Development of Assay to detect Silicase activity.

To determine if we could detect the liberation of cations such as calcium and magnesium from silica we first sough to determine if we could simply detect the breakdown of silicon dioxide which would release free silica. We used a commercially available silicic acid assay. To test this assay we incubated an enzyme known to be a silicase with silicon dioxide (sand), we included in our study proper negative controls. What we found was that all of our samples were positive for free silica, simply adding sand to water and shaking was sufficient to yield enough signal to max out the assay. We tried dilutions but were unable to ever detect any differences between our experimental conditions and negative control. We then sought to determine if we could detect the liberation of calcium from calcium containing silicate rocks. We again ran into a similar roadblock. We used the newest version of calcium indicators our lab has generated, gCaMP8, however this sensor is far too sensitive, and the abundance of calcium simply in the environment maxed the sensor out. We are now trying earlier generations of the gCaMP sensor.