Eliminating the hook effect in allergen testing

This article comes from Neogen’s Allergen Insider newsletter, a resource for the latest developments and news in allergen testing and related technology. See here to read the rest of the latest issue of Allergen Insider.

When testing for allergens with rapid methods, such as lateral flow test kits, there’s always a risk that a grossly contaminated sample will return a false negative result by overloading the test strip. Known as “the hook effect,” this occurs when the amount of target allergen exceeds the amount of color-labeled antibody material present in the strip’s reagent pad.

The mechanics of the hook effect are simple: Excess target allergen migrates across the membrane quicker than the color-labeled antibody-antigen complex and saturates the binding sites on the capture antibody at the test line. When the color-labeled complex arrives, it has no binding sites available, so it continues to travel up the membrane to the waste reservoir at the end of the device. Without binding sites available, the color-labeled antibody-antigen complex cannot create the colored test line indicative of a positive result. This presents the user with a false negative result, despite high levels of the target allergen.

Neogen’s Reveal 3-D lateral flow device includes a third line in addition to the test and control lines used in conventional lateral flow devices (see fig. 1). This overload line is simply a reagent striped onto the device. It doesn’t form in the presence of a highly positive sample. This means that it’s possible to instantly and easily differentiate a high positive result from a negative result — if the overload line doesn’t show up, the sample is overloaded with the target allergen. A negative result will show the overload and control lines, while a positive result will show the test, overload and control lines (see fig. 2).