title: Comparison of three methods of protein extraction from Dermatophagoides pteronyssinus for two-dimensional electrophoresis.
abstract: OBJECTIVE
  To explore an effective method of Dermatophagoides pteronyssinus protein extraction suitable for two-dimensional electrophoresis (2-DE) analysis.
  
  
  METHODS
  The extracts of Dermatophagoides pteronyssinus were prepared with Coca's solution, lysis buffer of 2-DE, and Trizol reagent, respectively. Bicinchoninic acid (BCA) assay was used to determine the total protein concentration of the samples. The efficiency of different protein extraction methods were evaluated with 2-DE analysis.
  
  
  RESULTS
  The concentrations of extracted protein by methods of Coca's solution, lysis buffer, and Trizol reagent were 0.63 g/L, 0.90 g/L, and 0.80 g/L, respectively. The 2-DE analysis results showed that some protein spots in low molecular weight (LMW) range could be detected with the Coca's solution method. With the lysis buffer of 2-DE method, more protein spots in LMW range could be detected, while the medium molecular weight (MMW) protein spots were absent. Several MMW protein spots (174-178 kD and 133 kD) and more LMW protein spots were detected with Trizol reagent method.
  
  
  CONCLUSIONS
  Among Coca's solution, lysis buffer of 2-DE, and Trizol reagent, the concentration of extracted protein of Dermatophagoides pteronyssinus by lysis buffer of 2-DE is the highest. However, most protein components of Dermatophagoides pteronyssinus purified mite bodies can be extracted by Trizol reagent, which may generally reflect the whole profile of Dermatophagoides pteronyssinus allergens.
authors.affiliation: Department of Allergy, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing, China. Sunjl5@yahoo.com
authors.forename: Jin-lu
authors.initials: JL
authors.lastname: Sun
authors.forename: Hong-yu
authors.initials: HY
authors.lastname: Zhang
authors.forename: Zhi-yi
authors.initials: ZY
authors.lastname: Guo
authors.forename: Wan-tao
authors.initials: WT
authors.lastname: Ying
authors.forename: Xiao-hong
authors.initials: XH
authors.lastname: Qian
authors.forename: Jing-lan
authors.initials: JL
authors.lastname: Wang
mesh_terms: D000485:Allergens / Q000302:isolation & purification; D000818:Animals; D040002:Dermatophagoides pteronyssinus / Q000737:chemistry*; D015180:Electrophoresis, Gel, Two-Dimensional; D006146:Guanidines / Q000737:chemistry; D010636:Phenols / Q000737:chemistry; D011506:Proteins / Q000302:isolation & purification*
chemical_list: D000485:Allergens; D006146:Guanidines; D010636:Phenols; D011506:Proteins; C411644:trizol