Electrochemical detection of HbA1c, a marker [correction of maker] for diabetes, using a flow immunoassay system.
An on-chip electrochemical flow immunoassay system for the detection of hemoglobin A1c (HbA1c) was developed using anti-human hemoglobin (Hb) IgG labeled with ferrocene monocarboxylic acid (Fc-COOH) and boronate-affinity chromatography. An on-chip column packed with boronate-activated agarose beads was used for the separation of HbA1c from both non-glycated Hb and free antibody. Anti-human Hb IgG conjugated to Fc-COOH (Fc-IgG) was used for the electrochemical detection of HbA1c. The assay procedure included immunoreactions with Fc-IgG and HbA1c, separation of immunocomplexes by boronate affinity, and electrochemical detection of Fc-IgG-HbA1c immunocomplexes. The immunoreaction mixtures were injected onto a boronate-affinity column. HbA1c-antibody complexes were then trapped onto the column by the affinity of HbA1c to boronic acid. Subsequently, elution buffer containing sorbitol was applied to elute HbA1c-antibody complexes and a current was detected by applying 600 mV versus Ag/AgCl. The elution signal was an estimation of the HbA1c amount. A linear correlation between the increase of current and HbA1c concentration was obtained up to an HbA1c concentration of 500 microg/ml. The HbA1c flow immunoassay was successfully achieved using hemolysates. This electrochemical flow immunoassay system enabled us to construct a novel point-of-care testing device for the monitoring of glycated proteins including HbA1c.
Tanaka
Tsuyoshi
T
Department of Biotechnology, Tokyo University of Agriculture and Technology, Koganei, Tokyo 184-8588, Japan.
Tsukube
Shoko
S
Izawa
Kojiro
K
Okochi
Mina
M
Lim
Tae-Kyu
TK
Watanabe
Shugo
S
Harada
Manabu
M
Matsunaga
Tadashi
T
D000818:Animals; D015415:Biomarkers / Q000032:analysis; D002846:Chromatography, Affinity; D003920:Diabetes Mellitus / Q000175:diagnosis* / Q000378:metabolism; D004563:Electrochemistry; D006442:Glycated Hemoglobin; D006454:Hemoglobins / Q000032:analysis* / Q000378:metabolism; D006801:Humans; D007118:Immunoassay
D015415:Biomarkers; D006442:Glycated Hemoglobin; D006454:Hemoglobins; C517652:hemoglobin A1c protein, human