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Development of Indirect Competitive Immuno-Assay Method Using SPR Detection for Rapid and Highly Sensitive Measurement of Salivary Cortisol Levels.

Tahara Y, Huang Z, Kiritoshi T, Onodera T, Toko K - Front Bioeng Biotechnol (2014)

Bottom Line: The surface of an Au chip was modified with PEG6-COOH aromatic dialkanethiol self-assembled monolayers and hydrocortisone 3-(O-carboxymethyl) oxime (hydrocortisone 3-CMO) as a cortisol analog.A detection limit of 38 ppt range with a measurement range of 10 ppt-100 ppb was accomplished without the incubation of a mixing solution consisting of standard cortisol and an anti-cortisol antibody, and the time for quantification of cortisol concentration was 8 min from the sample injection.It was found that the results obtained by the cortisol immuno-assay had a good correlation with those obtained by ELISA assay (R = 0.96).

View Article: PubMed Central - PubMed

Affiliation: Graduate School of Information Science and Electrical Engineering, Kyushu University , Fukuoka , Japan.

ABSTRACT
The monitoring of salivary cortisol as a key biomarker of an individual's stress response has been increasingly focused on. This paper describes the development of a novel cortisol immuno-assay method based on an indirect competitive method using a commercially available surface plasmon resonance instrument. The surface of an Au chip was modified with PEG6-COOH aromatic dialkanethiol self-assembled monolayers and hydrocortisone 3-(O-carboxymethyl) oxime (hydrocortisone 3-CMO) as a cortisol analog. A detection limit of 38 ppt range with a measurement range of 10 ppt-100 ppb was accomplished without the incubation of a mixing solution consisting of standard cortisol and an anti-cortisol antibody, and the time for quantification of cortisol concentration was 8 min from the sample injection. We experimentally compared our immuno-assay with a commercialized salivary cortisol enzyme-linked immunosorbent assay (ELISA) kit using human saliva samples. It was found that the results obtained by the cortisol immuno-assay had a good correlation with those obtained by ELISA assay (R = 0.96). Our findings indicate the potential utility of the cortisol immuno-assay for measurements of human salivary cortisol levels.

No MeSH data available.


Surface plasmon resonance sensorgram obtained from indirect competitive assay.
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Figure 2: Surface plasmon resonance sensorgram obtained from indirect competitive assay.

Mentions: A calibration curve for the cortisol measurement was obtained using a standard cortisol solution consisting of cortisol with various concentrations, HBS, and 2.5% ethanol (n = 3). Figure 2 shows an SPR sensorgram obtained by this measurement. When a mixture of 600 ppb antibody and HBS (final concentration, 300 ppb) was injected onto the chip, the change in the sensor response (Δθ1) was 207.4 ± 4.7 RU (n = 18). Here, 1000 RU corresponds to approximately 1 ng/mm2, and the amount of immobilized antibody was 207.4 pg/mm2. Dissociation of the bound antibody from the sensing surface is required for the next immunocycle. These results indicate that the regeneration conditions are suitable, because the sensor responses (Δθ1) were nearly unchanged.


Development of Indirect Competitive Immuno-Assay Method Using SPR Detection for Rapid and Highly Sensitive Measurement of Salivary Cortisol Levels.

Tahara Y, Huang Z, Kiritoshi T, Onodera T, Toko K - Front Bioeng Biotechnol (2014)

Surface plasmon resonance sensorgram obtained from indirect competitive assay.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4126460&req=5

Figure 2: Surface plasmon resonance sensorgram obtained from indirect competitive assay.
Mentions: A calibration curve for the cortisol measurement was obtained using a standard cortisol solution consisting of cortisol with various concentrations, HBS, and 2.5% ethanol (n = 3). Figure 2 shows an SPR sensorgram obtained by this measurement. When a mixture of 600 ppb antibody and HBS (final concentration, 300 ppb) was injected onto the chip, the change in the sensor response (Δθ1) was 207.4 ± 4.7 RU (n = 18). Here, 1000 RU corresponds to approximately 1 ng/mm2, and the amount of immobilized antibody was 207.4 pg/mm2. Dissociation of the bound antibody from the sensing surface is required for the next immunocycle. These results indicate that the regeneration conditions are suitable, because the sensor responses (Δθ1) were nearly unchanged.

Bottom Line: The surface of an Au chip was modified with PEG6-COOH aromatic dialkanethiol self-assembled monolayers and hydrocortisone 3-(O-carboxymethyl) oxime (hydrocortisone 3-CMO) as a cortisol analog.A detection limit of 38 ppt range with a measurement range of 10 ppt-100 ppb was accomplished without the incubation of a mixing solution consisting of standard cortisol and an anti-cortisol antibody, and the time for quantification of cortisol concentration was 8 min from the sample injection.It was found that the results obtained by the cortisol immuno-assay had a good correlation with those obtained by ELISA assay (R = 0.96).

View Article: PubMed Central - PubMed

Affiliation: Graduate School of Information Science and Electrical Engineering, Kyushu University , Fukuoka , Japan.

ABSTRACT
The monitoring of salivary cortisol as a key biomarker of an individual's stress response has been increasingly focused on. This paper describes the development of a novel cortisol immuno-assay method based on an indirect competitive method using a commercially available surface plasmon resonance instrument. The surface of an Au chip was modified with PEG6-COOH aromatic dialkanethiol self-assembled monolayers and hydrocortisone 3-(O-carboxymethyl) oxime (hydrocortisone 3-CMO) as a cortisol analog. A detection limit of 38 ppt range with a measurement range of 10 ppt-100 ppb was accomplished without the incubation of a mixing solution consisting of standard cortisol and an anti-cortisol antibody, and the time for quantification of cortisol concentration was 8 min from the sample injection. We experimentally compared our immuno-assay with a commercialized salivary cortisol enzyme-linked immunosorbent assay (ELISA) kit using human saliva samples. It was found that the results obtained by the cortisol immuno-assay had a good correlation with those obtained by ELISA assay (R = 0.96). Our findings indicate the potential utility of the cortisol immuno-assay for measurements of human salivary cortisol levels.

No MeSH data available.