Validation of serum protein profiles by a dual antibody array approach |
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| Affiliation: | 1. Institute of Chemical Biology and Fundamental Medicine SB RAS, 8, Lavrentiev Avenue, Novosibirsk, 630090, Russia;2. M.M. Shemyakin and Yu.A. Ovchinnikov Institute of Bioorganic Chemistry RAS, 16/10, Miklukho-Maklaya str., Moscow, 117997, Russia;3. Institute of Fundamental Medicine and Biology, Kazan Federal University, 18 Kremlyovskaya str., Kazan, Tatarstan, 420008 Russia;1. Robert Koch Institute, Division 16 Mycotic and Parasitic Agents and Mycobacteria, Nordufer 20, 13353 Berlin, Germany;2. Freie Universität Berlin, Centre for Infection Medicine, Institute of Microbiology and Epizootics, Robert-von-Ostertag-Str. 7-13, 14163 Berlin, Germany;1. Center for Genetic Epidemiology and Genomics, School of Public Health, Medical College of Soochow University, Suzhou, Jiangsu 215123, China;2. Jiangsu Key Laboratory of Preventive and Translational Medicine for Geriatric Diseases, Soochow University, Suzhou, Jiangsu 215123, China;3. Suzhou Center for Disease Prevention and Control, Suzhou, Jiangsu, China;4. Department of Orthopedics, Sihong People''s Hospital, Suqian, Jiangsu 223900, China;5. Loujiang Community Health Service Center, Suzhou Gusu District, Suzhou, Jiangsu, China;6. Disease Prevention and Control Center of Suzhou High Tech Zone, Suzhou, Jiangsu, China;7. Shishan Community Health Service Center, Suzhou High Tech Zone, Suzhou, Jiangsu, China;1. Department of Food Safety and Infection Biology, Center for Epidemiology and Biostatistics, Norwegian School of Veterinary Science, P.O. Box 8146 Dep., 0033 Oslo, Norway;2. Department of Epidemiology and Biostatistics/Population Health, College of Public Health, 132 Coverdell Center, University of Georgia Athens, GA 30602-7396, USA;3. Department of Veterinary Public Health, School of Veterinary Medicine, Makerere University, P.O. Box 7062, Kampala, Uganda;4. Norwegian Veterinary Institute, P.O. Box 750, N-0106 Oslo, Norway;5. Department of Food Safety and Infection Biology, Section of Arctic Veterinary Medicine, Norwegian School of Veterinary Science, Stakkevollveien 23, 9010 Tromsø, Norway;6. The Roslin Institute, College of Medicine and Veterinary Medicine, University of Edinburgh, Easter Bush, Roslin, Midlothian EH25 9RG, United Kingdom |
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| Abstract: | In recent years, affinity-based technologies have become important tools for serum profiling to uncover protein expression patterns linked to disease state or therapeutic effects. In this study, we describe a path towards the production of an antibody microarray to allow protein profiling of biotinylated human serum samples with reproducible sensitivity in the picomolar range. With the availability of growing numbers of affinity reagents, protein profiles are to be validated in efficient manners and we describe a cross-platform strategy based on data concordance with a suspension bead array to interrogate the identical set of antibodies with the same cohort of serum samples. Comparative analysis enabled to screen for high-performing antibodies, which were displaying consistent results across the two platforms and targeting known serum components. Moreover, data processing methods such as sample referencing and normalization were evaluated for their effects on inter-platform agreement. Our work suggests that mutual validation of protein expression profiles using alternative microarray platforms holds great potential in becoming an important and valuable component in affinity-based high-throughput proteomic screenings as it allows to narrow down the number of discovered targets prior to orthogonal, uniplexed validation approaches. |
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