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 IROA Phenotypic Quantitation Kit

There are many cases where it is not practical or possible to label the experimental sample, for example, when working with tissue samples, performing large fermentation runs, or propagating field-grown plants.  In these instances, the IROA Phenotypic protocol may be applied. In the Phenotypic IROA protocol (Figure 3-3), the control or “Internal Standard” is isotopically labeled using U-13C 95% IROA media. 
 
An ideal Internal Standard is one that represents the entire metabolome of the fluid or tissue under study. For example, if interrogating the metabolome in liver tissue, quantification of metabolites can be achieved using an IROA-grown liver cell line as a complex Internal Standard. For Phenotypic analyses, the ClusterFinder software will find all IROA-labeled compounds where the IROA peak is present in one sample but not the other. This allows for a complex targeted analysis whereby the 13C IROA peaks in the labeled Control sample/Internal Standard are used to identify their associated natural abundance peaks in the experimental sample.