GSM3379641 |
Fig 5 - 24 hrs -3 |
| GSM3379642 |
Fig 5 - 96 hrs -1 |
| GSM3379643 |
Fig 5 - 96 hrs -2 |
| GSM3379644 |
Fig 5 - 96 hrs -3 |
| GSM3379645 |
Fig 5 - 192 hrs -1 |
| GSM3379646 |
Fig 5 - 192 hrs -2 |
| GSM3379647 |
Fig 5 - 192 hrs -3 |
| GSM3379648 |
Fig 4 colon 1-86-1 |
| GSM3379649 |
Fig 4 colon 1-86-2 |
| GSM3379650 |
Fig 4 colon 1-86-3 |
| GSM3379651 |
Fig 4 colon 2-86-1 |
| GSM3379652 |
Fig 4 colon 2-86-2 |
| GSM3379653 |
Fig 4 colon 2-86-3 |
| GSM3379654 |
Fig 4 kidney 1-94-1 |
| GSM3379655 |
Fig 4 kidney 1-94-2 |
| GSM3379656 |
Fig 4 kidney 1-94-3 |
| GSM3379657 |
Fig 4 kidney 2-94-1 |
| GSM3379658 |
Fig 4 kidney 2-94-2 |
| GSM3379659 |
Fig 4 kidney 2-94-3 |
| GSM3379660 |
Fig 4 kidney 1-88-1 |
| GSM3379661 |
Fig 4 kidney 1-88-2 |
| GSM3379662 |
Fig 4 kidney 1-88-3 |
| GSM3379663 |
Fig 4 kidney 2-88-1 |
| GSM3379664 |
Fig 4 kidney 2-88-2 |
| GSM3379665 |
Fig 4 kidney 2-88-3 |
| GSM3379666 |
Fig 4 liver 1-93-1 |
| GSM3379667 |
Fig 4 liver 1-93-2 |
| GSM3379668 |
Fig 4 liver 1-93-3 |
| GSM3379669 |
Fig 4 liver 2-93-1 |
| GSM3379670 |
Fig 4 liver 2-93-2 |
| GSM3379671 |
Fig 4 liver 2-93-3 |
| GSM3379672 |
Fig 3 liver 10-1 |
| GSM3379673 |
Fig 3 liver 10-2 |
| GSM3379674 |
Fig 3 liver 10-3 |
| GSM3379675 |
Fig 3 liver 2-1 |
| GSM3379676 |
Fig 3 liver 2-2 |
| GSM3379677 |
Fig 3 liver 2-3 |
| GSM3379678 |
Fig 6a ffpe mcf-1 |
| GSM3379679 |
Fig 6a ffpe mcf-2 |
| GSM3379680 |
Fig 6a ffpe mcf-3 |
| GSM3379681 |
Fig 6a ffpe mda-1 |
| GSM3379682 |
Fig 6a ffpe mda-2 |
| GSM3379683 |
Fig 6a ffpe mda-3 |
| GSM3379684 |
Fig 6a fresh mcf-1 |
| GSM3379685 |
Fig 6a fresh mcf-2 |
| GSM3379686 |
Fig 6a fresh mcf-3 |
| GSM3379687 |
Fig 6a fresh mda-1 |
| GSM3379688 |
Fig 6a fresh mda-2 |
| GSM3379689 |
Fig 6a fresh mda-3 |
| GSM3379690 |
Fig 6b mcf20-1 |
| GSM3379691 |
Fig 6b mcf20-2 |
| GSM3379692 |
Fig 6b mcf20-3 |
| GSM3379693 |
Fig 6b mda20-1 |
| GSM3379694 |
Fig 6b mda20-2 |
| GSM3379695 |
Fig 6b mda20-3 |
| GSM3477439 |
Patient 1, normal colon tissue, biological replicate 1, technical replicate 1 |
| GSM3477440 |
Patient 1, normal colon tissue, biological replicate 1, technical replicate 2 |
| GSM3477441 |
Patient 1, normal colon tissue, biological replicate 1, technical replicate 3 |
| GSM3477442 |
Patient 1, cancerous colon tissue, biological replicate 1, technical replicate 1 |
| GSM3477443 |
Patient 1, cancerous colon tissue, biological replicate 1, technical replicate 2 |
| GSM3477444 |
Patient 1, cancerous colon tissue, biological replicate 1, technical replicate 3 |
| GSM3477445 |
Patient 1, normal colon tissue, biological replicate 2, technical replicate 1 |
| GSM3477446 |
Patient 1, normal colon tissue, biological replicate 2, technical replicate 2 |
| GSM3477447 |
Patient 1, normal colon tissue, biological replicate 2, technical replicate 3 |
| GSM3477448 |
Patient 1, cancerous colon tissue, biological replicate 2, technical replicate 1 |
| GSM3477449 |
Patient 1, cancerous colon tissue, biological replicate 2, technical replicate 2 |
| GSM3477450 |
Patient 1, cancerous colon tissue, biological replicate 2, technical replicate 3 |
| GSM3477451 |
Patient 2, normal colon tissue, biological replicate 1, technical replicate 1 |
| GSM3477452 |
Patient 2, normal colon tissue, biological replicate 1, technical replicate 2 |
| GSM3477453 |
Patient 2, normal colon tissue, biological replicate 1, technical replicate 3 |
| GSM3477454 |
Patient 2, cancerous colon tissue, biological replicate 1, technical replicate 1 |
| GSM3477455 |
Patient 2, cancerous colon tissue, biological replicate 1, technical replicate 2 |
| GSM3477456 |
Patient 2, cancerous colon tissue, biological replicate 1, technical replicate 3 |
| GSM3477457 |
Patient 2, normal colon tissue, biological replicate 2, technical replicate 1 |
| GSM3477458 |
Patient 2, normal colon tissue, biological replicate 2, technical replicate 2 |
| GSM3477459 |
Patient 2, normal colon tissue, biological replicate 2, technical replicate 3 |
| GSM3477460 |
Patient 2, cancerous colon tissue, biological replicate 2, technical replicate 1 |
| GSM3477461 |
