Development of a protein microarray for profiling circulating autoantibodies in human diseases

Abstract

Purpose To develop a robust microarray platform to detect thousands of serological autoantibodies (AAbs) simultaneously in different diseases. Experimental Design An AAbMap microarray was prepared by printing a total of 4032 purified His‐tagged human proteins and peptide probes on a chemically‐modified slide. The sensitivity, dynamic range, and the inter‐ and intra‐array reproducibility of the AAb microarray were then systematically tested and optimized. Finally, the large‐scale profiling of AAbs in the serum of patients with different human diseases using the AAbMap microarray was demonstrated. Results The dynamic range of antibody (Ab) detection was 2 to 3 orders of magnitude with the lowest limit of detection (LOD) of 68 pg/mL. The intra‐array (r) correlation of duplicate spots was 1.00, whereas the inter‐array correlations between different arrays and batches were 0.99 and 0.97 to 0.98, respectively. Notably, 132, 266, 171, and 84 AAbs were detected in pooled serum from healthy controls (HCs) or patients with rheumatoid arthritis (RA), systemic lupus erythematosus (SLE), or lung cancer (LC), respectively. These AAbs included antibodies that target well‐known disease biomarkers, such as anti‐cyclic citrullinated peptide, anti‐ribonucleoprotein, and anti‐nucleosome. Conclusions and Clinical Relevance We developed a microarray platform to measure thousands of serological AAbs simultaneously with high sensitivity and reproducibility. The array can help study autoimmunity and complement genomics, proteomics, and metabolomics data for systematic investigations of human diseases.

Advances in plasma proteomics: Moving from technology to precision medicine

“…Unique autoantibody profiles were identified for the healthy control, systemic lupus erythematosus, rheumatoid arthritis and lung cancer. [12] Using protein microarray, Banerjee, et al profiled the expression of autoantibodies in the serum of four healthy controls, four Acromegaly, three Cushing's and three Nonfunctional Pituitary Adenomas (NFPAs) patients. The results identified autoantibodies to five proteins in Acromegaly, five proteins in Cushing's patients, two proteins in NFPA patients.…”

“…Unique autoantibody profiles were identified for the healthy control, systemic lupus erythematosus, rheumatoid arthritis and lung cancer. [ 12 ]…”

A protein microarray‐based serum proteomic investigation reveals distinct autoantibody signature in colorectal cancer

“…The study was performed using the Human Proteome Array (HuProt™) to identify tumor-associated autoantibody signatures in colorectal cancer [19,20]. The arrays contain one of the largest collections of full-length human proteins expressed in the eukaryotic expression model yeast and hence provide a unique platform for identifying and detecting a huge number of autoantibodies generated in response to cancer-associated antigens [21].…”