title: Development of homogeneous 384-well high-throughput screening assays for Abeta1-40 and Abeta1-42 using AlphaScreen technology.
abstract: Amyloid beta (Abeta) peptides are the major constituent of amyloid plaques, one of the hallmark pathologies of Alzheimer's disease. Accurate and precise quantitation of these peptides in biological fluids is a critical component of Alzheimer's disease research. The current most established assay for analysis of Abeta peptides in preclinical research is enzyme-linked immunosorbent assay (ELISA), which, although sensitive and of proven utility, is a multistep, labor-intensive assay that is difficult to automate completely. To overcome these limitations, the authors have developed and optimized simple, sensitive, homogeneous 384-well assays for Abeta1-42 and Abeta1-40 using AlphaScreen technology. The assays are capable of detecting Abeta peptides at concentrations <2 pg/mL and, using a final assay volume of 20 microL, routinely generate Z' values >0.85. The AlphaScreen format has the following key advantages: substantially less hands-on time to run, easier to automate, higher throughput, and less expensive to run than the traditional ELISA. The results presented here show that AlphaScreen technology permits robust, efficient, and cost-effective quantitation of Abeta peptides.
authors.affiliation: Eli Lilly and Co. Ltd, Lilly Research Centre, Windlesham, Surrey, UK. szekeresp@lilly.com
authors.forename: Philip G
authors.initials: PG
authors.lastname: Szekeres
authors.forename: Kaitlin
authors.initials: K
authors.lastname: Leong
authors.forename: Theresa A
authors.initials: TA
authors.lastname: Day
authors.forename: Ann E
authors.initials: AE
authors.lastname: Kingston
authors.forename: Eric H
authors.initials: EH
authors.lastname: Karran
mesh_terms: D000544:Alzheimer Disease / Q000175:diagnosis*; D000595:Amino Acid Sequence; D016229:Amyloid beta-Peptides / Q000032:analysis / Q000276:immunology / Q000302:isolation & purification*; D000818:Animals; D000911:Antibodies, Monoclonal / Q000494:pharmacology; D001923:Brain Chemistry; D002138:Calibration; D002478:Cells, Cultured; D003362:Cost-Benefit Analysis; D003470:Culture Media / Q000737:chemistry; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D005260:Female; D006801:Humans; D008403:Mass Screening / Q000191:economics / Q000379:methods*; D051379:Mice; D008810:Mice, Inbred C57BL; D008954:Models, Biological; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000032:analysis / Q000276:immunology / Q000302:isolation & purification*; D012680:Sensitivity and Specificity; D014020:Tissue Extracts / Q000032:analysis / Q000737:chemistry
chemical_list: D016229:Amyloid beta-Peptides; D000911:Antibodies, Monoclonal; D003470:Culture Media; D010446:Peptide Fragments; D014020:Tissue Extracts; C078592:amyloid beta-protein (1-40); C075222:amyloid beta-protein (1-42)