PDF《(G-Series)》

Patent Pending Technology User Manual (Revised June 6, 2014) RayBio? Human Cytokine Antibody Array G-Series

3 Cat# AAH-CYT-G3-4 RayBio? Human Cytokine Antibody Array G-Series

3 Cat# AAH-CYT-G3-8 RayBio? Human Cytokine Antibody Array G-Series Testing Service Cat# AAG-SERV-G Please read manual carefully before starting experiment We provide you with excellent Protein Array systems and services Tel: (Toll Free) 1-888-494-8555 or +1-770-729-2992;

Fax: +1-770-206-2393;

Website: www.

raybiotech.com Email: [email protected] RayBio? Human Cytokine Antibody Array

3 (G-Series) RayBiotech, Inc., the Protein Array Pioneer Company, strives to research and develop new products to meet demands of the biomedical community. RayBiotech'

s patent-pending technology allows detection of up to 1,000 cytokines, chemokines and other proteins in a single experiment. Our format is simple, sensitive, reliable, reproducible and cost-effective. Our product offerings include: 1. Protein (antigen) Arrays 2. RayBio ? Cytokine Antibody Arrays o C Series (Membrane, chemiluminescence detection) o G-Series (Glass chip, fluorescence detection) 3. Pathway- and disease-focused antibody arrays o Angiogenesis Antibody Arrays o Apoptosis Antibody Arrays o Atherosclerosis Antibody Arrays o Chemokine Antibody Arrays o Growth Factor Antibody Arrays o Inflammation Antibody Arrays o MMP Antibody Arrays o Obesity Antibody Arrays 4. Quantibody? Multiplex ELISA Arrays 5. RayBio ? L-Series Biotin Label-based Antibody Arrays 6. RayBio ? E-Series Competition-based Antibody Arrays 7. RayBio ? Phosphorylation Antibody Arrays o Receptor Tyrosine Kinases o EGFR and ErbB family (site-specific phosphorylation) 8. Over 1,300 different ELISA kits 9. EIA (Competitive ELISA) kits 10. Cell-based Phosphorylation Assay 11. Over 20,000 different antibodies 12. Recombinant proteins 13. Peptide 14. Recombinant antibodies

1 RayBio? Human Cytokine Antibody Array G-Series

3 Protocol for RayBio? Human Cytokine Antibody Array G-Series

3 TABLE OF CONTENTS I. Introduction・1 II. Product Information・5 A. Storage Recommendations・5 B. RayBio? G-Series Glass Chip Layout

6 C. Materials Provided

6 D. Additional Materials Required・6 E. How It Works

7 III. Helpful Tips and General Considerations・7 A. Preparation and Storage of Samples・7 B. Handling Glass Chips・9 C. Incubations and Washes・9 D. Data Extraction Tips・9 IV. Protocol・10 A. Preparation and Storage of Reagents・10 B. Blocking and Incubations・11 C. Fluorescence Detection・14 V. Interpretation of Results・15 A. Explanation of Control Spots・15 B. Typical Results using G-Series Arrays・16 C. Background Subtraction・17 D. Normalization of Array Data・17 E. Threshold of Significance・18 VI. Antibody Array Map・19 VII. Troubleshooting Guide・20 VIII. Selected References・22 RayBio? Cytokine Antibody Arrays are patent-pending technology. RayBio? is the trademark of RayBiotech, Inc.

2 RayBio? Human Cytokine Antibody Array G-Series

3 I. Introduction New techniques such as cDNA microarrays have enabled us to analyze global gene expression1-3 . However, almost all cell functions are executed by proteins, which cannot be studied simply through DNA and RNA techniques. Experimental analysis clearly shows disparity can exist between the relative expression levels of mRNA and their corresponding proteins4 . Therefore, analysis of the proteomic profile is critical. The conventional approach to analyzing multiple protein expression levels has been to use 2-D SDS-PAGE coupled with mass spectrometry5,6 . However, these methods are slow, expensive, labor- intensive and require specialized equipment7 . Thus, effective study of multiple protein expression levels can be complicated, costly and time-consuming. Moreover, these traditional methods of proteomics are not sensitive enough to detect most cytokines (typically at pg/ml concentrations). Cytokines, broadly defined as secreted cellCcell signaling proteins distinct from classic hormones or neurotransmitters, play important roles in inflammation, innate immunity, apoptosis, angiogenesis, cell growth and differentiation7 . They are involved in most diseases, including cancer, obesity and inflammatory and cardiac diseases. Simultaneous detection of multiple cytokines undoubtedly provides a powerful tool to study cytokines. Regulation of cellular processes by cytokines is a complex, dynamic process, often involving multiple proteins. Positive and negative feedback loops, pleiotropic effects and redundant functions, spatial and temporal expression of or synergistic interactions between multiple cytokines, even regulation via release of soluble forms of membrane-bound receptors, all are common mechanisms modulating the effects of cytokine signaling8-14 . As such, unraveling the role of individual cytokines in physiologic or pathologic processes generally requires consideration and detection of multiple cytokines rather than of a single cytokine.