Evaluation of commercial α‐amylase enzyme‐linked immunosorbent assay (ELISA) test kits for wheat

Abstract

Background and objectives There are few methods in wheat to directly measure α‐amylase, which is the result of preharvest sprouting (PHS) and late maturity α‐amylase (LMA). Both of these conditions are associated with reduced wheat and flour quality. Those methods that do exist for measuring α‐amylase in wheat are based on enzyme activity. The objective of this study was to evaluate commercially available enzyme‐linked immunosorbent assay (ELISA) kits on sound grain, sprouted grain, and grain that had both PHS and LMA. Findings Three kits were evaluated; human AMY1 and AMY2, and a “plant α‐amylase” kit. The wheat samples were also assayed for α‐amylase activity using the Megazyme SD α‐amylase method. All three ELISA kits functioned very well based on calibration curves (R2 > .99). Of the three, the AMY1 and plant α‐amylase kits appeared to show no useful detection of wheat α‐amylase. The AMY2 kit, however, detected a 2.7‐fold increase in α‐amylase protein in the PHS and PHS/LMA grain lots, similar to a 2.7‐fold difference in activity. Conclusions It appears that the particular human AMY2A ELISA kit, and perhaps others, can detect α‐amylase in wheat and may be a useful alternative technology to enzyme kits. These kits may also be useful in predicting PHS and LMA potential damage. Significance and novelty Thus far, α‐amylase studies have focused on enzyme kits, whereas this study examined three commercially available ELISA kits to measure α‐amylase content in sound, sprouted, and PHS/LMA grain. Both PHS and LMA are highly detrimental to grain and result in low prices paid to growers.

Breeding, Selection, and Quality Characteristics of Soft White Wheat

“…Most of the testing protocols follow AACC Approved Methods (1). The WWQL has been involved in evaluating, modifying, and establishing methods for measuring soft white wheat quality, including gluten strength, enzymatic discoloration, solvent retention capacities (SRCs), arabinoxylans, and enzyme assays (2,5,9,13,20,21,23,33,38,39,41).…”

“…Primarily a function of its uniformly weak gluten, which is largely independent of protein content (9), it has the ability to lessen any adverse effect that gluten structure might have on product texture. These subtle but measurable differences between club and common soft white wheats are maintained through breeding efforts and phenotypic and genetic selection (20,21).…”