Technology overview

GeneCopoeia’s OmicsArray™ antigen microarrays contain up to 120 purified proteins spotted onto nitrocellulose filters, which are adhered to glass slides. In addition, 8 spots are included for normalization. Each slide carries 16 identical arrays, and so can be used to process up to 15 samples simultaneously as well as a negative control. As little as 1 ul serum or 50 ul of other bio fluids are needed for each sample.

As shown in Figure 1, arrays are incubated with patient samples, and any antibodies in the  samples bind to their cognate antigens on the array. The arrays are washed to remove unbound antibodies and other proteins, then co-incubated with Cy3- and Cy5-labeled secondary antibodies. The dual labeling strategy is intended to distinguish between immunoglobulin (Ig) subtypes present within samples. For example. a Cy3-labeled anti-IgG secondary antibody is used to detect IgG antibodies, and a Cy5-labeled anti-IgM secondary antibody is used to detect IgM antibodies. Fluorophore-labeled secondary antibodies are available for detecting IgA, IgD, IgE, IgG and IgM immunoglubulins, as well as IgG subclasses IgG1, IgG2, IgG3, and IgG4.

After washing to remove unbound secondary antibodies, signals are detected using a microarray scanner (e.g., GenePix® 4000B, InnoScan 710, or equivalents). The raw data is then be analyzed using GenePix® Pro 7.0, or Mapix software.

Data analysis

GeneCopoeia provides data analysis as part of its custom antigen microarray services. After array scanning, raw data are collected and analyzed using GenePix® 7.0 software. The standard service includes data normalization to include fold changes of, for example, disease vs. healthy control, and a heat map showing each antigen ranked by level of fold change. Our service undergoes the following steps:

1. Collection of raw data. After scanning, raw signal intensities are collected in an Excel file, as shown below.

2. Next, the raw data are normalized to controls on the array, and the Net Signal Intensity (NSI) values, as well as the signal-to-noise ratios (SNRs) are tabulated in an Excel file,

3. The final step of the standard analysis package is to determine the “Antibody score” which is the relative enrichment of a given antibody in a sample. The antibody score is determined numerically and displayed in a heat map. A higher score (shifted toward red on the heat map) suggests a stronger antibody-antigen interaction, as shown in the following example:

4. In addition to the standard data analysis package, customers can also choose custom data analysis services. One such service is to classify positive antigens on the array using Gene Ontology (GO) analysis, which classifies genes and proteins based on known biolgical functions. An example of the readout from GO analysis for molecular function is shown below:

Further, GO analysis for biological processes is shown in the following example:

5. Another widely-used tool for bioinformatic analysis is KEGG (Kyoto Enclycopedia for Genes and Genomes) Pathway Analysis, which we use to group positive antibody-antigen interactions on the basis of defined biologica pathways, as displayed in the example below:

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Frequently Asked Questions

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Publications

View publications using GeneCopoeia antigen microarray products and services

Webinars

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Title:Antigen Protein Microarrays For Effective Antibody Profiling.

Presented Wednesday, September 22, 2021

GeneCopoeia’s OmicsArray™ antigen microarrays are protein microarrays engineered for several powerful applications, including autoantibody detection, biomarker profiling, and characterization of protein-molecule interactions. Antigen microarrays enable parallel detection of antibodies or other biomarkers present in patient samples for diseases including rheumatoid arthritis, muscular dystrophy, systemic lupus erythematosus, and type 1 diabetes. In this webinar, we will describe the powerful application and advantages for using GeneCopoeia’s OmicsArray™ antigen microarrays for autoantibody detection, cancer immune checkpoint inhibition autoantibody profiling, and SARS-CoV-2 serological analysis.

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Title: OmicsArray™ Antigen Microarrays for high-throughput Autoantibody Screening for Autoimmune and Immune-associated Disorders.

Dr. Quanzhen Li from GeneCopoeia throws light on OmicsArray™ Antigen Microarrays for high-throughput Autoantibody Screening for Autoimmune and Immune-associated Disorders through an informative webinar hosted by GeneCopoeia.

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