Monitoring the Switching of Single BSA-ATTO 488 Molecules Covalently End-Attached to a pH-Responsive PAA Brush

TY - JOUR

T1 - Monitoring the Switching of Single BSA-ATTO 488 Molecules Covalently End-Attached to a pH-Responsive PAA Brush

AU - Akkilic, Namik

AU - Molenaar, Robert

AU - Claessens, Mireille Maria Anna Elisabeth

AU - Blum, Christian

AU - de Vos, Wiebe Matthijs

PY - 2016

Y1 - 2016

N2 - We describe a novel combination of a responsive polymer brush and a fluorescently labeled biomolecule, where the position of the biomolecule can be switched from inside to outside the brush and vice versa by a change in pH. For this, we grafted ultrathin, amino-terminated poly(acrylic acid) brushes to glass and silicon substrates. Individual bovine serum albumin (BSA) molecules labeled with fluorophore ATTO 488 were covalently end-attached to the polymers in this brush using a bis-N-succinimidyl-(pentaethylene glycol) linker. We investigated the dry layer properties of the brush–protein ensemble, and it is swelling behavior using spectroscopic ellipsometry. Total internal reflection fluorescence (TIRF) microscopy enabled us to study the distance-dependent switching of the fluorescently labeled protein molecules. The fluorescence emission from the labeled proteins ceased (out-state) when the polymer chains stretched away from the interface under basic pH conditions, and fluorescence recurred (in-state) when the chains collapsed under acidic conditions. Moreover, TIRF allowed us to study the fluorescence switching behavior of fluorescently labeled BSA molecules down to the single-molecule level, and we demonstrate that this switching is fast but that the exact intensity during the in-state is the result of a more random process. Control experiments verify that the switching behavior is directly correlated to the responsive behavior of the polymer brush. We propose this system as a platform for switchable sensor applications but also as a method to study the swelling and collapse of individual polymer chains in a responsive polymer brush.

AB - We describe a novel combination of a responsive polymer brush and a fluorescently labeled biomolecule, where the position of the biomolecule can be switched from inside to outside the brush and vice versa by a change in pH. For this, we grafted ultrathin, amino-terminated poly(acrylic acid) brushes to glass and silicon substrates. Individual bovine serum albumin (BSA) molecules labeled with fluorophore ATTO 488 were covalently end-attached to the polymers in this brush using a bis-N-succinimidyl-(pentaethylene glycol) linker. We investigated the dry layer properties of the brush–protein ensemble, and it is swelling behavior using spectroscopic ellipsometry. Total internal reflection fluorescence (TIRF) microscopy enabled us to study the distance-dependent switching of the fluorescently labeled protein molecules. The fluorescence emission from the labeled proteins ceased (out-state) when the polymer chains stretched away from the interface under basic pH conditions, and fluorescence recurred (in-state) when the chains collapsed under acidic conditions. Moreover, TIRF allowed us to study the fluorescence switching behavior of fluorescently labeled BSA molecules down to the single-molecule level, and we demonstrate that this switching is fast but that the exact intensity during the in-state is the result of a more random process. Control experiments verify that the switching behavior is directly correlated to the responsive behavior of the polymer brush. We propose this system as a platform for switchable sensor applications but also as a method to study the swelling and collapse of individual polymer chains in a responsive polymer brush.

KW - IR-102145

KW - METIS-318953

U2 - 10.1021/acs.langmuir.6b01064

DO - 10.1021/acs.langmuir.6b01064

M3 - Article

VL - 32

SP - 8803

EP - 8811

JO - Langmuir

JF - Langmuir

SN - 0743-7463

IS - 35

ER -