Surface modified capillary electrophoresis combined with in solution isoelectric focusing and MALDI-TOF/TOF MS: a gel-free multidimensional electrophoresis approach for proteomic profiling--exemplified on human follicular fluid.
Development of miniaturized analytical tools continues to be of great interest to face the challenges in proteomic analysis of complex biological samples such as human body fluids. In the light of these challenges, special emphasis is put on the speed and simplicity of newly designed technological approaches as well as the need for cost efficiency and low sample consumption. In this study, we present an alternative multidimensional bottom-up approach for proteomic profiling for fast, efficient and sensitive protein analysis in complex biological matrices. The presented setup was based on sample pre-fractionation using microscale in solution isoelectric focusing (IEF) followed by tryptic digestion and subsequent capillary electrophoresis (CE) coupled off-line to matrix assisted laser desorption/ionization time of flight tandem mass spectrometry (MALDI TOF MS/MS). For high performance CE-separation, PolyE-323 modified capillaries were applied to minimize analyte-wall interactions. The potential of the analytical setup was demonstrated on human follicular fluid (hFF) representing a typical complex human body fluid with clinical implication. The obtained results show significant identification of 73 unique proteins (identified at 95% significance level), including mostly acute phase proteins but also protein identities that are well known to be extensively involved in follicular development.
Hanrieder
Jörg
J
Department of Physical and Analytical Chemistry, Analytical Chemistry, Uppsala University, Uppsala, Sweden.
Zuberovic
Aida
A
Bergquist
Jonas
J
D019075:Electrophoresis, Capillary / Q000191:economics / Q000295:instrumentation / Q000379:methods*; D005260:Female; D015571:Follicular Fluid / Q000737:chemistry*; D006801:Humans; D007525:Isoelectric Focusing / Q000191:economics / Q000379:methods*; D011506:Proteins / Q000032:analysis*; D040901:Proteomics / Q000191:economics / Q000295:instrumentation / Q000379:methods*; D015203:Reproducibility of Results; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*; D053719:Tandem Mass Spectrometry / Q000379:methods
D011506:Proteins