Application and establishment of two-dimensional gel electrophoresis from proteome analysis of human osteosarcoma / 吉林大学学报(医学版)

ABSTRACT

Objective To establish analysis methods of two-dimensional(2-DE)gel electrophoresis for human osteosarcoma.Methods A series of methods,including Immobilized pH gradient were used as ID.Some applications,such as sample preparation used as choice of IPG gel,were improved.Coomassie brilliant blue staining,ImageMaster 2D Elite 3.01 analysis software,MALDI-TOF/ TOF MS and SWISS-PROT database serching were used to separate and indentify the proteins of human osteosarcoma.Results The good use pattern including repetitive experiments showed that in three experiments,the amount of protein spots of the same team sample deviates from the relative standard as following,the average of variation coefficient(%) 23.00?10.11 and 20.33?9.90;and the range of variation coefficient(%) were3.80-6.89 and 2.706.89 from osteosarcoma and normal group respectively.The isoelectric point and molecular weigh of the same protein spots in three experiments deviated from relative standard as following(8.93%?1.17)%,(10.16?2.02)%,(10.87?3.86)%,respectively.Therefore,better resolution and repetitive 2-DE atlas were obtained.The proteins from 11 pairs of sample were analysed by mass spectrometry,9 identified proteins(transthyretin precursor, Triosephosphate isomerase,slow skeletal muscle,cardiac muscle Troponin T,Cofilin-1,Myosin light chain 1,Calgranulin B,Heat-shock protein,Annexin A5, Fanconi anemia group D2 proteins) were more abundant in osteosarcoma tisstues and 2 proteins manganese SOD and carbonic dehydratase appeared down-regulation in osteosarcoma tisstues.Conclusion This optimized 2-DE map is an important tool for further study on osteosarcoma,and these identified proteins were related-proteins with osteosarcoma.It is suggested that the changes of the proteome are involved in the pathology processe of osteosarcoma.