Total protein concentration and stability of Amb a 1 in glycerinated ragweed sublingual immunotherapy stored at room temperature and refrigerated cold temperature.

Abstract

Background: Few studies have investigated optimal storage conditions or expiration dates for sublingual immunotherapy (SLIT) formulations prepared from glycerinated allergen extracts.

Hypothesis/objectives: The objective of this study was to compare concentrations of short ragweed major allergen (Amb a 1) and total protein in SLIT formulations stored at two different temperatures. It was hypothesised that protein concentrations would show greater decline over time in a formulation stored at room temperature (RT) than in one stored under refrigeration.

Materials and methods: Two SLIT samples containing equal volumes of 20,000 PNU Amb a 1 extract were prepared and stored at refrigerated cold (CT) (2-8°C) or RT (20-24°C) for 140 days. Changes in total protein and major allergen concentration and composition were measured by Bradford assay, two-site enzyme-linked immunosorbent assay and SDS-PAGE. Presence of Amb a 1 was confirmed with Western immunoblot. Data were analysed using an analysis of covariance, with p < 0.05 considered significant.

Results: SDS-PAGE showed compositional changes in a ~26-30 kDa protein band under RT and not CT storage. The Amb a 1 concentration of the RT SLIT sample declined significantly over time, compared to that of the CT SLIT sample (F_(1,8) = 47.69, p < 0.0001). There was no significant difference in total protein concentration over time between groups (F_(1,8) = 1.79, p = 0.22).

Conclusions and clinical relevance: These results demonstrate that storage of glycerinated SLIT formulations in refrigerated CT preserved the highest concentration of the specific allergen Amb a 1, suggesting that SLIT formulations containing short ragweed should be stored under refrigeration.