Patient 2, cancerous colon tissue, biological replicate 2, technical replicate 2 |
| GSM3477462 |
Patient 2, cancerous colon tissue, biological replicate 2, technical replicate 3 |
| GSM3477463 |
Patient 3, normal colon tissue, biological replicate 1, technical replicate 1 |
| GSM3477464 |
Patient 3, normal colon tissue, biological replicate 1, technical replicate 2 |
| GSM3477465 |
Patient 3, normal colon tissue, biological replicate 1, technical replicate 3 |
| GSM3477466 |
Patient 3, cancerous colon tissue, biological replicate 1, technical replicate 1 |
| GSM3477467 |
Patient 3, cancerous colon tissue, biological replicate 1, technical replicate 2 |
| GSM3477468 |
Patient 3, cancerous colon tissue, biological replicate 1, technical replicate 3 |
| GSM3477469 |
Patient 3, normal colon tissue, biological replicate 2, technical replicate 1 |
| GSM3477470 |
Patient 3, normal colon tissue, biological replicate 2, technical replicate 2 |
| GSM3477471 |
Patient 3, normal colon tissue, biological replicate 2, technical replicate 3 |
| GSM3477472 |
Patient 3, cancerous colon tissue, biological replicate 2, technical replicate 1 |
| GSM3477473 |
Patient 3, cancerous colon tissue, biological replicate 2, technical replicate 2 |
| GSM3477474 |
Patient 3, cancerous colon tissue, biological replicate 2, technical replicate 3 |
| GSM3477475 |
Patient 4, normal colon tissue, biological replicate 1, technical replicate 1 |
| GSM3477476 |
Patient 4, normal colon tissue, biological replicate 1, technical replicate 2 |
| GSM3477477 |
Patient 4, normal colon tissue, biological replicate 1, technical replicate 3 |
| GSM3477478 |
Patient 4, cancerous colon tissue, biological replicate 1, technical replicate 1 |
| GSM3477479 |
Patient 4, cancerous colon tissue, biological replicate 1, technical replicate 2 |
| GSM3477480 |
Patient 4, cancerous colon tissue, biological replicate 1, technical replicate 3 |
| GSM3477481 |
Patient 4, normal colon tissue, biological replicate 2, technical replicate 1 |
| GSM3477482 |
Patient 4, normal colon tissue, biological replicate 2, technical replicate 2 |
| GSM3477483 |
Patient 4, normal colon tissue, biological replicate 2, technical replicate 3 |
| GSM3477484 |
Patient 4, cancerous colon tissue, biological replicate 2, technical replicate 1 |
| GSM3477485 |
Patient 4, cancerous colon tissue, biological replicate 2, technical replicate 2 |
| GSM3477486 |
Patient 4, cancerous colon tissue, biological replicate 2, technical replicate 3 |
| GSM3477487 |
Patient 5, normal colon tissue, biological replicate 1, technical replicate 1 |
| GSM3477488 |
Patient 5, normal colon tissue, biological replicate 1, technical replicate 2 |
| GSM3477489 |
Patient 5, normal colon tissue, biological replicate 1, technical replicate 3 |
| GSM3477490 |
Patient 5, cancerous colon tissue, biological replicate 1, technical replicate 1 |
| GSM3477491 |
Patient 5, cancerous colon tissue, biological replicate 1, technical replicate 2 |
| GSM3477492 |
Patient 5, cancerous colon tissue, biological replicate 1, technical replicate 3 |
| GSM3477493 |
Patient 5, normal colon tissue, biological replicate 2, technical replicate 1 |
| GSM3477494 |
Patient 5, normal colon tissue, biological replicate 2, technical replicate 2 |
| GSM3477495 |
Patient 5, normal colon tissue, biological replicate 2, technical replicate 3 |
| GSM3477496 |
Patient 5, cancerous colon tissue, biological replicate 2, technical replicate 1 |
| GSM3477497 |
Patient 5, cancerous colon tissue, biological replicate 2, technical replicate 2 |
| GSM3477498 |
Patient 5, cancerous colon tissue, biological replicate 2, technical replicate 3 |
|
| Relations |
| BioProject |
PRJNA489794 |
| SRA |
SRP160108 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE119630_ColonCancerReplicatesMaster.csv.gz |
1.9 Mb |
(ftp)(http) |
CSV |
| GSE119630_HumanGeneCountsMaster.csv.gz |
1.6 Mb |
(ftp)(http) |
CSV |
| GSE119630_MouseGeneCountsMaster.csv.gz |
1.2 Mb |
(ftp)(http) |
CSV |
| GSE119630_RatGeneCountsMaster.csv.gz |
1.0 Mb |
(ftp)(http) |
CSV |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
|
|
|
|
 |
Less...
More...