Structural identification of N-linked carbohydrates using the GUcal application: A tutorial

In recent years, analytical glycomics gained a significant role due to the rapidly increasing number of glycoproteins introduced as biopharmaceuticals. One of the frequently used methods for the analysis of complex carbohydrates is capillary electrophoresis with laser induced fluorescent detection (CE-LIF). CE-LIF is a high resolution separation technique with excellent sensitivity capable of discriminating between closely related positional and linkage carbohydrate isomers. Individual glycan structures corresponding to the separated peaks in an electropherogram are identified according to their glucose unit (GU) values by mining the built in database. This tutorial introduces the practical use of the GUcal application, a recently developed glycoinformatics tool, which automatically calculates GU values for all sample components of interest in an electropherogram using either the bracketing standard approach or the recently published triple internal standard method. Furthermore, a worked example demonstrates the way glycan structural elucidation of human immunoglobulin G is processed with the help of this simple and rapid GU value calculation application.Biopharmaceuticals have seen something of tremendous development in recent years, which governs the parallel blooming of analytical glycomics. One of the frequently used methods for the analysis of complex carbohydrates is capillary electrophoresis with laser induced fluorescent detection (CE-LIF). CE-LIF is a high resolution separation technique with excellent sensitivity capable of discriminating between closely related positional and linkage carbohydrate isomers. While CE instrumentation is well developed, the related bioinformatics tools are lagging behind. According to our best knowledge, this is the first tutorial paper on the recently disseminated GUcal application, which automatically calculates GU values for all sample components of interest in an electropherogram using either the bracketing standard approach or the new triple internal standard method. On the top the step-by-step instruction how to use the application, the paper includes a worked example demonstrates the way glycan structure elucidation of human immunoglobulin G is processed with the help of the simple and rapid GU value calculation of this new application. After very short training period, the software can be used readily for cutting-edge glycomics of exploratory research or high-throughput routine analysis.