Protein Function Microarrays: Design, Use and Bioinformatic Analysis in Cancer Biomarker Discovery and Quantitation
Protein microarrays have many potential applications in the systematic, quantitative analysis of protein function, including in biomarker discovery applications. In this chapter, we review available methodologies relevant to this field and describe a simple approach to the design and fabrication of cancer-antigen arrays suitable for cancer biomarker discovery through serological analysis of cancer patients. We consider general issues that arise in antigen content generation, microarray fabrication and microarray-based assays and provide practical examples of experimental approaches that address these. We then focus on general issues that arise in raw data extraction, raw data preprocessing and analysis of the resultant preprocessed data to determine its biological significance, and we describe computational approaches to address these that enable quantitative assessment of serological protein microarray data. We exemplify this overall approach by reference to the creation of a multiplexed cancer-antigen microarray that contains 100 unique, purified, immobilised antigens in a spatially defined array, and we describe specific methods for serological assay and data analysis on such microarrays, including test cases with data originated from a malignant melanoma cohort.
KeywordsProtein microarrays Cancer–testis antigens Cancer biomarker discovery Bioinformatic analysis Pipeline
The authors thank Dr Aubrey Shoko, Dr Natasha Beeton-Kempen and Dr Judit Kumuthini for their help in generating the data herein. We thank the Centre for Proteomic & Genomic Research, Cape Town, for access to equipment and assistance in developing the CT100 array. JMB thanks the National Research Foundation (NRF), South Africa, for a Research Chair. The research was supported by grants from the NRF, University of Cape Town (UCT) and Marion Beatrice Waddel.
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