We show you how to detect and analyze acetylcholine in brain microdialysis samples in 18 minutes, with minimal or no need to add an acetylcholine esterase inhibitor or AEI.
For this analysis, we use high-performance liquid chromatography or HPLC, coupled with an immobilized enzymatic reactor column and an electrochemical detector. It’s fast and highly sensitive, down to the femtomole range.
Acetylcholine is separated by a polymer-based reverse phase column, which has been selected specifically for this application. Not only is the column we’ve selected great at separating acetylcholine, but it can also handle the higher pH required for optimal enzyme activity. After exiting the separation column, acetylcholine and other compounds enter an immobilized-enzyme reactor column. Inside the reactor column, compounds are broken down to produce hydrogen peroxide, which is then selectively oxidized by an applied voltage across the flow path and changes to current are detected with a platinum electrode.
Optimizing the right conditions can take time and resources. That’s why many labs require highly-trained specialists in HPLC. However, we’ve created an application, which includes everything you need to run the acetylcholine analysis right away.
For this application, we’re going to use the HTEC, which includes everything you need integrated into one single unit. Let’s take a look inside:
-Dual-piston pump with a unique algorithm to reduce noise without any pulse damper.
-Degasser to remove small air bubbles for better pump performance
-Temperature control for consistent results
-Separation column uniquely selected for acetylcholine
-Enzymatic reactor column
-Electrochemical detector cell with a platinum working electrode for selective detection of the enzymatic reaction products.
We’ve optimized conditions to make it as easy as possible. Simply inject the sample using a manual injector or autosampler if you’re using laboratory automation. You can monitor signal response in real-time using the dedicated chromatography software. In 18 minutes you will have your results and you’re ready for the next sample. It’s highly sensitive down to the femtomole range. Here’s an example of a chromatogram showing acetylcholine.
For more information on acetylcholine analysis, contact us today!