for a good recovery, whereas no effect was observed from protease inhibitors. No detectable loss was observed in the lyophilized product. In a subsequent experiment, the stability of alb-OC was analyzed under optimal conditions for prolonged periods of time. Storage conditions for the diluted product were as follows: alb-OC concentration, 375 mg/L in PBS containing 4% (w/v) ovalbumin; 0.1% (w/v) kathon; sealed plastic tubes. The lyophilized product was prepared from the same solution and was analyzed for up to 4 weeks at 37ยฐC. The data are given in Fig. 2 and demonstrate that at 4ยฐC alb-OC was stable for at least 1 month. At higher temperatures, substantial losses were observed after a few days. The lyophilized product was stable at 37ยฐC for at least 1 month.
In this paper, we demonstrate that peptides linked to a carrier protein may be used as a calibrator in enzyme assays for biomarkers. The method was validated with the bone Gla-protein osteocalcin, but may be used in a wide variety of comparable applications. In the case of alb-OC, the chimeric molecule mimicked the immunochemical properties of osteocalcin but was more than 10 times larger than the native molecule. The preparation procedure for albumin-linked peptides is simple and reproducible, and may be further improved by more rigid standardization or automated procedures. The stability of lyophilized material at temperatures up to 37ยฐC and that of soluble material at 4ยฐC were satisfactory; at higher temperatures, soluble alb-OC degraded with a biphasic curve, leading to a loss of approximately 50% after 4 weeks. If required, the stability may be improved, e.g., by sterile filtration of the solvent or by adding protease inhibitor cocktails (3). We conclude that albumin-linked peptides form a good alternative in cases in which the authentic protein cannot be used as a reference material in test kits, for instance because it is expensive, difficult to obtain, unstable, poorly soluble in saline, or associated with infection risks.