TaqMan PCR assay in the control of RNA normalization in human post-mortem brain tissue.

Author(s): Barrachina M, Castaño E, Ferrer I

Publication: Neurochem Int, 2006, Vol. 49, Page 276

PubMed ID: 16522342 PubMed Review Paper? No

Purpose of Paper

Conclusion of Paper

Studies

1. Study Purpose

   The purpose of this study was to compare two methods of RNA evaluation in the context of patient age, agonal state, brain pH, and postmortem interval.

   Summary of Findings:

   The authors report that electropherograms generated with the Agilent bioanalyzer were more sensitive in detecting RNA degradation than was conventional electrophoresis. No correlation was seen between RNA degradation and patient age, agonal state, brain pH, or postmortem interval.

   Biospecimens

   • Tissue - Brain

   Preservative Types

   • Frozen

   Diagnoses:

   • Not specified
   • Parkinson's Disease
   • Autopsy
   • Alzheimer's Disease
   • Lewy Body Disease

   Platform:

   Analyte	Technology Platform
   RNA	Spectrophotometry
   RNA	Automated electrophoresis/Bioanalyzer
   RNA	Electrophoresis

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   Preaquisition	Postmortem interval	2 h 3 h 4 h 5 h 6 h 7 h 8 h 10 h 12 h 13 h
   Preaquisition	Patient age	55 - 91 years
   Preaquisition	Diagnosis/ patient condition	Parkinson's disease Dementia with Lewy bodies, pure form Dementia with Lewy bodies, common form Alzheimer's disease Age-matched control
   Biospecimen Aliquots and Components	pH	Brain pH 6.11 - 7.02

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2. Study Purpose

   The purpose of this study was to assess several candidate control genes for normalization of RNA degradation in real-time quantitative reverse transcriptase polymerase chain reaction (qRT-PCR): human acidic ribosomal protein, beta-actin, cyclophilin, glyceraldehyde-3-phosphate dehydrogenase, phosphoglycerokinase, beta-2-microglobulin, beta-glucoronidase (GUS), hypoxanthine phosphoribosyltransferase, transcriptin factor IID TATA binding protein, and transferrin receptor. These potential control genes were compared to SOD1 and AMAD22 at a variety of postmortem intervals.

   Summary of Findings:

   While GUS and beta-actin were identified as candidate genes for normalization of qRT-PCR expression results to assess candidate gene degradation, GUS outperformed beta-actin with regards to inter-sample variability, stability with increasing PMI (beta-actin degradation was observed after 25 h), and susceptibility to degradation, although both genes did not vary among disease states.

   Biospecimens

   • Tissue - Brain

   Preservative Types

   • Frozen

   Diagnoses:

   • Parkinson's Disease
   • Lewy Body Disease
   • Autopsy
   • Not specified
   • Alzheimer's Disease

   Platform:

   Analyte	Technology Platform
   RNA	Real-time qRT-PCR

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   null Specific	Targeted nucleic acid	Human acidic ribosomal protein Beta-actin Cyclophilin Glyceraldehyde-3-phosphate dehydrogenase Phosphoglycerokinase Beta-2-microglobulin Beta-glucoronidase Hypoxanthine phosphoribosyltransferase Transcriptin factor IID TATA binding protein Transferrin receptor
   Preaquisition	Postmortem interval	5 h 9 h 25 h

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