Ulose membranes and incubated with sera from cancer patients and respective control donors. The clones reactive to sera are identified by sequencing [142]. Using this method >1000 TAA have been identified, including NY-ESO-1, which was discovered from an esophageal cancer cDNA library. Next to the use in human patients and clinical trials, SEREX has been alsoStroncek et al. Journal for ImmunoTherapy of Cancer (2017) 5:Page 11 ofemployed in murine transgenic models to predict TAA. Furthermore, a sera database has been established, which is for public access and allows the addition of data from other centers. However, one major limitation of this technology is the failure to detect post-translational modifications.PROTEOMEX/SERPAIn addition to SEREX, PROTEOMEX also termed SERPA was developed using two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) followed by Western blot analysis of the gels followed by their incubation with sera of patients and GGTI298MedChemExpress GGTI298 healthy volunteers [143, 144]. In addition, two-dimensional immune affinity chromatography followed by proteolysis and mass spectrometry has been used to identify novel TAA or respective biomarkers. Although these tools PubMed ID:https://www.ncbi.nlm.nih.gov/pubmed/28506461 are very robust, the disadvantages of these proteome-based technologies are their labor intensity with limitations in sample capacity, while the mapping could be automated in the future.Protein arraysOther proteome-based high-throughput analyses include automated protein microarrays of serum antibodies from cancer patients versus healthy controls. With this technology, a large series of proteins can be evaluated, which are either derived from cDNA or peptide phage display libraries [145]. By using protein arrays with a known panel of proteins, an induction of antibody responses against TAA has been recently demonstrated in a study using ipilimumab associated with GM-CSF treatment [146]. The development of antibodies to NY-ESO-1 posttreatment was identified in one clinical responder and one non-responder, suggesting that immunotherapy can induce immune responses to other known TAA. Other sources for immune genomic arrays apart from libraries include recombinant proteins or tumor lysates. The implementations of recombinant proteins for the array are multiple, but also more costly and may not account for post-translational modifications like SEREX. In order to take into account the effects of post-translational modifications on epitope recognition with respect to aberrant glycosylation of the tumor protein, high-throughput analysis using a glycopeptide discovery platform for proteomics profiling has been developed [147]. Although this glycopeptide platform allows high-throughput analyses, it has yet to be validated in particular regarding reproducibility and stability of this technique.SomaScanplasma (or other biological fluids). The SOMAscan assay is highly multiplexed, sensitive and quantitative. This assay is based on the use of a new generation of protein-capture SOMAmer reagent [148]. Native proteins contained in biological samples are captured by SOMAmers immobilized on streptavidin-agarose beads via a photo-cleavable biotin linker. Unbound proteins are washed away. The proteins captured are then biotinylated, the complex biotinylated protein/SOMAmer is released from the capture beads by the UV-induced photocleavage of the photosensitive linker. Magnetic-streptavidin beads capture the freed protein-SOMAmer complexes while the SOMAmers that did not bind to a protein ar.
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