C. elegans stands out as a powerful model organism for many reasons including its simple nervous system and relative optical transparency. Hence, techniques that enable researchers to manipulate the movement and position of C. elegans for the purposes of imaging and for behavioral studies are useful. We have developed a novel method of repositioning the nematode that involves feeding the worms magnetic beads mixed with bacteria, and then using a neodymium magnet to move the worms at will.
- Remove 25 μl of 2.5% w/v 1 μM paramagnetic iron oxide microparticles (Spherotech, AMX-10-10) from the stock solution.
- Use a neodymium magnet (Radio Shack, 640-1895) to separate magnetic particles from buffer, pipet out buffer containing sodium azide, and replace with 25 μl of desired buffer (e.g., M9).
- Mix magnetic particles with 25 μL of liquid culture OP50 bacteria (1:1 dilution).
- Seed this mixture onto an LB agar plate. Pick or chunk a small number of worms onto the plate.
- Let the worms grow on the plate at 16 oC for 7 days [wait longer and your efficiency of obtaining useable worms will increase].
- Using a stereoscope, ensure that you can see the reddish-tinted iron oxide beads throughout the digestive track of the worm (especially near the head and tail regions). Pick the appropriate worms onto an enclosed aqueous medium (e.g., onto a coverslip inserted into a chamber topped with M9 buffer).
- Place the neodymium magnet directly underneath the coverslip (preferably contacting it) at the location that you prefer the nematodes to move to (note that worm may not respond to magnet at distances greater than 25 mm from the center of the magnet).
Upon visual inspection, beads appeared confined to the digestive tract and no negative impact was observed on the development of worms grown on plates with magnetic beads (normal lifespan). The main caveat for this technique is that a long incubation time is required for a satisfactory percentage of worms to consume enough magnetic beads to be sufficiently affected by external magnets. After a 7 day incubation period, ~33% of worms picked at random respond to the magnetic field. Following 14 days of incubation, this percentage increases to ~75%. One can both increase this efficiency and decrease the incubation time by selecting only for worms that have an obvious reddish tint of iron oxide beads throughout their digestive tract.
Note that at shorter incubation times (1-3 days), one should only expect to find adult worms that have consumed a sufficient number of beads. Unfortunately, waiting longer for the nematodes to consume more beads leaves the user with a significantly higher percentage of worms having entered the dauer stage.
We also considered microinjecting beads into the worms or binding them to the surface coat of the worms. The first suggestion was undesirable because of its invasive nature. Binding the magnetic beads to the surface coat of the worm either via ionic bonding (Concanavalin A binding to abundant proteoglycans) or covalent antibody attachment (worm surface coat has the O-glycosylated target for antibody M-38) would be less invasive. However, we speculate that the bond forces holding the beads to the worm would be orders of magnitude less than the forces generated by the living organism’s muscles. Feeding the beads to the worms seemed like the most promising method due to C. elegans’ indiscriminant suction powered eating machinery, the technique’s minimally invasive nature, and the fact that previously nematodes have successfully been fed 0.8 µM latex beads to study food transport through the pharynx (Avery and Shtonda, 2003).
This technology can be used to direct one or more worms to a particular field of view. Microfluidic devices provide the same functionality but take more effort to create, and also introduce a ceiling layer on top of the worm which may be problematic for certain imaging techniques such as inverted selective plane illumination microscopy, in which water dipping objectives are immersed in the aqueous medium from above, to minimize optical aberrations. Another potential use includes behavioral analysis in which researchers need to physically move the worms. With future optimization, this technique may enable other applications.
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