Anchoring of Aminophosphonates on Titanium Oxide for Biomolecular Coupling

Aminophosphonates were chosen for a first step functionalization of TiO2 grown on titanium, as they possess a phosphonate group on one end, that can be exploited for coupling with the oxide surface, and an amino group on the other end to enable further functionalization of the surface. The deposition of aminophosphonates with different chain lengths (6 and 12 methylenes) was investigated. Oxygen plasma treatment proved useful in increasing the number of −OH groups at the TiO2 surface, thus helping to anchor the aminophosphonates. By combining different surface-sensitive experimental techniques, we found the existence of a discontinuous monolayer where the molecules are covalently coupled to the TiO2 surface. For the molecules with longer chains, we find evidence of their covalent coupling to the surface through Ti–O–P bond formation, of the exposure of the amino groups at the outer surface, and of an increase in the order of the layer upon thermal annealing

Reproducibility warning: The curious case of polyethylene glycol 6000 and spheroid cell culture - Fig 3

MALDI measurements to determine the mass of PEG 6000 produced by S.A. (green), Merck (blue), C.E. (red).</p

Reproducibility warning: The curious case of polyethylene glycol 6000 and spheroid cell culture - Fig 6

Micrographs of HT-29 (A-C) and Hela (D-F) cells without treatment (A, D), or incubated for 5 mins in PEG6000 from C.E. (B, E) and S.A. (C, F) and imaged 48 h after seeding. The scale bars indicate 200 μm.</p

Triple SEC measurement with PEG6000 from S.A., C.E., and Merck.

Triple SEC measurement with PEG6000 from S.A., C.E., and Merck.</p

Reproducibility warning: The curious case of polyethylene glycol 6000 and spheroid cell culture - Fig 2

Micrographs of (A) HeLa, (B) PANC-1; and (C) Caco-2 cells imaged 72 h after seeding on PEG60000 (C.E.) pre-treated surfaces.</p

Reproducibility warning: The curious case of polyethylene glycol 6000 and spheroid cell culture - Fig 5

AFM micrograph of Petri dish surfaces incubated for 1 h with 3% PEG6000 from (A) C.E.; (B) Merck; and (C) S.A. The images were recorded after replacing the PEG solution with Milli-Q water. The scale bars indicate 5 μm.</p

Reproducibility warning: The curious case of polyethylene glycol 6000 and spheroid cell culture - Fig 4

HT29 imaged by transmission light microscopy 96 h after plating in a 96 well plate treated for 1 h at 37°C with (A) PEG6000 from C.E., (B) PEG4000 (4000), (C) PEG6000 from S.A. (D) a mixture of 4000/S.A. 1/5 PRE diluted, (E) a mixture of 4000/S.A. 1/5, (F) mixture of 4000/S.A. 1/10, (G) PEG6000 from MERCK, (H) 4000/MERCK 1/5 pre-diluted, (J) 4000/MERCK 1/5, (K) 4000/MERCK 1/10, and (L) as control HT29 cells on an untreated surface. The scale bar in a) is valid for all images.</p

Reproducibility warning: The curious case of polyethylene glycol 6000 and spheroid cell culture - Fig 1

Microscopic images of HT29 cells on surfaces (1 h; 37°C) treated with PEG6000 from A) C.E.; B) Acros; C) Merck; and D) S.A.. The images were recorded 48 h after cell plating.</p