3′,3′-Cyclic GAMP ELISA Kits

Research Areas

Documentation

Specifications

• Assay Type Competitive ELISA
• Sample Types Cell Lysates, Tissue Extracts, Tissue Culture Media
• Sensitivity 0.128 pmol/mL
• Species 3',3'-Cyclic GAMP is identical across species
• Assay Duration 1.5 Hours
• Samples/Plate 39 in Duplicate
• Readout Colorimetric, 450 nm
• Standard Curve
• Description

  Assay Principle: 

  The 3′,3′-Cyclic GAMP ELISA Kit quantitatively measures 3’,3’-cGAMP present in cell lysates, tissue extracts, and tissue culture media. The 3′,3′-Cyclic GAMP ELISA Kit is a competitive ELISA. Please read the complete kit insert before performing this assay. 

  Use our provided 3’,3’-Cyclic GAMP (cGAMP) standard to generate a standard curve for the assay. Pipette the standards or diluted samples into a transparent microtiter plate coated with our goat anti-rabbit IgG antibody. Add the 3’,3’-cGAMP-peroxidase conjugate and the 3’,3’-cGAMP polyclonal rabbit antibody. Then incubate the mixture for 1 hour, shaking at room temperature. The immunological reaction occurs between the anti-3’,3’-cGAMP polyclonal antibody, the 3’,3’-cGAMP antigen in the sample or standard, and the 3’,3’-cGAMP-peroxidase conjugate. As the 3’,3’-cGAMP concentration in the sample increases, the bound 3’,3’-cGAMP-peroxidase conjugate decreases, causing a decrease in signal and vice versa. 

  After the 1 hour incubation, wash away the excess 3’,3’-cGAMP-peroxidase and add the TMB substrate. The TMB substrate reacts with the bound 3’,3’-cGAMP-peroxidase conjugate generating a signal detected by a plate reader at 450nm. Use the intensity and the standard curve to calculate the 3’,3’-cGAMP concentration in the samples. 

  Background:

  3’,3’-Cyclic guanosine monophosphate–adenosine monophosphate (2’,3’-cGAMP)is a mammalian messenger molecule. It is a crucial mediator of bacterial signal transduction and regulation, controlling a range of diverse targets, including transcription, enzyme activity, and cell cycle progression. In bacteria, gene-regulatory RNA elements called riboswitches bind and respond to cGAMP with high affinity and specificity, regulating 3’,3’-cGAMP signaling. The riboswitches regulate genes involved in motility, biofilm formation, colonization, and virulence. The cyclic nucleotides have emerged as critical players in bacterial physiology, and inhibition studies of cGAMP signaling are ongoing as an anti-microbial strategy.

  In mammalian cells, 3’,3’-cGAMP and its eukaryotic analog 2’,3’-cGAMP produced by cGAS, bind STING (stimulator of IFN genes). This induces TBK1-IRF3-dependent production of IFN-β. Here, the cGAS-cGAMP-STING DNA sensing pathway is a crucial activator of the innate immune response to foreign or harmful native DNA. The cGAS-cGAMP-STING system is critical in antiviral and antitumor immunity, mediating autoimmune responses. Dysregulation or aberrant activation of the pathway by self-DNA has emerged as an underlying cause of tumorigenesis and autoimmune disorders.