Immunoaffinity separation of plasma proteins by IgY microbeads: meeting the needs of proteomic sample preparation and analysis.
Separation of complex protein mixtures that have a wide dynamic range of concentration, such as plasma or serum, is a challenge for proteomic analysis. Sample preparation to remove high-abundant proteins is essential for proteomics analysis. Immunoglobulin yolk (IgY) antibodies have unique and advantageous features that enable specific protein removal to aid in the detection of low-abundant proteins and biomarker discovery. This report describes the efficiency and effectiveness of IgY microbeads in separating 12 abundant proteins from plasma with an immunoaffinity spin column or LC column. The protein separation and sample preparation process was monitored via SDS-PAGE, 2-DE, LC-MS/MS, or clinical protein assays. The data demonstrate the high specificity of the protein separation, with removal of 95-99.5% of the abundant proteins. IgY microbeads against human proteins can also selectively remove orthologous proteins of other mammals such as mouse, rat, etc. Besides the specificity and reproducibility of the IgY microbeads, the report discusses the factors that may cause potential variations in protein separation such as protein-protein interactions (known as "Interactome"), binding and washing conditions of immunoaffinity reagents, etc. A novel concept of Seppromics is introduced to address methodologies and science of protein separation in a context of proteomics.
Huang
Lei
L
GenWay Biotech, San Diego, CA 92121, USA.
Harvie
Gulia
G
Feitelson
Jerald S
JS
Gramatikoff
Kosi
K
Herold
David A
DA
Allen
David L
DL
Amunngama
Ravi
R
Hagler
Rachel A
RA
Pisano
Michael R
MR
Zhang
Wei-Wei
WW
Fang
Xiangming
X
D000418:Albumins / Q000737:chemistry; D000818:Animals; D015415:Biomarkers / Q000737:chemistry; D001798:Blood Proteins / Q000737:chemistry* / Q000302:isolation & purification*; D002853:Chromatography, Liquid; D004492:Edetic Acid / Q000737:chemistry; D015180:Electrophoresis, Gel, Two-Dimensional; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D007136:Immunoglobulins / Q000737:chemistry*; D013058:Mass Spectrometry; D051379:Mice; D008863:Microspheres; D040901:Proteomics / Q000379:methods*
D000418:Albumins; D015415:Biomarkers; D001798:Blood Proteins; C016512:IgY; D007136:Immunoglobulins; D004492:Edetic Acid