Table 4. Advantages and disadvantages of platformsa

PlatformAdvantages for Biomarker DiscoveryDisadvantages for Biomarker Discovery
a 2-D DIGE, two-dimensional difference gel electrophoresis; TOF, time of flight mass spectrometry; SELDI, surface-enhanced laser desorption ionization; ICAT, isotope-coded affinity tags; SNP, single nucleotide polymorphism.
b Proteins from normal and diseased samples are labelled with different flurescent dyes and then separated by two dimensional electrophoresis. Size of peptide (mass to charge ratio) is calculated based on the length of time for the peptide to travel through a vacuum.
c Proteins from a sample(s) bind to a chip if the coating of the chip allows an adequate protein-surface affinity. For example, hydrophobic proteins bind to a hydrophobic chip surface. Then the proteins are identified by a TOF mass spectrometer.
d Complex peptide mixtures are separated by chromatography (e.g., reverse phase, cation exchange), then the chromatography fractions are analysed by TOF mass spectrometry. When two TOF mass spectrometers are used in “series,” this is referred to as MS/MS. This allows actual peptide sequencing.
e Proteins from two different sources (e.g., disease versus normal) can be labelled with “light” and “heavy” tags. After LC/MS/MS, the relative abundances of different peptides in the two samples can be calculated.
Measurement of mRNA expression (e.g., differential display, SAGE, microarray)Able to screen large number of “genes”RNA levels may not directly relate to protein levels
Commercially availableProvide no information about posttranslational protein modifications
Difficult to handle large volume of data
2-D DIGEbAssay of the actual biomarker not mRNAPoor technique for difficult-to-solubilize proteins (e.g., membrane proteins), low-abundance proteins, and low-molecular-weight proteins
Allows identification of previously unknown biomarkers
Can quantify amplitude of change in biomarker
Well established techniqueNot high throughput, i.e., labor intensive
SELDIcWell suited to generating a pattern of peptide peaks corresponding to a disease biomarkerDifficult to identify proteins
Difficult to measure protein abundance
High throughput, less labor intensive, and cheaper than 2-D electrophoresisSpecimen handling can have large impact on quality
Can focus on certain subsets of proteins
LC/MS/MSdHigher throughput than 2D DIGENeed to use ICATe to measure biomarker abundance
Can identify protein by amino acid sequencing
Increased yield of membrane proteins and low-abundance proteins
Tissue microarrayHigh-throughput validation and prioritization of tissue biomarkers (Pepe stage 1b)Immunohistochemistry: need antibody; cannot detect “unknown” proteins.
Obtain protein location by immunohistochemistryIn situ hybridization—detects mRNA only
Quantitation issues
Specimen quality issues
SNP detectionMay produce unexpected new leads about pathogenesis of and biomarkers for diseaseOnly gives information about an individual’s risk of disease, not presence of disease per se
Provides no information about expression of protein