Stepwise silver-staining-based immunosorbent assay for amyloid-beta autoantibody detection.
OBJECTIVE
To detect amyloid-beta (Abeta) autoantibodies in a reliable and high-throughput fashion, we developed a stepwise silver-staining-based immunosorbent assay in a 96-well-plate platform.


METHODS
Abeta autoantibodies were incubated in an Abeta-immobilized 96-well microplate. Antihuman IgG-modified gold nanoparticle probes were then used to bind to the autoantibodies and signal enhancement was carried out with stepwise silver-staining on immobilized gold nanoparticle probes. A microplate reader was used to quantify silver-stained gold nanoparticles on a well-plate surface.


CONCLUSIONS
Stepwise silver-staining at low temperature enables long-term silver-staining with minimal increase of background signal. This stepwise staining method helps solve the problems of one-step staining, such as nonspecific binding or nonuniformity in silver precipitation after prolonged silver-staining for signal enhancement.


CONCLUSIONS
A stepwise silver-staining strategy could be useful in minimizing nonspecific background signals. This 96-well-plate-based Abeta antibody detection assay could be useful in studying and diagnosing Alzheimer's disease.
Shim
So-Youn
SY
Department of Chemistry, Seoul National University, Seoul, South Korea.
Woo
Jung-Reem
JR
Nam
Eun-Ju
EJ
Hong
Hyun-Joo
HJ
Mook-Jung
Inhee
I
Kim
Young-Ho
YH
Nam
Jwa-Min
JM
D016229:Amyloid beta-Peptides / Q000276:immunology*; D001323:Autoantibodies / Q000032:analysis* / Q000276:immunology; D006801:Humans; D007118:Immunoassay / Q000379:methods; D008855:Microscopy, Electron, Scanning; D036103:Nanotechnology / Q000379:methods; D015203:Reproducibility of Results; D016622:Silver Staining / Q000379:methods*
D016229:Amyloid beta-Peptides; D001323:Autoantibodies