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Biomolecular Interaction Analysis of Gestrinone-anti-Gestrinone Using Arrays of High Aspect Ratio SU-8 Nanopillars.

Ortega FJ, Bañuls MJ, Sanza FJ, Casquel R, Laguna MF, Holgado M, López-Romero D, Barrios CA, Maquieira Á, Puchades R - Biosensors (Basel) (2012)

Bottom Line: After gestrinone antigen immobilization on the BICELLs, the immunorecognition was performed.The cells were interrogated vertically by using micron spot size Fourier transform visible and IR spectrometry (FT-VIS-IR), and the dip wavenumber shift was monitored.The biosensing assay exhibited good reproducibility and sensitivity (LOD = 0.75 ng/mL).

View Article: PubMed Central - PubMed

Affiliation: Centro de Reconocimiento Molecular y Desarrollo Tecnológico, Departamento de Química, Universitat Politècnica de València, Camino de Vera s/n, Valencia 46022, Spain. fraorhi1@upvnet.upv.es.

ABSTRACT
In this paper, label-free biosensing for antibody screening by periodic lattices of high-aspect ratio SU-8 nano-pillars (BICELLs) is presented. As a demonstration, the determination of anti-gestrinone antibodies from whole rabbit serum is carried out, and for the first time, the dissociation constant (KD = 6 nM) of antigen-antibody recognition process is calculated using this sensing system. After gestrinone antigen immobilization on the BICELLs, the immunorecognition was performed. The cells were interrogated vertically by using micron spot size Fourier transform visible and IR spectrometry (FT-VIS-IR), and the dip wavenumber shift was monitored. The biosensing assay exhibited good reproducibility and sensitivity (LOD = 0.75 ng/mL).

No MeSH data available.


Related in: MedlinePlus

Synthesis of gestrinone oxime hapten (a) and gestrinone oxime hapten-horseradish peroxidase (HRP) conjugate (b).
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biosensors-02-00291-f003: Synthesis of gestrinone oxime hapten (a) and gestrinone oxime hapten-horseradish peroxidase (HRP) conjugate (b).

Mentions: Once the successful immobilization of bioreceptors on the SU-8 polymer was demonstrated, their bioavailability for a recognition event was studied. For that, the model system gestrinone/anti-gestrinone was employed. The synthetic steroid gestrinone has anti-estrogenic and anti-progesterone properties, and it is involved in several pharmacological applications including uterine pathologies [29], contraception [30,31], and endometriosis [32]. Due to gestrinone’s anabolic effects, it is the only marketed progestin included in the banned list of performance-enhancing drugs in sports (The world anti-doping code, www.wada-ama.org). In our case, gestrinone oxime was used as steroid hapten conjugated to HRP (HRP-h-G) (Figure 3), which is more appropriate to obtain specific assays because antibodies are raised against a particular moiety of a selected steroid. First of all, the immobilization of HRP was checked using the protocol used with BSA, employing TMB as enzymatic substrate for the assay development. Thus, the presence of the enzyme was detected through the appearance of a dark blue precipitate whose intensity was proportional to the amount of immobilized HRP.


Biomolecular Interaction Analysis of Gestrinone-anti-Gestrinone Using Arrays of High Aspect Ratio SU-8 Nanopillars.

Ortega FJ, Bañuls MJ, Sanza FJ, Casquel R, Laguna MF, Holgado M, López-Romero D, Barrios CA, Maquieira Á, Puchades R - Biosensors (Basel) (2012)

Synthesis of gestrinone oxime hapten (a) and gestrinone oxime hapten-horseradish peroxidase (HRP) conjugate (b).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4263551&req=5

biosensors-02-00291-f003: Synthesis of gestrinone oxime hapten (a) and gestrinone oxime hapten-horseradish peroxidase (HRP) conjugate (b).
Mentions: Once the successful immobilization of bioreceptors on the SU-8 polymer was demonstrated, their bioavailability for a recognition event was studied. For that, the model system gestrinone/anti-gestrinone was employed. The synthetic steroid gestrinone has anti-estrogenic and anti-progesterone properties, and it is involved in several pharmacological applications including uterine pathologies [29], contraception [30,31], and endometriosis [32]. Due to gestrinone’s anabolic effects, it is the only marketed progestin included in the banned list of performance-enhancing drugs in sports (The world anti-doping code, www.wada-ama.org). In our case, gestrinone oxime was used as steroid hapten conjugated to HRP (HRP-h-G) (Figure 3), which is more appropriate to obtain specific assays because antibodies are raised against a particular moiety of a selected steroid. First of all, the immobilization of HRP was checked using the protocol used with BSA, employing TMB as enzymatic substrate for the assay development. Thus, the presence of the enzyme was detected through the appearance of a dark blue precipitate whose intensity was proportional to the amount of immobilized HRP.

Bottom Line: After gestrinone antigen immobilization on the BICELLs, the immunorecognition was performed.The cells were interrogated vertically by using micron spot size Fourier transform visible and IR spectrometry (FT-VIS-IR), and the dip wavenumber shift was monitored.The biosensing assay exhibited good reproducibility and sensitivity (LOD = 0.75 ng/mL).

View Article: PubMed Central - PubMed

Affiliation: Centro de Reconocimiento Molecular y Desarrollo Tecnológico, Departamento de Química, Universitat Politècnica de València, Camino de Vera s/n, Valencia 46022, Spain. fraorhi1@upvnet.upv.es.

ABSTRACT
In this paper, label-free biosensing for antibody screening by periodic lattices of high-aspect ratio SU-8 nano-pillars (BICELLs) is presented. As a demonstration, the determination of anti-gestrinone antibodies from whole rabbit serum is carried out, and for the first time, the dissociation constant (KD = 6 nM) of antigen-antibody recognition process is calculated using this sensing system. After gestrinone antigen immobilization on the BICELLs, the immunorecognition was performed. The cells were interrogated vertically by using micron spot size Fourier transform visible and IR spectrometry (FT-VIS-IR), and the dip wavenumber shift was monitored. The biosensing assay exhibited good reproducibility and sensitivity (LOD = 0.75 ng/mL).

No MeSH data available.


Related in: MedlinePlus