3d localization and diffusion of proteins in polyelectrolyte multilayers

Abstract

The interaction of diverse biomaterials with surfaces is more crucial than ever for biomedical applications to ensure efficiency and reproducibility. Very interesting surface materials are micrometer-thick polyelectrolyte multilayers. Not only their surface but also the bulk can be loaded with biomaterials like proteins or DNA for various purposes. Therefore, we established a method to analyze the lateral and vertical distribution of fluorescently labelled proteins of various size and charge in polyelectrolyte films composed of poly(L-lysine) and hyaluronic acid by confocal laser scanning microscopy. This approach enables us to measure the diffusion coefficients of the proteins via fluorescence recovery after photobleaching as a function of their vertical position in the film and facilitates the understanding of molecular interactions in the film with a high resolution in both space and time. As a result, we confirm that protein loading in the film is driven by electrostatic interactions - uncharged dextran molecules of 10 and 500 kDa do not diffuse into the film. Proteins of different sizes (3-11 nm) can diffuse relatively fast (D = 2-4 mm(2) s(-1)) independent of their net charge, indicating complex interpolymer interactions. This approach is a new powerful experimental tool to design the polyelectrolyte multilayers for bio-applications by finding a relationship between intermolecular interactions and mobility and availability of biomolecules to biological samples (e.g. cells) or detection units (e.g. biosensors)

Biodegradation-Resistant Multilayers Coated with Gold Nanoparticles. Toward a Tailor-made Artificial Extracellular Matrix

“…There is also a lack in understanding the mechanism of biomolecule-multilayer interaction that obviously defines biomolecule presentation/release from multilayers to cells and, as a result, cellular response. [23][24] In this work a new approach for the design of artificial ECM with controlled biodegradation based on the coating of multilayers with gold nanoparticles (GNPs) is present. The multilayers were assembled from glycosaminoglycan hyaluronic acid (HA) and polyaminoacid poly-L-lysine 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 5 To prove this assumption, transport of the molecules of different siz...…”

“…In the case of charged molecules (CF, PLL) electrostatics may play a significant role because of a large number of both positive and negative uncompensated charges in multilayers. 23 Negative charge on citric acid stabilized nanoparticles may thus also play a role.Therefore, uncharged molecules -dextrans -of various molecular masses (10, 70, and 500 kDa) have been employed to evaluate the impact of molecular mass to permeability. Figure 2B 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 6 shows that the cut-off of the uncoated films is between 70 and 500 kDa.…”

Temperature-induced molecular transport through polymer multilayers coated with PNIPAM microgels

“…Nowadays these multilayer films are extensively used as reservoirs to host bioactive molecules aiming at biological applications. [6][7][8][9][10] A variety of molecules can be embedded into the films; they include small drugs and dyes, [11][12][13][14][15][16][17] proteins and peptides, [18][19][20][21] or nucleic acids. 22,23 Extensive reviews on the reservoir properties of the multilayers can be found elsewhere.…”

Polyelectrolyte Multilayers: Towards Single Cell Studies

“…Because of the reason mentioned above, Uhlig and co-workers have developed a method enabling the examination of the multilayer film composition and molecular diffusion in the film with high precision in space and time [86]. CLSM has been adopted for the monitoring of molecular transport in thick (micrometer-sized) multilayer films.…”

“…The insets show the square width at half minimum of the Gaussian w 2 at different time intervals. Adopted from [86].…”