Abstract 4625: A biotin-label-based antibody array for high-content profiling of protein expression

Abstract

Profiling protein expression on a global scale will have significant impact on biomedical research, particularly in the discovery and development of drugs and biomarkers. Through the years, several antibody array systems have been invented and developed for multiple protein detection. However, a reliable and high-content system for protein profiling from many biological samples has yet developed. To address this problem, we have developed a high density antibody array and used this technology to uncover the potential biomarkers of ovarian cancer. In this approach, biological samples are conjugated with biotin. Antibody arrays are then used to capture the biotinylated proteins. The presence of proteins captured by the antibody chip is detected using streptavidin-conjugated fluorescent dye (Cy3 equivalent) as a reporter. The signals, which are visualized by laser scanning, are normalized using positive, negative, and internal controls. Using this biotin-label-based antibody array technology, the expression levels of 507 human, 308 mouse, 90 rat target proteins can be simultaneously detected, including cytokines, chemokines, adipokines, growth factors, angiogenic factors, proteases, soluble receptors, soluble adhesion molecules, and other proteins in a variety of samples. Most proteins can be detected at pg/ml and ng/ml levels with a CV of less than 20%. Using human biotin-based antibody arrays, we screened the serum expression profiles of 507 proteins in 47 ovarian cancer patients and 39 normal subjects. A panel of protein expression showed significant difference between normal and cancer (P<0.05). By Classification Analysis and Split-Point Score Analysis of these two groups, a small group of proteins can be used to distinguish ovarian cancer patients from normal subjects and between ovarian cancer patients and normal subjects. Our results suggest the biotin label-based antibody arrays that we have developed have great potential in applications for biomarker discovery. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4625.

Cytokine Antibody Arrays in Biomarker Discovery and Validation

“…Because the cross-reactivity is eliminated in the label-based format, array densities can be scaled up to a theoretically unlimited size. Both the label-based method [33] and the sandwich-based method [34] have proven valuable in the discovery of disease biomarkers.…”

“…A panel of protein expression showed significant difference between normal and cancer (P<0.05). By classification analysis and split-point score analysis of these two groups, a six-marker panel of proteins (IL-2R , endothelin, osteoprotegerin, vascular endothelial growth factor D (VEGF-D) and betacellulin (BTC)) could be used to distinguish ovarian cancer patients from normal subjects [33]. Those studies strongly suggest that antibody array technology has great promise in the discovery and development of ovarian cancer biomarkers.…”

Identification of Five Serum Protein Markers for Detection of Ovarian Cancer by Antibody Arrays

“…Multiple studies have been reported to identify serum ovarian cancer biomarkers using multiplex antibody array technology [7-9]. Dr. Lokshin’s group identified a group of 6 serum protein markers, including interleukin-6 (IL-6), interleukin-8 (IL-8), epidermal growth factor (EGF), vascular endothelial growth factor (VEGF), monocyte chemoattractant protein-1 (MCP-1), and CA-125, which displayed significant difference in serum concentrations between ovarian cancer and control groups with 84% sensitivity at 95% specificity [7].…”

“…Using human biotin-based antibody arrays, we screened the serum expression profiles of 507 proteins in serum samples from 47 patients with ovarian cancers, 33 patients with benign ovarian masses and 39 healthy, age-matched controls and identified significant differences in protein expression between normal controls and patients with ovarian cancer ( P <0.05). By classification analysis and split-point score analysis of these 2 groups, a 6-marker panel of proteins, which consisted of interleukin-2 receptor alpha (IL2Rα), endothelin, osteoprotegerin (OPG), vascular endothelial growth factor D (VEGF-D) and betacellulin (BTC), can be used to distinguish ovarian cancer patients from normal subjects [9]. These studies strongly suggest that antibody array technology has shown great promise in the discovery and development of serum ovarian cancer biomarker profiles and strongly suggest that serum cytokine panels may be useful as biomarkers for early detection of ovarian cancers.…”

Unifying the integration, analysis and interpretation of multi-omic datasets: Exploration of the disease networks of Obstructive Nephropathy in children

“…Thus, nowadays microarrays are used to measure events at the post-transcriptional level of gene regulation (e.g. miRNA expression [1]), at the post-translational level by measuring active protein levels (antibody arrays [2]) or I. Valavanis is with the Institute of Biological Research and Biotechnology, National Hellenic Research Foundation, Athens, Greece (email: ivalavan@eie.gr).…”