Development of an Immunoassay for the Detection of Amyloid Beta 1-42 and Its Application in Urine Samples

Amyloid beta peptides (A beta 1-42) have been found to be associated with the cause of Alzheimer's disease (AD) and dementia. Currently, methods for detecting A beta 1-42 are complicated and expensive. The present study is aimed at developing an indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) to detect A beta 1-42 by using a polyclonal antibody from alpaca, an application used in urine samples. The serum was collected from the alpaca after immunizing it with A beta 1-42 at 500 mu g/injection 5 times. The ic-ELISA was developed and showed a half-maximal inhibitory concentration (IC50) of 103.20 ng/ml. The limit of detection (LOD) was 0.39 ng/100 mu l. The cross-reactivity was tested with A beta 1-40 and 8 synthesized peptides that had sequence similarities to parts of A beta 1-42. The cross-reactivity of A beta 1-40 and peptide 1 (DAEFRHDSGYE) was 55% and 69.4%, respectively. The ic-ELISA was applied to analyze A beta 1-42 in the urine and precipitated protein urine samples. This method can be used for detecting a normal level of total soluble A beta (approximately 1 ng in 5 mg of precipitated urine protein) and can be used for detecting the early stages of AD. It is considered to be an easy and inexpensive method for monitoring and diagnosing AD.