title: Investigating effects of sample pretreatment on protein stability using size-exclusion chromatography and high-resolution continuum source atomic absorption spectrometry.
abstract: In this study, size-exclusion chromatography and high-resolution atomic absorption spectrometry methods have been developed and evaluated to test the stability of proteins during sample pretreatment. This especially includes different storage conditions but also adsorption before or even during the chromatographic process. For the development of the size exclusion method, a Biosep S3000 5 μm column was used for investigating a series of representative model proteins, namely bovine serum albumin, ovalbumin, monoclonal immunoglobulin G antibody, and myoglobin. Ambient temperature storage was found to be harmful to all model proteins, whereas short-term storage up to 14 days could be done in an ordinary refrigerator. Freezing the protein solutions was always complicated and had to be evaluated for each protein in the corresponding solvent. To keep the proteins in their native state a gentle freezing temperature should be chosen, hence liquid nitrogen should be avoided. Furthermore, a high-resolution continuum source atomic absorption spectrometry method was developed to observe the adsorption of proteins on container material and chromatographic columns. Adsorption to any container led to a sample loss and lowered the recovery rates. During the pretreatment and high-performance size-exclusion chromatography, adsorption caused sample losses of up to 33%.
authors: Institute of Medicinal and Pharmaceutical Chemistry, TU Braunschweig, Braunschweig, Germany.
authors: Tobias
authors: T
authors: Rakow
authors: Sami
authors: S
authors: El Deeb
authors: Thomas
authors: T
authors: Hahne
authors: Deia Abd
authors: DA
authors: El-Hady
authors: Hassan M
authors: HM
authors: AlBishri
authors: Hermann
authors: H
authors: Wätzig
mesh_terms: D000327:Adsorption; D053000:Analytic Sample Preparation Methods; D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry; D002417:Cattle; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D007074:Immunoglobulin G / Q000737:chemistry; D009211:Myoglobin / Q000737:chemistry; D010047:Ovalbumin / Q000737:chemistry; D010316:Particle Size; D055550:Protein Stability; D011506:Proteins / Q000032:analysis / Q000737:chemistry*; D012710:Serum Albumin, Bovine / Q000737:chemistry; D013054:Spectrophotometry, Atomic; D013696:Temperature
chemical_list: D000911:Antibodies, Monoclonal; D007074:Immunoglobulin G; D009211:Myoglobin; D011506:Proteins; D012710:Serum Albumin, Bovine; D010047:Ovalbumin
keywords: Adsorption; Atomic absorption spectrometry; Protein stability; Size exclusion chromatography