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Once I’ve got labeled DNA to play with, the next step, if I’m trying to measure force-extension relations anyway, is to incubate DNA and polystyrene beads coated with the appropriate anti-body. It just so happens that the streptavidin coated beads we have are 3.17 μm in diameter, while the anti-dig beads are 2.1 μm in diameter. The larger beads better cover the tip of a micropipette, so residual suction is less of an issue. That means the 2.1 μm beads will be the ones held in the trap, and so it is these beads which I incubate with DNA.
After an hour or so the beads are covered with DNA, so I flow in with the 3.17 μm streptavidin beads, stick one on the tip of the pipette, and then flush the chamber with the DNA-covered beads. When one is trapped, I start a process we lovingly call fishing. Not entirely unlike real fishing, I move the streptavidin bead held on the micropipette away and toward the trapped bead. Normally nothing happens, but when you get a “bite”, when the biotin-labeled end of a piece of DNA finds the binding site on a streptavidin molecule, the bead being held in the trap will start getting pulled away from the trap centre, and I know I’ve got a (or perhaps many) connection.
Like real fishing, sometimes you never get a bite at all, and other days there seems to be an overabundance of them. This image is taken from one of those days when things are working too well, and while it makes for a cute image,
(and note the bead size alternating back and forth, as it has to if the labeling reaction worked as planned!), it is actually quite frustrating when this happens since I need to start all over again, flushing the chambers clean and wasting countless DNA-covered beads. Even so, these days are better than the no-bite days, without a doubt.
Biocurious is written by Andre Brown and Philip Johnson, since 2005. Content of the weblog is licensed under a Creative Commons Attribution-Share Alike 3.0 License.
