Micro OS-ELISA: Rapid noncompetitive detection of a small biomarker peptide by open-sandwich enzyme-linked immunosorbent assay (OS-ELISA) integrated into microfluidic device.
A novel detection system that combines the merits of open-sandwich (OS) enzyme-linked immunoadsorbent assay (ELISA) and a microfluidic sensor chip system, and which enables rapid and noncompetitive immunodetection of small antigens of less than 1000 in molecular weight, has been proposed. Equipped with a sensitive thermal lens microscope, a minute amount of the carboxyl-terminal peptide of human osteocalcin (BGP), a biomarker for bone metabolism, was quantified utilizing antigen-dependent stabilization of an antibody variable region (OS principle). In a short analysis time (approximately 12 min), we could attain a detection limit comparable to that of the microplate-based OS ELISA (1 microg L(-1)). In addition, the effects of several pretreatments for serum-derived samples were investigated: an albumin absorption resin, addition of a protease inhibitor cocktail and heat treatment. Each pretreatment was found to be effective. Consequently, intrinsic BGP and its fragments could be detected in healthy human serum with a superior detection limit and working range compared to those of the conventional competitive ELISA method.
Ihara
Masaki
M
Department of Bioengineering, School of Engineering, The University of Tokyo, Bunkyo, Tokyo, 113-8656, Japan.
Yoshikawa
Amane
A
Wu
Yushu
Y
Takahashi
Hiroko
H
Mawatari
Kazuma
K
Shimura
Kiyohito
K
Sato
Kae
K
Kitamori
Takehiko
T
Ueda
Hiroshi
H
D055910:Antibodies, Immobilized / Q000737:chemistry; D000911:Antibodies, Monoclonal / Q000737:chemistry; D015415:Biomarkers / Q000032:analysis*; D004797:Enzyme-Linked Immunosorbent Assay / Q000295:instrumentation / Q000379:methods*; D004867:Equipment Design; D006801:Humans; D007135:Immunoglobulin Variable Region / Q000737:chemistry; D056656:Lab-On-A-Chip Devices; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation / Q000379:methods*; D015675:Osteocalcin / Q000032:analysis; D010446:Peptide Fragments / Q000032:analysis*; D013997:Time Factors
D055910:Antibodies, Immobilized; D000911:Antibodies, Monoclonal; D015415:Biomarkers; D007135:Immunoglobulin Variable Region; D010446:Peptide Fragments; D015675:Osteocalcin