Differential proteome analysis of the preeclamptic placenta using optimized protein extraction.
The human placenta is a difficult tissue to work with using proteomic technology since it contains large amounts of lipids and glycogen. Both lipids and glycogen are known to interfere with the first step in the two-dimensional polyacrylamide gel electrophoresis (2D-PAGE), the isoelectric focusing. In order to gain the best possible protein separation on 2D-PAGE, an optimized sample preparation protocol for placental proteins was developed. Two different buffers, urea/CHAPS and Hepes, were used for solubilization in combination with six different precipitation methods. The removal of glycogen from the samples by centrifugation was crucial for the final proteome maps. Solubilization with urea/CHAPS in combination with dichloromethane/methanol or acidified acetone proved to be the best precipitation procedures. When applied to clinical placenta samples apolipoprotein A1 was found to be accumulated in the preeclamptic placenta, where it may either have a nutritional effect or act as a modifier of signal transduction.
Centlow
Magnus
M
Department of Obstetrics & Gynecology, Lund University, BMC C14, 22184 Lund, Sweden.
Hansson
Stefan R
SR
Welinder
Charlotte
C
D000328:Adult; D016632:Apolipoprotein A-I / Q000032:analysis; D011232:Chemical Precipitation; D002793:Cholic Acids / Q000737:chemistry; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D005260:Female; D006003:Glycogen / Q000737:chemistry; D006531:HEPES / Q000737:chemistry; D006801:Humans; D007150:Immunohistochemistry; D008055:Lipids / Q000737:chemistry; D008875:Middle Aged; D010920:Placenta / Q000737:chemistry* / Q000378:metabolism; D011225:Pre-Eclampsia / Q000378:metabolism*; D011247:Pregnancy; D011257:Pregnancy Proteins / Q000032:analysis*; D020543:Proteome / Q000032:analysis*; D040901:Proteomics / Q000379:methods*; D015203:Reproducibility of Results; D012995:Solubility; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014508:Urea / Q000737:chemistry
D016632:Apolipoprotein A-I; D002793:Cholic Acids; D008055:Lipids; D011257:Pregnancy Proteins; D020543:Proteome; D014508:Urea; D006003:Glycogen; C028213:3-((3-cholamidopropyl)dimethylammonium)-1-propanesulfonate; D006531:HEPES