Name: a microfluidic platform for longitudinal imaging in caenorhabditis elegans
Uploaded: 2018-05-02T13:30:00
Description: Watch this Scientific Journal Video about A Microfluidic Platform for Longitudinal Imaging in Caenorhabditis elegans at JoVE.com

This research was supported by the National Science Foundation through grants PHY-1205494 and MCB-1413134 (EL) and by the National Research Foundation of Korea grant 2017R1D1A1B03035671 (KSL).

The protocol below uses WormSpa5, a previously-described microfluidic device for longitudinal imaging of worms. Fabrication of WormSpa (starting with CAD files that can be obtained from the authors upon request) is straightforward but requires some expertise. In most cases, fabrication can readily be done by a core facility or by a commercial company that provides such services. When fabricating the device, make sure to specify that the height of the features is 50 &#181;m.
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<ol>
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Microfluidic tools provide multiple benefits in studying worms. Imaging in a PDMS device offers higher imaging quality as compared with a standard NGM agar plate. Multiple images can be taken from a single worm, in contrast with traditional methods in which animals are picked from the plate and mounted on a microscope slide for imaging. In addition, the microenvironment in which worms reside can be kept constant or modulated as desired, permitting precise mapping between the composition of the environment and the respons...

Changes in environmental conditions may lead to activation of genetic programs accompanied by induction and repression of the expression of specific genes1,2. These kinetic changes may be variable among tissues in the same animals and between different animals. Studies of such genetic programs therefore call for methods that allow longitudinal imaging of individual animals and provide precise dynamical control of environmental conditions.
In recent years, microfabricated fluidic devices have been used to study many aspects of response and behavior in small animals, including worms, flies, water bears and more3,4,5,6,7. Applications include, for example, deep phenotyping, optogenetic recording of neuronal activity in response to chemical stimuli, and tracking of motor behaviors such as locomotion and pumping8,9,10,11.
Microfluidic-based approaches hold many properties that could benefit long-term longitudinal imaging of response to environmental cues, including precise dynamical control of the local microenvironment, flexible design that allows maintenance of individual animals in separate quarters, and favorable attributes for imaging. However, maintaining animals in a microfluidic chamber for a long time with minimal adverse impact on their well-beings is a challenge, which requires particular care in the design of the microfluidic device as well as in the execution of the experiment.
Here we demonstrate the use of WormSpa, a microfluidic device for longitudinal imaging of Caenorhabditis elegans.5 Individual worms are confined in chambers. A constant low flow of liquid and bacterial suspension guarantees that worms are well-fed and sufficiently active to maintain good health and alleviate stress, and the structure of the chambers allows worms to lay eggs. The simplicity of the design and operation of WormSpa should allow researchers with no previous experience in microfluidics to incorporate this device into their own research plans.

<table border="0" cellpadding="0" cellspacing="0" style="width:1012px;" width="1011">
	<colgroup>
		<col />
		<col />
		<col />
		<col />
	</colgroup>
	<tbody>
		<tr height="40">
			<td height="40" style="height:40px;width:216px;">WormSpa</td>
			<td style="width:123px;">N/A</td>
			<td style="width:344px;">N/A</td>
			<td style="width:329px;">The CAD file for WormSpa is available from the Levine lab.</td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">Compound Microscope</td>
			<td>Zeiss</td>
			<td>AxioObserver Z1</td>
			<td>An inverted fluorescence microscope with a motorized stage</td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">Syringe Pump</td>
			<td>New Era Pump Systems</td>
			<td>NE-501</td>
			<td></td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">Tubing</td>
			<td>SCI Scientific Commodities Inc.</td>
			<td>BB31695-PE/5</td>
			<td>0.034&#8221; (0.86 mm) I.D. x 0.052&#8221; (1.32 mm) O.D</td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">Syringe Tip</td>
			<td>CMLsupply</td>
			<td>901-20-050</td>
			<td>20 Gauge x 1/2&#8221; blunt tip stainless steel canula</td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">Syringe Filter</td>
			<td>PALL</td>
			<td>4650</td>
			<td>Acrodisc 32 mm Syringe Filter with 5 um Supor Membrane</td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">Syringe</td>
			<td>Qosina</td>
			<td>C3307</td>
			<td>10 mL Male Luer Lock Syringe</td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">3 Way Valve</td>
			<td>ColeParmer</td>
			<td>FF-30600-23</td>
			<td>Large-bore 3-way, male-lock, stopcocks, 10/pack, Non-sterile</td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">Dowel Pin</td>
			<td>McMaster-Carr</td>
			<td>90145A317</td>
			<td>18-8 Stainless Steel Dowel Pins (1/32&#34; Dia. x 1/2&#34; Lg.)</td>
		</tr>
		<tr height="40">
			<td height="40" style="height:40px;">Low Binding Microcentrifuge Tube</td>
			<td style="width:123px;">Corning</td>
			<td>CL S3206</td>
			<td>0.65 mL low binding snap cap microcentrifuge tube</td>
		</tr>
	</tbody>
</table>

a microfluidic platform for longitudinal imaging in caenorhabditis elegans

In the last decade, microfluidic techniques have been applied to study small animals, including the nematode Caenorhabditis elegans, and have proved useful as a convenient live imaging platform providing capabilities for precise control of experimental conditions in real time. In this article, we demonstrate live imaging of individual worms employing WormSpa, a previously-published custom microfluidic device. In the device, multiple worms are individually confined to separate chambers, allowing multiplexed longitudinal surveillance of various biological processes. To illustrate the capability, we performed proof-of-principle experiments in which worms were infected in the device with pathogenic bacteria, and the dynamics of expression of immune response genes and egg laying were monitored continuously in individual animals. The simple design and operation of this device make it suitable for users with no previous experience with microfluidic-based experiments. We propose that this approach will be useful for many researchers interested in longitudinal observations of biological processes under well-defined conditions.

Age-synchronized young adult worms (46 hours post L1 larval arrest at 25 &#176;C)12 were loaded in the device, as described in the protocol. The worms were individually located in separate channels, enabling longitudinal measurement of animals&#39; response to the pathogen. When the experiment is successful, most worms remain in their channels for the duration of the experiment. In this case, images of individual worms are taken simply by placing their channel in t...

Watch this Scientific Journal Video about A Microfluidic Platform for Longitudinal Imaging in Caenorhabditis elegans at JoVE.com

A Microfluidic Platform for Longitudinal Imaging in Caenorhabditis elegans

In this article, we demonstrate live imaging of individual worms employing a custom microfluidic device. In the device, multiple worms are individually confined to separate chambers, allowing multiplexed longitudinal surveillance of various biological processes.

A Microfluidic Platform for Longitudinal Imaging in Caenorhabditis elegans

In the last decade, microfluidic techniques have been applied to study small animals, including the nematode Caenorhabditis elegans, and have proved ...

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