Sample non-linearity can be defined as a sample that when measured at different dilutions gives a substantially different result (Section 7.3). Sample non-linearity should not be confused with assay non-linearity; if an assay is non-linear all samples will give substantially different results when measured at different dilutions. Assessment of Hevylite assay linearity forms an important part of immunoassay development and validation (Sections 9.5 and 9.6).

A linearity study by Eckold et al. [895] reported that IgA, IgG and IgM Hevylite assays are linear over a wide range of sample dilutions and are suitable for routine application. However, as with all immunoassays, Hevylite assays may be potentially prone to occasional sample non-linearity. Sample-specific non-linearity may occur in some samples when measured with Hevylite assays due to: 1) non-specific interference (matrix effects); or 2) the inherent structural variability of monoclonal immunoglobulins (Chapter 3).

11.3.1. Managing non-linearity

Binding Site recommend following the sample dilution protocol as shown in each product insert and reporting the first plausible result only. The use of non-standard sample dilutions, or skipping dilutions should be avoided. Examples of non-linear samples are shown in Table 11.7.

Dilution IgAκ HLC (g/L) IgAλ HLC (g/L)
1/100 >11.3 >10.4
1/400 21.48.0
1/2000 19.1 11.2

Table 11.7. Examples of non-linear IgA Hevylite results using a Siemens BNII analyser. The first plausible result should be reported (in bold).