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Enabling RNA Analyses from Formaldehyde-fixed Tissues

Award Information

Agency:
Department of Health and Human Services
Branch:
N/A
Award ID:
71759
Program Year/Program:
2004 / SBIR
Agency Tracking Number:
NS046958
Solicitation Year:
N/A
Solicitation Topic Code:
N/A
Solicitation Number:
N/A
Small Business Information
AMBION DIAGNOSTICS, INC.
Asuragen, Inc. 2150 WOODWARD ST AUSTIN, TX -
View profile »
Woman-Owned: No
Minority-Owned: No
HUBZone-Owned: No
 
Phase 1
Fiscal Year: 2004
Title: Enabling RNA Analyses from Formaldehyde-fixed Tissues
Agency: HHS
Contract: 1R43NS046958-01A1
Award Amount: $242,086.00
 

Abstract:

DESCRIPTION (provided by applicant): The overall goal is to develop a robust procedure for extracting RNA from tissues that have been fixed in formaldehyde, either neutral buffered formalin or buffered paraformaldehyde. Clinically interesting tissue from biopsies and necropsies is routinely archived after formaldehyde fixation by medical institutions. The primary use for these tissues is histological, as the RNA and other macromolecular components are so heavily crosslinked that they cannot be retrieved intact. The ability to use these samples in mRNA expression studies presents a huge opportunity. Procedures currently employed are inefficient and often are difficult to reproduce, so improvements are desired. A procedure is sought that provides RNA that is fully functional as a template for reverse transcription and can function in more classical analyses like Northern blots and ribonuclease protection assays (RPA's). Enhancement of the current extraction procedures by utilizing a mixture of hydrolytic enzymes and denaturants, to obtain RNA that has a normal appearance on electrophoresis will be pursued, as well as investigating methodologies for reversing the formaldehyde reaction. In the latter endeavor, methylene-N6-bis-adenosine, a common formaldehyde-generated crosslink, will be synthesized to be used as a test compound under controlled chemical tests to determine conditions for specifically removing the crosslink. This procedure can then be applied to RNA, either in situ (posthomogenization) or after extraction from tissue. Downstream procedures to analyze the RNA obtained will be extensively investigated, concentrating on reverse transcription (cDNA)-based methods. Much of this work will involve comparing the relative levels of different mRNA species inferred by these methods, especially between tissues split before fixing or freezing, to ensure the procedure does not bias the populational representation.

Principal Investigator:

Richard C. Conrad
5126510200
RCONRAD@AMBION.COM

Business Contact:

Rigo Vallejo
5126510200
RVALLEJO@AMBION.COM
Small Business Information at Submission:

AMBION, INC.
2130 WOODWARD STREET AUSTIN, TX 78744

EIN/Tax ID: 742515119
DUNS: N/A
Number of Employees: N/A
Woman-Owned: No
Minority-Owned: No
HUBZone-Owned: No