2D DIGE analysis of serum after fractionation by ProteoMiner™ beads.
Serum is a popular biofluid used for many protein biomarker discovery projects since the collection and processing of serum/plasma is relatively noninvasive and inexpensive. Unfortunately, the downstream analysis of serum/plasma is hampered severely by several high-abundant proteins which often interfere with the separation and detection of many of the proteins of lower abundance. Thus, a number of prefractionation methods have recently been developed with the view to reduce the dynamic range of these proteins. These include both dye- and immunoaffinity-based methods that are specifically designed to remove serum albumin. In this chapter, we describe an alternative method using ProteoMiner™ or Equalizer beads that is aimed at overcoming this problem in serum. This method uses a combinatorial library of hexapeptides bound to beads and works by binding proteins until saturation is reached. Thus, the high-abundant proteins will reach saturation quickly, while the lower-abundant proteins continue to bind. This results in a dramatic depletion of the most abundant proteins, with a concurrent concentration of the middle- to low-abundant proteins.
Department of Biological Science, Faculty of Science, National University of Singapore, Singapore, Singapore.
Cynthia
C
Liang
Gek San
GS
Tan
Maxey C M
MC
Chung
D001798:Blood Proteins / Q000032:analysis* / Q000737:chemistry / Q000302:isolation & purification; D005591:Chemical Fractionation / Q000379:methods*; D020650:Combinatorial Chemistry Techniques; D005590:Fractional Precipitation; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007091:Image Processing, Computer-Assisted; D007525:Isoelectric Focusing; D008863:Microspheres; D020543:Proteome / Q000032:analysis* / Q000737:chemistry / Q000302:isolation & purification; D057825:Two-Dimensional Difference Gel Electrophoresis / Q000379:methods*
D001798:Blood Proteins; D020543:Proteome