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Antibody suspension bead arrays with multi-parallel CSF profiling for biomarker discovery

The antibody suspension bead array technology has been shown to be highly applicable for discovery of potential biomarkers in blood derived samples [1]. For analysis in neurodegenerative disorders like multiple sclerosis (MS), cerebrospinal fluid (CSF) is another highly interesting body fluid for protein profiling efforts [2]. With the antibody array approach, labelled samples are used to create relative protein profiles in a highly multiplexed way. Currently, up to 500 different antibodies can be combined and simultaneously analyzed in one sample and the samples can be processed in 96 or 384 well formats, thus enabling screening of 96 or 384 samples in parallel. The procedure is thereby also adaptable to standard robotics and instrumentation and another advantage is that only small volumes of samples are required. The availability of well validated capture agents is an important matter for large scale profiling studies. This issue is being addressed by the Human Protein Atlas project, which aims at producing mono-specific antibodies targeting all proteins in the human genome in a high-throughput manner [3]. The presented study aimed at developing a protocol for protein profiling of CSF and to use this assay to screen for differentially expressed proteins in samples from MS patients. Antibodies from the Human Protein Atlas project were selected, coupled to the surface of microspheres and assembled into a 76plex array in suspension. The samples analyzed were obtained from two sample sets from MS patients and patients with other neurological diseases (OND). An initial screening of 135 samples indicated that proteins differing between the sample groups could be found. Statistical analysis resulted in around 20 proteins showing clear significant differences in at least one of the sample sets. As a second stage, two antibodies for each of these candidates were used to reveal concordant results for both sample sets and antibodies. This further supports the possibility that these proteins could be regarded as biomarker candidates. Results from further validations, including data from a set of 400 new CSF samples together with the outcome of analysis on other technical platforms will be presented. Also, co-profiling of plasma from the same individuals will be made to investigate the possible presence of a correlating profile in the different body fluids.

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