Comparison of Different Buffers for Protein Extraction from Formalin-Fixed and Paraffin-Embedded Tissue Specimens

We determined the best extraction buffer for proteomic investigation using formalin-fixation and paraffin-embedded (FFPE) specimens. A Zwittergent 3-16 based buffer, sodium dodecyl sulfate (SDS)-containing buffer with/without polyethylene glycol 20000 (PEG20000), urea-containing buffer, and FFPE-FAS...

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Bibliographic Details
Published in:PloS one 2015-11, Vol.10 (11), p.e0142650-e0142650
Main Authors: Shen, Kaini, Sun, Jian, Cao, Xinxin, Zhou, Daobin, Li, Jian
Format: Article
Language:English
Subjects:
Alkanes
Animals
Antigens
Brain
Brain Chemistry
Buffers
Chromatography, Liquid
Formaldehyde
Formaldehyde - chemistry
Gene expression
Hematology
Histology
Kidney - chemistry
Kidneys
Laboratory animals
Liver
Liver - chemistry
Lung - chemistry
Lungs
Mass spectrometry
Mass spectroscopy
Methods
Molecular weight
Morphology (Biology)
Paraffin
Paraffin Embedding
Pathology
Peptides
Polyethylene glycol
Proteins
Proteins - chemistry
Proteins - isolation & purification
Proteomics
Rats
Scientific imaging
Sodium
Sodium dodecyl sulfate
Sodium lauryl sulfate
Tandem Mass Spectrometry
Tissues
Urea
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Summary:We determined the best extraction buffer for proteomic investigation using formalin-fixation and paraffin-embedded (FFPE) specimens. A Zwittergent 3-16 based buffer, sodium dodecyl sulfate (SDS)-containing buffer with/without polyethylene glycol 20000 (PEG20000), urea-containing buffer, and FFPE-FASP protein preparation kit were compared for protein extraction from different types of rat FFPE tissues, including the heart, brain, liver, lung, and kidney. All of the samples were divided into two groups of laser microdissected (LMD) and non-LMD specimens. For both kinds of specimens, Zwittergent was the most efficient buffer for identifying peptides and proteins, was broadly applicable to different tissues without impairing the enzymatic digestion, and was well compatible with mass spectrometry analysis. As a high molecular weight carrier substance, PEG20000 improved the identification of peptides and proteins; however, such an advantage is limited to tissues containing submicrograms to micrograms of protein. Considering its low lytic strength, urea-containing buffer would not be the first alternative for protein recovery. In conclusion, Zwittergent 3-16 is an effective buffer for extracting proteins from FFPE specimens for downstream proteomics analysis.
ISSN:1932-6203
1932-6203
DOI:10.1371/journal.pone.0142650