AFM

Main Page LM's Logbook RM's Logbook (click to view my milestone report, too) AL's Logbook - Milestone Map on front logbook page.


 * AFM stands for Atomic Force Microscopy**.

|| Here is a sample AFM image I took at the Notre Dame lab some containing DNA. It is DNA on an APTES monolayer surface. || DNA has great potential for use as a structural foundation. It has been used to make a variety of two dimensional, three dimensional and mechanical devices. DNA is currently being studied on a semi-conducting silicon substrate, but the silicon oxide film on the silicon is negatively charged and repels the sugar-phosphate backbone of DNA. The silicon surfaces were treated with an aqueous **APTES** (aminopropyl triethoxysilane, C9H23NO3Si) solution to act as a positively charged adhesive. The goal of this project is to introduce mixed monolayers to manipulate and slightly offset the APTES solution which binds tightly with the silane surface of the silicon, loosening its grip on the DNA. The mixed monolayers on silicon were prepared by mixing various proportions of APTES and **N-Trimethoxysilylpropyl-N,N,N-trimethylammonium chloride** (C9H24ClNO3Si). Also used as monolayers for DNA attachment were **PEG** (Polyethylene glycol) and **OTS** (octadecyltrichlorosilane). Computer software connected to the AFM or Atomic Force Microscope, the main tool in the process, is used to analyze the DNA on the silicon.
 * media type="custom" key="454079"

This is a diagram of the DNA on the APTES surface. The diagram by Dr. Koshala Sarveswaran shows the chemical composition of **APTES**, **which is between the DNA (pink string) and the silicon substrate.**



Here is another image of that same DNA, but on a mica surface instead of a silicon surface. There is a major difference. For one you can clearly see the circular features of the DNA on the surface, in contrast to the image above, which is on silicon.



Also, this was my general project around the end of last year at the science fair at Notre Dame, which would help you get a good understanding of my project



media type="custom" key="454087"

Over the summer at Notre Dame, I tried a variety of other neutral solutions in conjunction with the APTES solution.

Mix 20 μL of APTES (Aminopropyltriethoxysilane)1980 μL of 18 mega ohm water and place into a clean vial. Then submerge silicon wafers for 10 to 15 minutes.
 * __APTES Deposition__**

Same procedure as APTES deposition on the silicon, but replace the word APTES with the word PEG (Polyethylene glycol). Also tried deposition for 1 hour and 2 hours, to see any changes in the DNA
 * __PEG Deposition in Water__**

I deposited 2 mL of Toluene and 40 μL of PEG on the silicon wafer and 10 μL of HCL to act as a catalyst. This is for each vial. I left mine in first for 42 hours, then 21 and 69 hours. Deposition time should be at least 18. The best results were from 21 hours.
 * __PEG deposition with Toluene__**

.04 mL of OTS mixed with 20 mL of solvents (1.5:1:10 ratio of CHCL3:CCL4: Isopar G) to form OTS solution in dry box. This amount was dispensed into each vial where each wafer would be soaked for 3-5 days.
 * __OTS Deposition__**

One problem I encountered in my project was how small the circular DNA plasmids were as seen in the AFM image. Because of this, it was hard to gauge in what ways the APTES and N-Trimethoxysilylpropyl-N,N,N-trimethylammonium chloride affected the DNA. At the Science Fair last year, one judge recommended that I use long-chain DNA rather than small plasmids in order to better view the features and positions of the DNA. What was suggested to me was to use **restriction enzymes** which would cut the DNA plasmid at a certain point which would cause it to relax and spread out. This image, [|from www.sangon.com] shows the circular DNA plasmid's restriction points for certain enzymes. Depending on the enzyme and plasmid you choose, the enzyme will cut the DNA at a different point. This specific plasmid type is not the kind I'm using however. I am using the pUC-19 plasmid so the site might be slightly different (using the kpn I enzyme).

The other part is finding restriction points on the DNA plasmid and finding the right enzymes to cut those points. The process is as follows:

I first take 5 uL (microliters) of the Kpn I restriction enzyme and mix with 10 uL of the buffer solution, 1 uL of a BSA solution and 5 uL of a 1ug/uL concentration, along with 79 uL of water for a 100 uL solution which is put into a tube. I then treat the mixture with chlorophorm, sodium acetate, and ethanol and put the tube in a centerfuge a number of times in a "cold room" at the lab. I can then store the mixture in a freezer for a period of time until I need it, which is after I clean my silicon samples. It is on //these// silicon wafers where I deposit the DNA mixture with the restriction enzyme.

The silicon wafers themselves also have their own cleaning process before the DNA deposition. The wafers first come in large circles which must then be cut into small 1cm x 1cm pieces and then cleaned in a variety of ways. Afterwards they are used as a substrate for the DNA and various monolayers.

The next part of the project is to make DNA necklaces.



This is an image of an AFM similar to the one in my PP presentation. The little cave in the upper part of the microscope is where you place the sample, which you analyze using a nearby computer hooked up to the AFM. The machine is quite sensitive to any sort of vibrations or outside disturbances. Simply touching the table where the machine is could ruin your image. At the lab in the AFM room, there is a machine that turns on that creates ripples in the image every once in a while. This AFM is from the [|Jim Schneider Research Group at Carnegie Mellon University.]