The doors on the oversized freight elevator slam shut. We slowly descend until we are forty feet below the surface of the earth. The doors open and we file out of the elevator into a long, artificially lit stretch of hall. We have reached the depths of Building 219 on the National Institute of Standards and Technology (NIST) campus in Gaithersburg, Maryland, where every aspect of the local environment – temperature, humidity, lighting, etc. – is strictly controlled.
“The researchers fight to work here,” mechanical engineer Jon Pratt remarks as we walk down the pristine hallway. Pratt and research chemist Gordon Shaw are creating reliable and accurate intrinsic (based on nature) reference standards as part of the Mass and Force Group of NIST’s Manufacturing Engineering Laboratory.
Shaw leads us to a room housing an atomic force microscope (AFM). We pass through one door, walk a few feet, and then pass through another door. This double isolation system helps keep the vibrations in the microscope room to a minimum and the temperature at 20 degrees Celsius plus or minus two-hundredths of a degree. Humidity is maintained at 30 percent plus or minus 1 percent. Not a bad place to work, especially during one of the hottest summers on record in the Washington, DC area.
Light is piped into the room along the ceiling so that no one light bulb creates a hot spot. “Thermogradients are the enemy,” Shaw says. “Even small temperature changes can cause the measurement system to be unstable enough that it won’t be able to measure what you’re interested in.”
With an AFM, you can view objects over 1,000 times smaller than with conventional light microscopy. The AFM uses a cantilever system to characterize materials at the atomic and molecular levels. A cantilever resembles a miniature diving board with a sharp silicone tip.
“As you bring the tip close to the surface, it may start to bend toward the surface because of the electrostatic force interactions with the surface,” Shaw notes.
The cantilever tip moves slowly back and forth across the surface of the sample. We look in amazement as the monitor starts to display a pattern that resembles an intertwined assortment of twigs. Shaw notes that the AFM is imaging collagen fibers. Collagen – the most abundant protein in mammals – is found in tendons, ligaments, skin, cartilage, and bone, among other tissues.
Shaw tells us that he would have preferred to image DNA this evening, but he acknowledges that it is rather “tricky” to do so.
Tricky, but not impossible. Actually, Shaw and Pratt have successfully used the AFM to measure the mechanical properties of a single DNA molecule so that it can serve as an intrinsic reference standard for force.
DNA normally forms random coils in solution. In a typical experiment, Shaw and Pratt would move the cantilever around, hunting for the DNA in a solution. They would coat the tip of the cantilever with a chemical that specifically binds to a substance found at one end of a DNA molecule. Eventually, they would get a molecule of DNA attached to the tip of the cantilever. The researchers would then begin to stretch the DNA molecule. At a certain point, typically at a force of approximately 65 piconewtons, the DNA molecule would stretch for a long distance with very little extra applied force.
“Some of the base pairs in the DNA are starting to come apart,” Shaw says. “You have a partial melting of the double helix at that point. That happens as far as we can tell at a fairly well-defined force.”
As Shaw and Pratt and another NIST colleague, Douglas Smith, note in a recent article published in the Proceedings of the SEM Annual Conference, a DNA molecule can therefore be used as a “force reference that will allow the calibration of a wide variety of force measuring instruments such as optical, magnetic, and dielectrophoretic tweezers.”
And thanks to modern polymerase chain reaction (PCR) methods, the same DNA molecule can be made over and over again. “In an afternoon’s work you can make enough to give everyone on the planet 5,000 force references,” Shaw notes as we exit through the double-door climate and vibration control system.
The freight elevator slowly rises to ground level. The doors open and natural light filters in through the first-floor windows. We have returned to a “normal” hallway, where a simple thermostat keeps the temperature at a comfortable level and rooms have just one door.
Still, I tread lightly as I walk down the hall.
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