Microfluidic Methods for Fixing and Positioning C. Elegans
The transparent body of C. elegans facilitates real-time imaging and analysis of molecules and cells in its body. The target protein can be fluorescently labeled and observed in vivo. The dynamic process of screening in C. elegans requires precise positioning or immobilization of the worm. Usually, glue or anesthesia is used for the worm in the experiment. However, this will destroy the normal function of the worm. Moreover, it is very difficult to recover from glued or anesthetized worms in most cases, which hinders further screening.
The microfluidic chips can be designed specifically to fix worms in order to perform high-resolution microscopic imaging on the chip without the use of anesthetics or glue. Different fixing methods can be used according to the specifications of the experiment. These microfluidic chip designs can be used in combination with side microchannels to stimulate worms through electrical, mechanical, optical and other means during immobilization. For some experiments or applications, it is necessary to move the worm to a specific location on the chip. The positioning techniques mentioned later may be helpful.
Immobilization Methods
The optical transparency and biocompatibility of the microfluidic chip make it an ideal choice for C. elegans manipulation and immobilization. The worm can be easily released from the chip for further analysis. Each method has different advantages, and the choice of method depends on the experimental requirements and complexity as well as the availability of the hardware and software required to implement these technologies.
Figure.1 Worm immobilization techniques in microfluidic devices. A) Physical immobilization: compression of a membrane deflected by pressure in the control channel. B) Chemical immobilization: CO2 diffusion through an adjacent channel. C) Physical restriction in worm clamp, top view of device design. D) Physical constraint gel. E) Cooling. (San-Miguel A, et al. 2013)
- Air pressure: The chip is made in three layers. The bottom layer through which the worm flows. The top layer is filled with air to compress the thin middle layer to fix the worms. This method will damage the worm.
- CO2 Immobilization: The chip is divided into three layers. The worm penetrates the bottom layer. CO2 passes through the top layer and through the thin porous middle layer. Exposure to CO2 will immobilize the worms for 2-3 hours.
- Tapering: The worms get stuck in the channel due to the back pressure caused by their flow, so they pass through the tapered channel. This method is suitable for high-throughput screening.
- Physical confinement gel: The worm is immobilized by gelling the surrounding liquid at physiological temperature.
- Cooling: The chip applies a flow of cooling fluid in the channel (blue) at the top of the worm imaging channel (red). The green channel represents the valve that stops fluids and worms.
Positioning Techniques
| Electrotaxis | Chemotaxis | Phototaxis |
|---|---|---|
| C. elegans respond to the direction and strength of the electric field. which can be used to move and locate worms within the channel. Placing the electrode in the desired position and applying voltage can help the worm automatically locate it. | C. elegans can recognize chemical cues through two organs located in the head and back, namely amphibians and phages. The response of worms to chemical gradients C. elegans has a response mechanism to various chemicals, odors and gases, which helps to locate the worm inside the microfluidic chip. | Although C. elegans does not carry light-sensitive neurons, it can respond to optical cues through different mechanisms. |
Reference
- San-Miguel A, et al. (2013). "Microfluidics as a tool for C. elegans research." WormBook. 1-19.
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