2004-01-17 23:16:00WuYung123
原子力顯微鏡掃瞄DNA
DNA imaging by AFM (原子力顯微鏡掃瞄DNA)
(Based on Bustamante et. al. 1992)
Scanning probe microscopies are based on the ability to detect a local property of a surface by means of a spatially controlled sensor or probe. The spatial distribution of the magnitude of the quantity measured on the surface provides a position-dependent signal or "image" of the surface. Depending on the sensor and the mode of operation,images depict the topography, the electronic structure, or the mechanical or thermal properties of the surface. The spatial resolution of these methods is critically dependent on the property measured and the type of interaction between the probe and the surface.
Commerical cantilevers are microfabricated from Si3N4, Si or SiO2 and have typical spring constants between 0.1 and 1 N/m. For comaprison, the spring constant between two atoms is about 10nN/m.
Poor reproducbility of DNA imaging by AFM is due to:
1. lack of a suitablbly flat, strongly adsorbing substrate,
2. less than optimal tip-moleule interactions,
3. lack of consistenly sharp styli,
4. the need to develop optimal ambient conditions for imaging
When samples are imaged in air, a water layer of varying thickness is present on the sample. In these conditions, the aqueous meniscus formed between the tip and the sample induces capillary forces that scale with the radius of curvature of the tip. Thus in air, the forces involved in the SFM experiment are usually higher than those estimated from pure van der Waals considerations. With shaper tips, it will minimise the dimensions of the air-liquid interface, therefore require a lower force to operate, therefore easier to control. Also lower relative humidity will also reduce the force required. At relative humidity of over 60%, the molecular-tip interactions become comparable to those binding the molecule to the mica surface, allowing the tip to detach and sweep the molecule during the scan. This could be due to the partial solvation of the molecule form the surface due to the screening of the electrostatic charges holding the molecule down and an increase in molecule-tip interaction due to the enhanced surface tension effects.
Routine and reproducible imaging of DNA in air with AFM can be done with:
1. the use of AFM tips with a consistent radius of curvature of 10nm or less to minimize image distortion introduced by the finite dimensions of the tips, and decease the liquid-air interface at the point of contact between the tip and sample.
2. the optimization of a method to deposit and immobilize DNA onto mica surface in a stable manner.
3. careful control of the sample humidity to prevent solvation of the molecules and detachment from the surface by the scanning tip.
(Based on Bustamante et. al. 1992)
Scanning probe microscopies are based on the ability to detect a local property of a surface by means of a spatially controlled sensor or probe. The spatial distribution of the magnitude of the quantity measured on the surface provides a position-dependent signal or "image" of the surface. Depending on the sensor and the mode of operation,images depict the topography, the electronic structure, or the mechanical or thermal properties of the surface. The spatial resolution of these methods is critically dependent on the property measured and the type of interaction between the probe and the surface.
Commerical cantilevers are microfabricated from Si3N4, Si or SiO2 and have typical spring constants between 0.1 and 1 N/m. For comaprison, the spring constant between two atoms is about 10nN/m.
Poor reproducbility of DNA imaging by AFM is due to:
1. lack of a suitablbly flat, strongly adsorbing substrate,
2. less than optimal tip-moleule interactions,
3. lack of consistenly sharp styli,
4. the need to develop optimal ambient conditions for imaging
When samples are imaged in air, a water layer of varying thickness is present on the sample. In these conditions, the aqueous meniscus formed between the tip and the sample induces capillary forces that scale with the radius of curvature of the tip. Thus in air, the forces involved in the SFM experiment are usually higher than those estimated from pure van der Waals considerations. With shaper tips, it will minimise the dimensions of the air-liquid interface, therefore require a lower force to operate, therefore easier to control. Also lower relative humidity will also reduce the force required. At relative humidity of over 60%, the molecular-tip interactions become comparable to those binding the molecule to the mica surface, allowing the tip to detach and sweep the molecule during the scan. This could be due to the partial solvation of the molecule form the surface due to the screening of the electrostatic charges holding the molecule down and an increase in molecule-tip interaction due to the enhanced surface tension effects.
Routine and reproducible imaging of DNA in air with AFM can be done with:
1. the use of AFM tips with a consistent radius of curvature of 10nm or less to minimize image distortion introduced by the finite dimensions of the tips, and decease the liquid-air interface at the point of contact between the tip and sample.
2. the optimization of a method to deposit and immobilize DNA onto mica surface in a stable manner.
3. careful control of the sample humidity to prevent solvation of the molecules and detachment from the surface by the scanning tip.