Multivariate analysis of RNA levels from postmortem human brains as measured by three different methods of RT-PCR.

Author(s): Johnston N L, Cervenak J, Shore A D, Torrey E F, Yolken R H

Publication: J Neurosci Methods, 1997, Vol. 77, Page 83

PubMed ID: 9402561 PubMed Review Paper? No

Purpose of Paper

Conclusion of Paper

Studies

1. Study Purpose

   The purpose of this paper was to evaluate the influence of pre-and postmortem factors on GAPDH mRNA levels in the occipital cortices as measured by reverse transcription-polymerase chain reaction (RT-PCR). Three RT-PCR priming methods were compared: oligo dT, GAP specific primer, and random primer.

   Summary of Findings:

   pH was consistent across the 10 brain regions examined. High brain pH was correlated with age at death, ventilation and CO poisoning, and a quick death (low agonal state). GAPDH levels did not vary between diagnostic groups. While priming method was the best predictor of quantifiable cDNA levels, method-specific differences in sensitivity to preanalytical variables were observed. Prolonged frozen storage (24 months) negatively impacted quantifiable cDNA levels in oligo dT primed samples at a rate of 7% per month. Cause of death was also correlated to priming method, with decreased cDNA levels measured in oligo dT primed samples obtained from suicide victims, and in random hexamer and gene-specific primed samples obtained from CO poisoning victims. An elevated pH translated to varying degrees of increase in measured cDNA levels from differentially primed samples, with oligo dT yielding the most robust increase of 25-fold for every pH point.

   Biospecimens

   • Tissue - Brain

   Preservative Types

   • Frozen

   Diagnoses:

   • Normal
   • Schizophrenia
   • Autopsy
   • Depression
   • Other diagnoses
   • Bipolar Disorder

   Platform:

   Analyte	Technology Platform
   RNA	Spectrophotometry
   RNA	RT-PCR

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   Biospecimen Aliquots and Components	pH	5.4 - 6.3
   Preaquisition	Cause of death	CO poisoning Other suicide Other, non-suicide
   Preaquisition	Diagnosis/ patient condition	Schizophrenia Bipolar disorder Depression Other
   Preaquisition	Rapidity of death	Instantaneous Within 24 h of cause, no hypoxia Over a > 24 h period
   Preaquisition	Patient age	17 - 83 years
   Preaquisition	Postmortem interval	5 - 133 h
   Storage	Storage duration	2 - 24 months
   RT-PCR Specific	Priming method	Oligo dT Random hexamer Gene-specific (GAPDH)

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