Research On Cell And Bacteria Morphology And Biomechanics Based On Atomic Force Microscope

Nanobiotechnology, as the cross-discipline of nanotechnology and life science, is a novel technology and approach in nanoscale life science study; it can provide many insightful views into biophysical behaviors of biomolecules and cells including the intramolecular force, intermolecular interaction, and molecule-molecule and cell-cell interaction mechanism. In this work, atomic force microscopy, flow hyetometer, laser scanning confocal microscopy and quantum dot labeling were used to investigate the biophysical properties (the parameters including morphology, membrane nanostructures, adhesion properties, elasticity, stiffiness, specific interaction of antigen-antibody, distribution mode of membrane receptors and so on) of T lymphocytes, DC cells, B16-F10 cells and bacteria. According to the experiments, we obtained many insightful results shown as the followings:1) The nanostructures and nanomechanics of resting and activated T lymphocytes were analyzed. The results indicate that cell diameter, cell height, cell volume and the mean height of membrane surface particles of T cells increased due to PHA or SEA stimulation.The AFM images also revealed that the surface of the T lymphocyte treated with PHA or SEA were rougher than that of resting T lymphocyte, and coated with an outer layer of extracellular nanocluster. High-spatially resolved force-distance curves indicated that the adhesion force values of T lymphocyte activated by PHA or SEA were approximately three to six times stronger than that of resting T lymphocyte, but the activated T cells were softer than resting T cells. These visualized results can improve our understanding the relationship between structures and functional properties of T cells.2) On one hand, the nanostructures and nano-mechanical properties of DCs were firstly evaluated. On other hand, based on AFM and tip modification technology, combined with fluorescent quantum dots (QDs) mark, and laser scanning confocal microscopy, the structure and function or the immune recognition of the biological molecules were studied. The results revealed that the specific binding force of CD28-CD86 was found to be approximately nine times that of the unspecific force. This research afforded useful and new data for studying the second signal about T cell's activation.3) Atomic force microscope was used to detect morphological and mechianical properties of K562 cells and neutrophilic granulocyte. The result indicated that the average roughness value of K562 cell was higher than that of normal neutrophilic. Moreover, analysis of non-specific cell adhesion showed surface adhesion of K562 cells was-30% less than that of normal neutrophilic granulocyte cells. We also found that K562 cells were-70% softer than normal neutrophilic granulocyte cells. Our findings indicated that AFM can be used to diagnose cancer in the future.4) The bacterial inhibition rate of Gram-positive bacteria Listeria monocytogenes and Gram-negative bacteria Salmonella by amoxicillin or hematoporyrin monomethyl ether was investigated by the reduction of colony unit and AFM. The AFM images and force-distance curves showed that the morphology, surface ultrastructure and nanomechanics of Gram-positive bacteria and Gram-negative bacteria changed evidently. It was observed morphologically that the bacterial surface was damaged seriously after treated with amoxicillion or hematoporyrin monomethyl ether. Nanomechanically, we found that treated with amoxicillion or hematoporyrin monomethyl ether caused an increase in the surface adhesion but a decrease in the rigidity while the cell surface changed from smooth to damaged, perhaps due to the changes of the peptidoglycan layer and the change of the bacterial envelope composition from homogenicity to heterogeneity.5) First, changes in the morphology, ultrastructure and nanomechanics of B16-F10 cells were studied by AFM both before and after treatment with the anti-cancer drug (DTIC).In response to DTIC, the microstructure of the cell membrane changed, that is, with increases in drug concentration and reaction time, the degree of morphological changes on the cell membrane increased. These changes included increases in the fluctuation of the surface components of the cell membrane, increase in shrinkage, or even the appearance of pores. The morphological changes on the cell membrane induced the decrease of cell surface adhesion and of cell elasticity. Second, we used the antibody CD44 functional modified AFM tip to detect specific cell surface receptor molecule interaction and to determine the nanoscale distribution of CD44 molecules on cell-membrane. We found that the strength of the specific binding force of the CD44 receptor-ligand was approximately two times that of the unspecific force. The adhesion forces were not randomly distributed over the surface of a B16-F10cell, but the CD44 molecules were concentrated into nanodomain. In addition, the CD44 molecules on the edge and pseudopodia of B16-F10 cells were more than that on the top of the cells. This research has great significance for the study of anti-cancer mechanisms about DTIC.