Application of Organosilane Monolayer Template to Quantitative Evaluation of Cancer Cell Adhesive Ability

Abstract

The adhesive ability of two human pancreatic cancer cell lines was evaluated using organosilane monolayer templates (OMTs). Using the OMT, the spreading area of adhered cells can be limited, and this enables us to focus on the initial attachment process of adhesion. Moreover, it becomes possible to arrange the cells in an array and to quantitatively evaluate the number of attached cells. The adhesive ability of the cancer cells cultured on the OMT was controlled by adding (À)-epigallocatechin-3-gallate (EGCG), which blocks a receptor that mediates cell adhesion and is overexpressed in cancer cells. Measurement of the relative ability of the cancer cells to attach to the OMT revealed that the ability for attachment decreased with increasing EGCG concentration. The results agreed well with the western blot analysis, indicating that the OMT can potentially be employed to evaluate the adhesive ability of various cancer cells. #

Quantitative Evaluation of Cancer Cell Adhesion to Self-Assembled Monolayer-Patterned Substrates by Reflection Interference Contrast Microscopy

“…To fabricate SAM-patterned glass substrates, we followed protocols described in our previous reports. 8,9 In brief, tetraethyl orthosilicate (TEOS)-coated coverslips (diameter Φ = 15 mm, thickness = 0.17 mm) were used as a glass substrate. The organosilane monolayer was patterned so as to array cell permissive spots regularly in nonpermissive background on the coverslip.…”

“…8 On the other hand, the outside regions were modified with an n-octadecyltrimethoxysilane (ODS) where a greater amount of albumin in FBS is adsorbed compared to APTES SAM and thus serves as cell-repellent surface. 9 The averaged diameter of cell-adhesive spots and their lattice spacing were estimated to be approximately 18 and 100 μm, respectively. The coverslip was fixed to a plastic cell culture dish with a hole (Φ = 10 mm) at the center.…”

“…Recently, Tanii and co-workers developed a unique method for quantifying cancer cell adhesion by using a self-assembled monolayer (SAM)-patterned substrate where the glass substrate surface was functionalized with aminosilane (cell-adhesive) and alkylsilane (cell-repellent) monolayers. 8,9 Since an array of cell- adhesive spots with optimized diameter can prevent cell−cell adhesion and thus allow single cells to adhere on each spot, the method enabled one to carry out quick in vitro assay of cancer cell adhesion by counting the number of spots occupied by cancer cells. However, such on−off classification of each spots by "occupied" or "nonoccupied" does not provide adhesion states of cancer cells and their dynamic development after attachment, which may be obtained by carefully observing cells that adhere on each spot.…”

Photopatterning Proteins and Cells in Aqueous Environment Using TiO₂ Photocatalysis

“…Use a stage micrometer and close the field diaphragm to set the size of the region to be irradiated, e.g., 200 μm. 4. After the 3 hr incubation, supplement the medium with 200 μl of 3.0 mg ml -1 type-IV collagen (Col-IV; final concentration 300 μg ml -1…”

“…Semiconductor lithography processes and its derivatives -such as photolithography 1,2 , electron-beam lithography [3][4][5][6] , and microcontact printing [7][8][9][10] -have now become an established tool in cell biology to grow living cells in a defined position and geometry. The patterning method relies on the use of microfabricated substrates, consisting of micro-island of cell permissive coating in a non-permissive background.…”