ABSTRACT
In this prospective study, for the first time we have separated and quantified amyloid beta (Abeta) peptides in the plasma of patients with Alzheimer's disease (AD, n = 8) and age- and environment-matched healthy controls (n = 9) with urea-based Abeta-sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE)/immunoblot. In addition to the Abeta peptides 1-37/38/39/40/42, which we recently identified as regular constituents of human cerebrospinal fluid (CSF), we have observed a novel electrophoretic band migrating slightly cathodically to Abeta1-42. Since a standard peptide with the amino acid sequence Abeta2-40 migrates in the same position, we hypothesize that this plasma-specific band may correspond to Abeta2-40. The concentration of Abeta peptides in the plasma has been approximately 100-fold lower compared to the CSF. Interestingly, the concentration of the two shortest peptides and the longest one of these considered here (i.e., Abeta1-37/38/42) have increased significantly when the samples have been frozen at -80 degrees C before immunoprecipitation, while the 'middle-length' peptides (i.e., Abeta1-39/40) have not been affected by this procedure. We have not observed significant differences of the Abeta peptides concentrations between AD and control subjects. Our method can be used to investigate the significance of plasma Abeta peptides in neurodegenerative disorders, and to monitor the efficiency of drugs with beta/gamma-secretase inhibitory potency.
Subject(s)
Alzheimer Disease/diagnosis , Amyloid beta-Peptides/blood , Electrophoresis, Polyacrylamide Gel , Peptide Fragments/blood , Aged , Alzheimer Disease/metabolism , Amyloid beta-Peptides/immunology , Amyloid beta-Peptides/isolation & purification , Humans , Immunoprecipitation , Male , Middle Aged , Peptide Fragments/immunology , Peptide Fragments/isolation & purificationABSTRACT
BACKGROUND: The precise measurement of osmolality is crucial in the differential diagnosis of disorders of water balance. Storage conditions, and freezing and thawing of serum or plasma samples before osmometry may influence the accuracy of measured values. METHODS: A series of serum and plasma samples of 25 healthy young individuals were stored under different conditions at different temperatures (room temperature (22 degrees C), 7 degrees C, -21 degrees C, -78 degrees C) for up to 56 days. Before freezing a protein-stabilizing agent (bacitracin) was added to one part of the samples. Osmolality was examined using the freezing point method. RESULTS: At room temperature osmolality was stable for up to 3 days but showed a tendency toward an increase that was significant on day 14. In contrast, at 7 degrees C an initial significant decrease in serum osmolality occurred (day 1), which was followed by a slow increase. Serum samples stored at -21 degrees C showed a significantly lower osmolality on the 14th day compared to baseline. Adding bacitracin before freezing reduced this decrease by more than half, but the deviation was still significant. In samples stored at -78 degrees C no significant alteration of osmolality from baseline was observed over the observation period of 56 days if samples were thawed in a 37 degrees C water bath. CONCLUSION: Immediate measurement of osmolality is most reliable in order to obtain accurate values, although storing at room temperature does not influence osmolality significantly during the first 3 days. If storage is necessary for longer, samples should be stored at -78 degrees C and must be thawed quickly (at 37 degrees C). Under these conditions reliable values can be obtained from frozen serum or plasma. Storage at 7 degrees C is not recommended. If samples are stored at -21 degrees C the addition of a protein-stabilizing agent may be useful.
