Effect of different coatings on immobilization of biomolecules in brush polymer cells

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Abstract

Biochips with protein and oligonucleotide probes are used to analyze protein and nucleic acid samples. The key challenges of the technology are the selection of substrate materials and surface functionalization. Polybutylene terephthalate substrates were modified by coating them with photoactive polymers: poly(ethylene-co-propylene-co-5-methylene-2-norbornene), acetylcellulose, polyvinyl acetate and polyvinyl butyral. The coatings were applied by centrifugation and dried. The effect of the coating on the biochip characteristics was investigated. A matrix of hydrophilic cells made of brush polymers with epoxy groups for immobilization of DNA probes and human immunoglobulins was prepared by photoinitiated radical polymerization. The functionality of probes was investigated by hybridization analysis and reaction with specific antibodies. The binding efficiency of probes to molecular targets was evaluated on biochips with different coatings. Cells on substrates coated with polyvinyl butyral and poly(ethylene-co-propylene-co-5-methylene-2-norbornene) showed the best binding efficiency and weak adsorption of targets, providing high contrast fluorescence images after probe binding. Biochips on such substrates are promising for lab-on-a-chip microanalysis technology.

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About the authors

G. F. Shtylev

Engelhardt Institute of Molecular Biology, Russian Academy of Sciences

Author for correspondence.
Email: gosha100799@mail.ru
Russian Federation, ul. Vavilova 32/1, Moscow, 119991 Russia

I. Yu. Shishkin

Engelhardt Institute of Molecular Biology, Russian Academy of Sciences

Email: gosha100799@mail.ru
Russian Federation, ul. Vavilova 32/1, Moscow, 119991 Russia

R. A. Miftakhov

Engelhardt Institute of Molecular Biology, Russian Academy of Sciences

Email: gosha100799@mail.ru
Russian Federation, ul. Vavilova 32/1, Moscow, 119991 Russia

S. A. Polyakov

Engelhardt Institute of Molecular Biology, Russian Academy of Sciences

Email: gosha100799@mail.ru
Russian Federation, ul. Vavilova 32/1, Moscow, 119991 Russia

V. E. Shershov

Engelhardt Institute of Molecular Biology, Russian Academy of Sciences

Email: gosha100799@mail.ru
Russian Federation, ul. Vavilova 32/1, Moscow, 119991 Russia

V. E. Kuznetsova

Engelhardt Institute of Molecular Biology, Russian Academy of Sciences

Email: gosha100799@mail.ru
ul. Vavilova 32/1, Moscow, 119991 Russia

S. A. Surzhikov

Engelhardt Institute of Molecular Biology, Russian Academy of Sciences

Email: gosha100799@mail.ru
Russian Federation, ul. Vavilova 32/1, Moscow, 119991 Russia

V. I. Butvilovskaya

Engelhardt Institute of Molecular Biology, Russian Academy of Sciences

Email: gosha100799@mail.ru
Russian Federation, ul. Vavilova 32/1, Moscow, 119991 Russia

V. E. Barsky

Engelhardt Institute of Molecular Biology, Russian Academy of Sciences

Email: gosha100799@mail.ru
Russian Federation, ul. Vavilova 32/1, Moscow, 119991 Russia

V. A. Vasiliskov

Engelhardt Institute of Molecular Biology, Russian Academy of Sciences

Email: gosha100799@mail.ru
Russian Federation, ul. Vavilova 32/1, Moscow, 119991 Russia

О. А. Zasedateleva

Engelhardt Institute of Molecular Biology, Russian Academy of Sciences

Email: gosha100799@mail.ru
Russian Federation, ul. Vavilova 32/1, Moscow, 119991 Russia

A. V. Chudinov

Engelhardt Institute of Molecular Biology, Russian Academy of Sciences

Email: gosha100799@mail.ru
Russian Federation, ul. Vavilova 32/1, Moscow, 119991 Russia

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Supplementary files

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2. Fig. 1. Scheme of chemical modification of polybutylene terephthalate.

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3. Fig. 2. Schematic diagram of the structure of fluorescent dyes.

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4. Fig. 3. Scheme of DNA probe immobilization and its hybridization with a synthetic target in biochip cells obtained by polymerization of a GMA–HEMA–DMAPS mixture.

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5. Fig. 4. Fluorescent patterns of biochip cells obtained on different photoactive surfaces from the GMA–HEMA–DMAP mixture after immobilization of the DNA probe on the Cy3 channel with an exposure of 10 s (rows 1 and 4 contain the DNA probe labeled with Cy3, rows 2 and 3 are empty cells). The graphs of signal distribution along the drawn lines are shown.

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6. Fig. 5. Fluorescent patterns of biochip cells obtained on different photoactive surfaces from a mixture of GMA–HEMA–DMAPs after hybridization with a synthetic target in the Cy5 channel with an exposure of 1 s. The graphs of signal distribution along the drawn lines are shown.

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7. Fig. 6. Scheme of immobilization of Human-IgG immunoglobulin and its specific binding to goat antibodies in biochip cells obtained by polymerization of a GMA–HEMA–DMAPS mixture.

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8. Fig. 7. Fluorescent pattern of biochip cells obtained by photopolymerization of a mixture of HEMA–DMAPS–GMA monomers after immobilization of Human IgG and Human IgG-Су3 with an exposure of 30 s. Graphs of signal distribution along the drawn lines are shown.

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9. Fig. 8. Fluorescent picture of biochip cells obtained by photopolymerization of a mixture of HEMA–DMAPS–GMA monomers after immobilization of Human IgG and binding to developing antibodies Goat anti-Human IgG-Cy5 in the Cy5 channel with an exposure of 1 s.

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