Identified the nonspecific binding proteins in depletion of Albumin and IgG from Human plasma
Wang Yundan1, Ning Yunshan1,3, Jiang Yin2, Deng Xinyu2, Fang Qinmei2, Hong Yanhua3,
Li Ming1,3
1 College of Biotechnology, Southern Medical University, Guangzhou, P. R. China, 510515
2 Beijing Institute of radiation Medicine, Beijing, P. R. China, 100850
3 Boang Antibody Company, Shanghai, P. R. China, 200233 to
mmy604@fimmu.com
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Depletion of high abundant proteins in plasma samples was necessary for the further study of new biomarkers mining in HPPP. We used the high specific mouse mAb against human albumin and Protein G to remove Albumin and IgG respectively from human plasma in denatured condition and native condition. We observed the different capacity of depletion in the presence of chaos reagents, non-ionic detergent and high concentration of salts. In native condition, the elution proteins were separated by 2DE and 104 spots in the gel were excised and trypsin digested for tandem mass spectrum (MS/MS) analysis. The binding proteins including Albumin, IgG, Fibrinogen, Vitamin D binding protein, Alpha-1 antitrypsin, transferrin, Transthyretin, Proapolipoprotein, Keratin, Complement component 3. The remained spots are albumin and IgG fragments. In denatured condition, the capacity of depletion for albumin become lower but IgG not affected. The concentration of nonspecific binding proteins including the fragments of Albumin in elution sample was lower. The results may explain the relation between low non-specific binding and presence of albumin fragments in condensed plasma samples processed by MARC or MARS system using commercial buffer.
Keywords:
High abundant protein / Depletion / 2-DE / MS / Nonspecific / Human plasma protein / Monoclonal antibody / Denature
References
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