Product Information
Myasthenia Gravis (MG) is a chronic autoimmune neuromuscular disorder in which autoantibodies destroy the communication between nerves and muscle, resulting in weakness of the skeletal muscles. Myasthenia gravis affects the voluntary muscles of the body, especially those that control the eyes, mouth, throat, and limbs. MG is most often caused by antibodies to the acetylcholine receptors, but antibodies to other proteins, such as MuSK (Muscle-Specific Kinase), LRP4 (Lipoprotein-receptor-related protein 4), RYR (Ryanodine receptor), also can impair transmission at the neuromuscular junction.
GeneCopoeia’s OmicsArray™ Myasthenia gravis (MG) antigen array carries 68 MG-specific and/or -associate antigens. The 68-Plex MG Antigen Array is by far the largest collection of antigens for MG study and provides a powerful tool for highly sensitive profiling of autoantibodies in myasthenia gravis and associated diseases.
The 68 antigens included in the MG panel were chosen based on a thorough literature search. The superior-quality purified antigens were coated onto the 3D surface of microchips to ensure that antigens retain their natural conformations. Internal Ig and anti-Ig controls are included in each array for normalization and quantification purposes. Additionally, the utilization of fluorescent 2nd antibodies in our microarrays allow for the detection of different types and sub-types of antibodies (IgG1, IgG2, IgG3, IgG4, IgM, IgA) per experiment, minimizing the requirement of sample needed.
In addition to this premade array, arrays containing customized sets of antigens are available, as well as array profiling services and data analysis. To order premade or custom arrays, please contact us.
GeneCopoeia’s OmicsArray™ MG 68-plex antigen array is part of the GeneCopoeia OmicsArray™ Antigen Microarray family.
References
- Leite MI, Waters P, Vincent A. Diagnostic use of autoantibodies in myasthenia gravis. Autoimmunity. 2010;43(5-6):371-9.
- Meriggioli MN, Sanders DB. Muscle autoantibodies in myasthenia gravis: beyond diagnosis? Expert Rev Clin Immunol. 2012;8(5):427-38.
- Evoli A, Alboini PE, Damato V, et al. Myasthenia gravis with antibodies to MuSK: an update. Ann N Y Acad Sci. 2018 Jan;1412(1):82-89.
- Yan M, Xing GL, Xiong WC, Mei L. Agrin and LRP4 antibodies as new biomarkers of myasthenia gravis. Ann N Y Acad Sci. 2018;1413(1):126-135.
- Zhang J, Chen Y, Chen J, et al. AChRAb and MuSKAb double-seropositive myasthenia gravis: a distinct subtype? Neurol Sci. 2021;42(3):863-869.
- Gilhus NE, Skeie GO, Romi F, et al. Myasthenia gravis – autoantibody characteristics and their implications for therapy. Nat Rev Neurol. 2016;12(5):259-68.
- Lazaridis K, Tzartos SJ. Autoantibody Specificities in Myasthenia Gravis; Implications for Improved Diagnostics and Therapeutics. Front Immunol. 2020;11:212.
- Mantegazza R, Bernasconi P, Cavalcante P. Myasthenia gravis: from autoantibodies to therapy. Curr Opin Neurol. 2018;31(5):517-525.
The listed chip prices apply to academic customers. For custom arrays and services pricing, please contact us.
Custom Services
GeneCopoeia offers custom antigen microarray services in the following areas:
- Custom array printing. GeneCopoeia will create custom antigen microarrays built to your specifications.
- Sample processing. Send us your blood, plasma, tissue, or other biological sample and we will prepare it for processing and incubation with any of our premade antigen microarrays or custom-built antigen microarrays for autoantibody profiling and other applications. For information on sample types to submit, consult the FAQ
- Data analysis. Once samples are processed and incubated with an antigen microarray, we will analyze the raw data. The standard analysis service includes: 1) An Excel file of the Net Signal Intensity (NSI) for each antigen on the array, normalized to internal controls; and 2) a heat map
Additional analysis services, including proteomic analysis, pathway analysis, and more, are also available.
To inquire about custom antigen microarray products and services, please fill out our custom quote request form.
Technology overview
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.
Figure 1. Workflow for detection of antibodies in samples using GeneCopoeia’s OmicsArray™ antigen microarrays.
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:
FAQ
Frequently Asked Questions
Answer: You can send us virtually any biological sample, including whole blood, serum, plasma, interstitial fluid, semen, urine, and saliva. To learn more, please contact us at
inquiry@genecopoeia.com.
Answer: Yes.
Answer: Yes.
Answer: Yes.
Answer: Each nitrocellulose membrane is surrounded by a gasket to prevent leakage and cross-contamination.
Answer: a. Each slide carries 16 identical arrays. One of these is reserved for a PBS control, while the remaining 15 are used for incubation of each sample. Therefore, the total number of samples sent by the customer is preferably a multiple of 15. However, if you have fewer than 15 samples, we will need to charge you for the full cost of an entire slide of 15 samples. b. Theoretically, each of the different test groups requires a minimum of 3 samples to meet statistical reproducibility requirements. On this basis, the greater the number of samples, the higher the reproducibility of the results in the test group. In order to ensure the reliability of the results, the sample size of different test groups is recommended to be ≥3.
Answer: The antigen microarray uses fluorescence detection, and its sensitivity is higher and more stable than ELISA (colorimetry) and Western (chemiluminescence). The technical repetition correlation coefficient R2 between the arrays can reach 0.9 or more. The dynamic detection range is 1-65000. The larger the dynamic range, the more layers of signals that can be detected.
Answer: Please fully communicate with our Technical Support staff before the experiment to explain the purpose of the experiment and the sample status, to determine whether we can meet your needs and determine the experimental plan.
Answer: The antigens on the antigen array are all derived from self-antigens reported in the literature. For each of the different types of antigenic arrays, we can provide you with all the names of the antigens upon reques Please do not disclose this information to third parties.
Answer: Proteins are usually expressed and purified from E. coli or mammalian cells.
Answer: We usually spot the entire full-length protein in its native conformation. However, we can also spot truncated proteins or peptides if needed.
Answer: If you need a custom antigen microarray, we need to fully communicate with you about the following issues: a. The intellectual property of the antigen (whether from a published paper or a patent, etc.). We can only provide customized detection of antigens that do not involve patent protection; b. Basic information about the antigen (whether it is whole protein or peptide, molecular weight, domain, whether it is a membrane protein, etc.); c. Whether you can provide an antigen that meets the requirements of a custom array; d. If we need to provide the antigen to be tested, the corresponding cost and experimental time frame need to be accounted for separately.
Answer: The antigens on our predesigned arrays are usually human. However, for custom arrays, we can use antigens from virtually any organism.In addition, the arrays have been used with human, mouse, and rat samples.
Answer: Antigens are usually spotted at a concentration between 0.1 and 1.0 mg/ml, but each individual protein concentration requires optimization.
Answer: a. This difference may be related to the detection method. The protein used on the array is a non-denatured protein, which is different from western blot. The hybrid system of the chip is an antigen-trapping antibody, and the western blot antigen is slightly different from the antibody after interacting with the antigen; b. Please provide a graph of the results of western blot verification, so that we can better analyze the image and data of your verified indicators on the array.
