Mouse-specific tandem IgY7-SuperMix immunoaffinity separations for improved LC-MS/MS coverage of the plasma proteome.
We report on a mouse specific SuperMix immunoaffinity separation system for separating low-abundance proteins from high and moderate abundance proteins in mouse plasma. When applied in tandem with a mouse IgY7 column that removes the seven most abundant proteins in plasma, the SuperMix column captures more than 100 additional moderate abundance proteins, thus allowing significant enrichment of low-abundance proteins in the flow-through fraction. A side-by-side comparison of results obtained from 2D-LC-MS/MS analyses of flow-through samples from IgY7 and SuperMix columns revealed a nearly 2-fold improvement in the overall proteome coverage. Detection of low-abundance proteins was also enhanced, as evidenced by a more than 2-fold increase in the coverage of cytokines, growth factors, and other low-abundance proteins. Moreover, the tandem separations are automated, reproducible, and allow effective identification of protein abundance differences from LC-MS/MS analyses. Considering the overall reproducibility and increased sensitivity using the IgY7-SuperMix separation system, we anticipate broad applications of this strategy for biomarker discovery using mouse models.
Zhou
Jian-Ying
JY
Biological Science Division and Environmental Molecular Sciences Laboratory, Pacific Northwest National Laboratory, Richland, Washington 99352, USA.
Petritis
Brianne O
BO
Petritis
Konstantinos
K
Norbeck
Angela D
AD
Weitz
Karl K
KK
Moore
Ronald J
RJ
Camp
David G
DG
Kulkarni
Rohit N
RN
Smith
Richard D
RD
Qian
Wei-Jun
WJ
D000818:Animals; D015415:Biomarkers / Q000737:chemistry / Q000378:metabolism; D002846:Chromatography, Affinity / Q000295:instrumentation / Q000379:methods; D002853:Chromatography, Liquid / Q000295:instrumentation / Q000379:methods*; D006801:Humans; D007118:Immunoassay / Q000295:instrumentation / Q000379:methods; D007136:Immunoglobulins / Q000276:immunology*; D008297:Male; D051379:Mice; D008969:Molecular Sequence Data; D010949:Plasma / Q000737:chemistry*; D020543:Proteome / Q000032:analysis*; D040901:Proteomics / Q000295:instrumentation / Q000379:methods; D053719:Tandem Mass Spectrometry / Q000295:instrumentation / Q000379:methods*
D015415:Biomarkers; C016512:IgY; D007136:Immunoglobulins; D020543:Proteome