Measurement of Cyclic Guanosine Monophosphate (cGMP) in Solid Tissues using Competitive Enzyme-Linked Immunosorbent...

A major pathway that produces cyclic guanosine monophosphate (cGMP) is the nitric oxide (NO)-cGMP pathway. In this pathway, NO stimulates the catalytic activity of soluble guanylyl cyclase (sGC), which converts guanosine triphosphate to cGMP. NO is of interest because, in mammals, it acts as the main vasodilator, an inhibitor of platelet aggregation, a mediator of immune function, a neurotransmitter, and to improve endurance during physical activity, among other physiological roles. Measuring NO's abundance in different tissues could help elucidate its functions. However, NO cannot be measured directly due to its less than seconds-long half-life. Typically, NO's metabolites, nitrate and nitrite, are measured to estimate NO's abundance. This is because these molecules can be interconverted in vivo through redox reactions. A less common method of estimating NO's abundance requires measuring the concentration of cGMP, a signaling molecule downstream of NO. Yet, measuring cGMP in tissues is seldom done, despite creating the possibility to better understand NO's functions in organs. Therefore, a need exists for a protocol to measure cGMP concentrations in solid organs and tissues. In this protocol, collected pig tissues were homogenized, competitive ELISA was performed on processed samples, and absorbances were converted into nanomoles of cGMP per gram of original tissue. In short, this protocol delineates a simplistic method to measure cGMP concentrations in solid tissues using the cyclic GMP ELISA kit.