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A Highly Sensitive Porous Silicon (P-Si)-Based Human Kallikrein 2 (hK2) Immunoassay Platform toward Accurate Diagnosis of Prostate Cancer.

Lee SW, Hosokawa K, Kim S, Jeong OC, Lilja H, Laurell T, Maeda M - Sensors (Basel) (2015)

Bottom Line: The tailored pore size of the porous silicon also allows efficient immobilization of antibodies by surface adsorption, and does not require chemical immobilization.In total 13 × 13 arrays (169 spots) were spotted on the chip with its single spot volume of 300 pL.The best performance of the microarray platform was seen at 100 µg/mL of the capture antibody concentration (LOD was 100 fg/mL).

View Article: PubMed Central - PubMed

Affiliation: Bioengineering Laboratory, RIKEN, Saitama 3510198, Japan. SangWook_L@chem.s.u-tokyo.ac.jp.

ABSTRACT
Levels of total human kallikrein 2 (hK2), a protein involved the pathology of prostate cancer (PCa), could be used as a biomarker to aid in the diagnosis of this disease. In this study, we report on a porous silicon antibody immunoassay platform for the detection of serum levels of total hK2. The surface of porous silicon has a 3-dimensional macro- and nanoporous structure, which offers a large binding capacity for capturing probe molecules. The tailored pore size of the porous silicon also allows efficient immobilization of antibodies by surface adsorption, and does not require chemical immobilization. Monoclonal hK2 capture antibody (6B7) was dispensed onto P-Si chip using a piezoelectric dispenser. In total 13 × 13 arrays (169 spots) were spotted on the chip with its single spot volume of 300 pL. For an optimization of capture antibody condition, we firstly performed an immunoassay of the P-Si microarray under a titration series of hK2 in pure buffer (PBS) at three different antibody densities (75, 100 and 145 µg/mL). The best performance of the microarray platform was seen at 100 µg/mL of the capture antibody concentration (LOD was 100 fg/mL). The platform then was subsequently evaluated for a titration series of serum-spiked hK2 samples. The developed platform utilizes only 15 µL of serum per test and the total assay time is about 3 h, including immobilization of the capture antibody. The detection limit of the hK2 assay was 100 fg/mL in PBS buffer and 1 pg/mL in serum with a dynamic range of 106 (10(-4) to 10(2) ng/mL).

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Titration series of hK2 in buffer (PBS) solution at three different concentrations of the capturing antibody 6B7 (75 µg/mL, 100 µg/mL and 145 µg/mL). The LOD was found to be 1 pg/mL when the capturing antibody was 75 µg/mL and was reduced to 100 fg/mL when the capturing antibody concentrations were 100 µg/mL. The LOD became 1 pg/mL again when concentration of the antibody was 145 µg/mL.
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sensors-15-11972-f003: Titration series of hK2 in buffer (PBS) solution at three different concentrations of the capturing antibody 6B7 (75 µg/mL, 100 µg/mL and 145 µg/mL). The LOD was found to be 1 pg/mL when the capturing antibody was 75 µg/mL and was reduced to 100 fg/mL when the capturing antibody concentrations were 100 µg/mL. The LOD became 1 pg/mL again when concentration of the antibody was 145 µg/mL.

Mentions: To optimize the P-Si hK2 immunoassay, we first evaluated the assay sensitivity by arraying the capture antibody, 6B7, at three different concentrations and performing the hK2 immunoassay in PBS buffer (10 mM, pH 7.2). Figure 3 shows a titration series of hK2 against three different capture antibody concentrations (75, 100 and 145 µg/mL).


A Highly Sensitive Porous Silicon (P-Si)-Based Human Kallikrein 2 (hK2) Immunoassay Platform toward Accurate Diagnosis of Prostate Cancer.

Lee SW, Hosokawa K, Kim S, Jeong OC, Lilja H, Laurell T, Maeda M - Sensors (Basel) (2015)

Titration series of hK2 in buffer (PBS) solution at three different concentrations of the capturing antibody 6B7 (75 µg/mL, 100 µg/mL and 145 µg/mL). The LOD was found to be 1 pg/mL when the capturing antibody was 75 µg/mL and was reduced to 100 fg/mL when the capturing antibody concentrations were 100 µg/mL. The LOD became 1 pg/mL again when concentration of the antibody was 145 µg/mL.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4481930&req=5

sensors-15-11972-f003: Titration series of hK2 in buffer (PBS) solution at three different concentrations of the capturing antibody 6B7 (75 µg/mL, 100 µg/mL and 145 µg/mL). The LOD was found to be 1 pg/mL when the capturing antibody was 75 µg/mL and was reduced to 100 fg/mL when the capturing antibody concentrations were 100 µg/mL. The LOD became 1 pg/mL again when concentration of the antibody was 145 µg/mL.
Mentions: To optimize the P-Si hK2 immunoassay, we first evaluated the assay sensitivity by arraying the capture antibody, 6B7, at three different concentrations and performing the hK2 immunoassay in PBS buffer (10 mM, pH 7.2). Figure 3 shows a titration series of hK2 against three different capture antibody concentrations (75, 100 and 145 µg/mL).

Bottom Line: The tailored pore size of the porous silicon also allows efficient immobilization of antibodies by surface adsorption, and does not require chemical immobilization.In total 13 × 13 arrays (169 spots) were spotted on the chip with its single spot volume of 300 pL.The best performance of the microarray platform was seen at 100 µg/mL of the capture antibody concentration (LOD was 100 fg/mL).

View Article: PubMed Central - PubMed

Affiliation: Bioengineering Laboratory, RIKEN, Saitama 3510198, Japan. SangWook_L@chem.s.u-tokyo.ac.jp.

ABSTRACT
Levels of total human kallikrein 2 (hK2), a protein involved the pathology of prostate cancer (PCa), could be used as a biomarker to aid in the diagnosis of this disease. In this study, we report on a porous silicon antibody immunoassay platform for the detection of serum levels of total hK2. The surface of porous silicon has a 3-dimensional macro- and nanoporous structure, which offers a large binding capacity for capturing probe molecules. The tailored pore size of the porous silicon also allows efficient immobilization of antibodies by surface adsorption, and does not require chemical immobilization. Monoclonal hK2 capture antibody (6B7) was dispensed onto P-Si chip using a piezoelectric dispenser. In total 13 × 13 arrays (169 spots) were spotted on the chip with its single spot volume of 300 pL. For an optimization of capture antibody condition, we firstly performed an immunoassay of the P-Si microarray under a titration series of hK2 in pure buffer (PBS) at three different antibody densities (75, 100 and 145 µg/mL). The best performance of the microarray platform was seen at 100 µg/mL of the capture antibody concentration (LOD was 100 fg/mL). The platform then was subsequently evaluated for a titration series of serum-spiked hK2 samples. The developed platform utilizes only 15 µL of serum per test and the total assay time is about 3 h, including immobilization of the capture antibody. The detection limit of the hK2 assay was 100 fg/mL in PBS buffer and 1 pg/mL in serum with a dynamic range of 106 (10(-4) to 10(2) ng/mL).

Show MeSH
Related in: MedlinePlus