Optimized protein extraction from cryopreserved brain tissue samples

Author(s): Ericsson C, Peredo I, Nistér M

Publication: Acta Oncol, 2007, Vol. 46, Page 10

PubMed ID: 17438701 PubMed Review Paper? No

Purpose of Paper

Conclusion of Paper

Studies

1. Study Purpose

   Comparative analysis of protein recovery using different methods of tissue homogenization (mortar grinding, shaking, sonication, mill grinding) followed by extraction by incubaction in SDS buffer.

   Summary of Findings:

   Extraction efficiency was greatest when tissues were processed by sonication or frozen ball mill grinding.

   Biospecimens

   • Tissue - Brain

   Preservative Types

   • Frozen

   Diagnoses:

   • Neoplastic - Carcinoma

   Platform:

   Analyte	Technology Platform
   Protein	1D/2D gels

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   Analyte Extraction and Purification	Tissue homogenization	Mill grinding Mortar grinding Shaking Sonication

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2. Study Purpose

   Comparative analysis of protein recovery with different extraction/ solubilization (1) temperatures, (2) volumes, and (3) durations.

   Summary of Findings:

   Protein recovery was equivalent at all extraction/ solubilization temperatures evaluated (70, 80, 95 degrees C). However, results of Western blot analysis for Na/K ATPase and SAPK/JNK indicated expression was greater when samples were solubilized at 70 degrees, while p44/42 MAPK levels were comparable across samples. Extraction efficiency plateaued at an extraction/ solubilization volume 10X the initial tissue volume. Extraction efficiency was not influenced by the duration of extraction/ solubilization.

   Biospecimens

   • Tissue - Brain

   Preservative Types

   • Frozen

   Diagnoses:

   • Neoplastic - Carcinoma

   Platform:

   Analyte	Technology Platform
   Protein	1D/2D gels
   Protein	Western blot

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   Analyte Extraction and Purification	Protein solubilization	2 min 5 min 10 min 30 min 60 min 2.5 X volume 5 X volume 10 X volume 40 X volume 70 degrees C 80 degrees C 95 degrees C

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3. Study Purpose

   To assess whether repeated exposure to freeze thaw cycles impacted the quantity of soluble protein or Na/K ATPase protein expression.

   Summary of Findings:

   Exposure of protein samples to 20 freeze thaw cycles did not affect the quantity of soluble protein of Na/K ATPase expression.

   Biospecimens

   • Tissue - Brain

   Preservative Types

   • Frozen

   Diagnoses:

   • Neoplastic - Carcinoma

   Platform:

   Analyte	Technology Platform
   Protein	Western blot
   Protein	1D/2D gels

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   Storage	Freeze/thaw cycling	0 cycles 20 cycles

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