Sample pre-treatment determines the clinical usefulness of acid-labile subunit immunoassays in the diagnosis of growth hormone deficiency and acromegaly

Abstract

Objective: The usefulness of measuring the GH-dependent acid-labile subunit (ALS) in the management of GH deficiency (GHD) and acromegaly remains in question and is investigated in this study, comparing several different immunoassays for ALS. Method: We compared the diagnostic accuracy of a commercially available polyclonal Ab-based ELISA with SDS pre-treatment (SDS-ELISA) with a monoclonal Ab-based immunofluorometric assay, using two unfolding methods (urea (UREA) and Glycine-HCl (Gly)). The corresponding molecular weight (MW) of ALS and IGFBP-3 immunoreactivity was determined. The clinical usefulness of each assay was examined in adult GH disorders. Results: ALS was lower in GHD and higher in acromegaly using all assays. In GHD, UREA had higher sensitivity and specificity than SDS-ELISA (59 and 69% versus 41 and 51% respectively). In acromegaly, sensitivity and specificity was 94 and 87% for UREA, 81 and 36% for Gly, and 44 and 44% for SDS-ELISA. After UREA, immunoreactivity for ALS and IGFBP-3 eluted at their predicted free MW using size-exclusion chromatography, whereas ALS immunoreactivity in SDS (300-600 kDa) and Gly (250-500 kDa) was at a high apparent MW consistent with aggregation. Conclusion: The diagnostic accuracy of ALS varies with assay choice and pre-treatment modality. UREA, which results in migration of ALS at the expected MW on a sizing column, has the highest specificity and sensitivity. Thus, if measured in an assay in which ALS is unfolded without aggregation, ALS is a clinically highly useful parameter for the assessment of GH.

Endogenous estradiol may influence IGF-I levels in acromegalic women treated with pegvisomant

“…To optimize immunorecognition, samples were pre-treated with 3 M urea and 0.05% SDS [13]. A serum pool taken from healthy male volunteers was used for calibration and assigned 1,000 U/l.…”

IGF-1, IGFBP-3 and ALS in adult patients with chronic kidney disease

“…For measurement of acid-labile subunit (ALS) levels pre-treatment of sample sera was performed as previously described [22].…”

Systemic response of the GH/IGF-I axis in timely versus delayed fracture healing

“…Serum ALS levels (mU/l) were measured in duplicate by sandwich immunometric assay using monoclonal antibodies directed against specific N-and C-terminal oligopeptides [16,17]. To optimize immunorecognition, samples were pre-treated with 3 M urea and 0.05% SDS.…”

“…Values for IGF-I and IGBP-3 are based on age matched data from 797 female and 787 male apparently healthy volunteers which have been detected by the assays used in our study [18]. Reference ranges for ALS were adapted from those published for a commercially available assay system [19] after extensive comparative analysis of samples by the inhouse assay and the commercial assay system [17].…”