[Nanomanipulation of sections of human tongue squamous cell carcinoma by atomic force microscopy].
OBJECTIVE
To investigate the appropriate nanomanipulation of slides of human tongue squamous cell carcinoma tissue and cultured cells by atomic force microscopy (AFM), and develop methods to improve the resolution and contrast of AFM images.


METHODS
Human tongue squamous carcinoma cells of the line Tca8113 were cultured. Cell masses were isolated and collected, and sectioned. Then the white lead and silvery white sections were transferred on mica slides, dried. Eighty slides were used and divided into 2 groups to be immersed into hydrogen dioxide of the concentrations 20% and 30% for 0, 5, 10, 15, 20, 25, 30, and 40 min respectively. Samples of human tongue squamous carcinoma were collected from 15 cases. Every sample was divided into 2 halves. Half of every sample underwent routine paraffin preparation of slide and adhered onto glass, dissolving of paraffin with dimethylbenzene for 0, 10, 15, 30, 45, or 60 min; and half of the sample underwent routine preparation of slides for electron microscopy, the white lead and silvery white sections were transferred on mica slides, dried. The slides were observed by contact mode AFM in air.


RESULTS
In the ultrathin sections the metastructure of the cell such as the cell membrane, nuclear membrane and nucleolus could be distinct from each other clearly. The profile of the cell and the nucleus inside could be revealed by AFM after the paraffin sections were treated in dimethylbenzene for 15-30 min. AFM manipulation could be achieved within nanometer range by precise modulation of scanning force, area, angle and so on. The appropriate needle point should contain elastic coefficient > 5.0 N/m. The force exerted on the needle point should be within the range 50-300 nN. After location the scanning direction should be within the range of 20 nm-1 microm.


CONCLUSIONS
A series of modified techniques of preparing and treating AFM samples, such as adhering the sections onto mica slide, dissolving of epoxy resin by hydrogen dioxide and removal of paraffin by dimethylbenzene, were developed, thus realizing high resolving power AFM imaging, especially for nuclear membrane, and nucleolus, and providing a base of AFM location, dissection, isolation and obtainment at nanometer scale.
Li
Xin-hui
XH
Nanobiology Laboratory, Bio-X Life Science Research Center, Shanghai Jiaotong University, Shanghai 200030, China.
Sun
Jie-lin
JL
Zhang
Ping
P
Hu
Jun
J
Zhang
Chen-ping
CP
Ji
Tong
T