Placental tissue as model for pilot study focused on RNA analysis from human foetal tissue.

Author(s): Hulkova, Zeman J

Publication: Prague Med Rep, 2011, Vol. 112, Page 93-101

PubMed ID: 21699758 PubMed Review Paper? No

Purpose of Paper

Conclusion of Paper

Studies

1. Study Purpose

   The aims of this study were to determine if a 1 or 2 h delay at 0 or 24 degrees C prior to snap freezing affects the yield or integrity of RNA, and to identify promising reference genes for semi-quantitative RT-PCR analysis of placental tissue. Placenta specimens were collected from 20 healthy individuals following the birth of infants with Apgar scores ranging from 3 to 7. Placenta specimens were rinsed in physiological buffer prior to immediate snap freezing in liquid nitrogen or a 1-2 h delay at 0 or 24 degrees C.

   Summary of Findings:

   For placenta specimens snap frozen immediately, RNA purity (determined by OD 260/280 ratio) was negatively correlated to RNA yield (r= -0.61, p<0.01), and although it is unclear whether specimen size was a controlled variable the authors report a positive correlation between RNA yield and sample weight (r=0.46, p<0.05). Placenta specimens subjected to 1 or 2 h of cold ischemia at 0 degrees C displayed a similar negatively correlation between RNA yield and purity (r= -0.94 and r= -0.83, respectively; p<0.05). Although data was not shown, the authors note that poor RNA quality was associated with a low Apgar score, while RNA yield remained unaffected. RIN was also adversely, although not significantly, affected by the longer ischemia times and the higher temperature, with lower RINs observed after 2 h compared to 1 h at 0 degrees C (5.0 versus 5.4, respectively), and at 24 degrees (3.5 versus 4.4, respectively). The abundance, but not rate of success, of a 378 bp RT-PCR amplicon was influenced by the duration and temperature of cold ischemia. Specimens subjected to 1 h of cold ischemia at 0 degrees C generated the most abundant amplicon, although the temperature of cold ischemia produced more robust differences in amplicon intensity than the duration.

   Biospecimens

   • Tissue - Placenta

   Preservative Types

   • Frozen

   Diagnoses:

   • Normal

   Platform:

   Analyte	Technology Platform
   RNA	Automated electrophoresis/Bioanalyzer
   RNA	Electrophoresis
   RNA	RT-PCR
   RNA	Spectrophotometry

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   Biospecimen Acquisition	Cold ischemia time	0 h 1 h 2 h
   Storage	Storage temperature	0 degrees C 24 degrees C
   Preaquisition	Diagnosis/ patient condition	Low Apgar score (3-7) High Apgar score (8-10)
   RT-PCR Specific	Targeted nucleic acid	MECP2

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