Interest of major serum protein removal for Surface-Enhanced Laser Desorption/Ionization – Time Of Flight (SELDI-TOF) proteomic blood profiling

Stéphane Roche,Marie-Thérèse Piva,Monique Provansal,Laurent Tiers,Sylvain Lehmann

doi:10.1186/1477-5956-4-20

Abstract

BackgroundSurface-Enhanced Laser Desorption/Ionization – Time Of Flight (SELDI-TOF) has been proposed as new approach for blood biomarker discovery. However, results obtained so far have been often disappointing as this technique still has difficulties to detect low-abundant plasma and serum proteins.ResultsWe used a serum depletion scheme using chicken antibodies against various abundant proteins to realized a pre-fractionation of serum prior to SELDI-TOF profiling. Depletion of major serum proteins by immunocapture was confirmed by 1D and 2D gel electrophoresis. SELDI-TOF analysis of bound and unbound (depleted) serum fractions revealed that this approach allows the detection of new low abundant protein peaks with satisfactory reproducibility.ConclusionThe combination of immunocapture and SELDI-TOF analysis opens new avenues into proteomic profiling for the discovery of blood biomarkers.

Highlights

Surface-Enhanced Laser Desorption/Ionization – Time Of Flight (SELDI-TOF) has been proposed as new approach for blood biomarker discovery
We evaluated the interest of the removal of major serum protein for SELDI-TOF analysis
While 1 μL of undepleted serum was analysed on each SELDI-TOF spot, the immunodepletion was performed on 10 μL of serum and 1/3 of the resulting depleted fraction was analysed, resulting in a 3.3 fold increase in the serum equivalent amount analysed

Summary

Introduction

Surface-Enhanced Laser Desorption/Ionization – Time Of Flight (SELDI-TOF) has been proposed as new approach for blood biomarker discovery. The few most abundant blood proteins constitute 95% of the bulk mass of proteins but they represent less than 0.1% of the total number of proteins These high abundant proteins, and in particular albumin, produce large signals in most proteomics approaches and they mask or interfere with the detection of the other low amount protein components. This situation explains why the discovery of new proteins or peptides biomarkers in blood is challenging. Many methods rely on a multidimensional separation scheme combining for example multidimensional chromatography or electrophoresis and mass spectrometry (MS) [4,5] This is the case of the Surface-Enhanced (page number not for citation purposes)

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Conclusion