Novel Type of Protein Chip for Multiplex Detection of Autoantibodies

Christer Wingren

Disclosures

Expert Rev Proteomics. 2013;10(5):417-420. 

In This Article

Abstract and Introduction

Abstract

Evaluation of: Akada J, Kamei S, Ito A et al. A new type of protein chip to detect hepatocellular carcinoma-related autoimmune antibodies in the sera of hepatitis C virus-positive patients. Proteome Sci. 11(1), 33 (2013).

Unlocking the proteome and delivering biomarkers to the clinic will be critical for early and improved diagnosis and prognosis. Conventional protein microarrays have evolved as a promising proteomic technology with great potential for protein expression profiling in health and disease. In this study, Akada et al. explore a new type of protein chip, interfaced with a dual-color fluorescence-based read-out, for screening of autoantibodies in serum. Uniquely, the recombinant antigens were microarray adapted by molecular design to contain a five-cysteine tag for immobilization and green fluorescent protein for detection (color 1). The engineered antigens were immobilized on in-house-designed maleimide-incorporated diamond-like carbon substrates and subsequently heat treated in a solution of denaturing and reducing agents before any specifically bound serum autoantibodies were detected (color 2). The authors used a 4-plex array targeting hepatocellular carcinoma-related autoantibodies in the sera of hepatitis C virus-positive patients as model system to demonstrate proof-of-concept.

Introduction

Protein microarrays have evolved as a promising proteomic technology with great potential for protein expression profiling in health and disease.[1–3] Depending on their applications, protein arrays can be classified into two types: analytical and functional protein arrays. Analytical microarrays, or affinity microarrays, are composed of highly specific binders, mainly antibodies, and used to perform protein expression profiling, biomarker discovery and so on.[1,3,4] Functional protein microarrays are based on printing a large number of individual proteins, and used to investigate biochemistry properties, antigenicity and/or activities of those immobilized proteins.[2]

Focusing on functional protein microarrays, a variety of proteins, ranging from allergens, autoantigens and enzymes to even whole proteomes, have been arrayed.[2,5] The proteins are normally deposited one-by-one in the format of intact, purified proteins, intended to be displayed as 'native' proteins with retained structural and functional properties. Functional protein microarrays have been successfully used to identify protein–ligand (e.g., protein, antibody, lipid, small molecules, DNA and RNA) interactions, and to identify substrates or enzymes in, for example, phosphorylation, ubiquitylation and acetylation, as well as to profile immune responses, including autoantibody repertoires[2] (and references therein).

The presence of autoantibodies in the circulation is a well-known fundamental feature of autoimmune diseases, making them appealing biomarker candidates.[6] In fact, autoantibodies have also been explored as potential biomarkers in several other diseases, including various cancers.[7] As for example, serum autoantibodies against tumor-associated antigens have been explored as diagnostic biomarkers in hepatocellular carcinoma and other solid tumors.[8] In this context, the use of functional protein microarrays for multiplex detection of autoantibodies represents an attractive high-throughput approach. Indeed, antigen microarrays have recently been shown useful for autoantibody profiling in a range of diseases, paving the way for biomarker discovery efforts.[9–13] But despite the success, there is still a great need for additional high-performing protein microarrays that display the arrayed proteins, microarray adapted by molecular design, in a format (conformation) best suitable for recognition of and binding to any autoantibodies present in the sample. To this end, Akada et al. has generated proof-of-concept for a new type of protein chip for multiplex detection of serum autoantibodies.[14]

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