We thank Dr. Krishanu Ray for Drosophila stocks, Tarjani Agarwal for maintaining a Drosophila cage, Peter Juo for nuIs25 and CGC for C. elegans strains. SPK made jsIs609 in Michael Nonet's laboratory. We thank Arpan Agnihotri (BITS Pilani) for his help in time-lapse imaging of mitochondria transport of jsIs609 animals in microfluidic devices. We are grateful to Dr. Vatsala Thirumalai and Surya Prakash for providing us with zebrafish embryos. We thank Dr. Krishna and CIFF at NCBS for use of the spinning disc confocal microscope supported by the Department of Science and Technology- Centre for Nanotechnology (No. SR/55/NM-36-2005). We also thank Kaustubh Rau, V. Venkatraman and Chetana Sachidanand for discussions. This work was funded by the DBT post-doctoral fellowship (S.M.), DST fast-track scheme (S.M.) and a DBT grant to (S.P.K.). S.A. was supported by DST and CSIR grants to SPK.

1. SU8 Master Fabrication
<ol>
 <li>Design the microfluidic structures using Clewin software and print it using 65,024 DPI laser plotter with minimum feature size of 8 &mu;m on circuit board film. </li>
 <li>Clean 2 cm X 2 cm silicon wafers with native oxide in 20% KOH for 1 min and rinse in deionized water; one wafer each for the flow layer and its corresponding control layer. </li>
 <li>Blow dry the pieces with nitrogen gas and dehydrate on a hot plate at 120 &deg;C for 4 hr. Allow the pieces to cool down to ro...

<ol>
	<li>Whitesides, G. M., Ostuni, E., Takayama, S., Jiang, X., Ingber, D. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Soft+lithography+in+biology+and+biochemistry.">Soft lithography in biology and biochemistry.</a> Annu. Rev. Biomed. Eng. 3, 335-373 (2001).</li><li>Guo, S. X. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Femtosecond+laser+nanoaxotomy+lab-on-a-chip+for+in+vivo+nerve+regeneration+studies.">Femtosecond laser nanoaxotomy lab-on-a-chip for in vivo nerve regeneration studies.</a> Nat. Methods. 5, 531-533 (2008).</li><li>Gilleland, C. L., Rohde, C. B., Zeng, F., Yanik, M. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microfluidic+immobilization+of+physiologically+active+Caenorhabditis+elegans.">Microfluidic immobilization of physiologically active Caenorhabditis elegans.</a> Nat. Protoc. 5, 1888-1902 (2010).</li><li>Mondal, S., Ahlawat, S., Rau, K., Venkataraman, V., Koushika, S. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Imaging+in+vivo+neuronal+transport+in+genetic+model+organisms+using+microfluidic+devices.">Imaging in vivo neuronal transport in genetic model organisms using microfluidic devices.</a> Traffic. 12, 372-385 (2011).</li><li>Chung, K., Crane, M. M., Lu, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Automated+on-chip+rapid+microscopy,+phenotyping+and+sorting+of+C.+elegans.">Automated on-chip rapid microscopy, phenotyping and sorting of C. elegans.</a> Nat. Methods. 5, 637-643 (2008).</li><li>Allen, P. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Single-synapse+ablation+and+long-term+imaging+in+live+C.+elegans.">Single-synapse ablation and long-term imaging in live C. elegans.</a> J. Neurosci. Methods. 173, 20-26 (2008).</li><li>Chronis, N., Zimmer, M., Bargmann, C. I. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microfluidics+for+in+vivo+imaging+of+neuronal+and+behavioral+activity+in+Caenorhabditis+elegans.">Microfluidics for in vivo imaging of neuronal and behavioral activity in Caenorhabditis elegans.</a> Nat. Methods. 4, 727-731 (2007).</li><li>Rohde, C. B., Zeng, F., Gonzalez-Rubio, R., Angel, M., Yanik, M. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microfluidic+system+for+on-chip+high-throughput+whole-animal+sorting+and+screening+at+subcellular+resolution.">Microfluidic system for on-chip high-throughput whole-animal sorting and screening at subcellular resolution.</a> Proc. Natl. Acad. Sci. U.S.A. 104, 13891-13895 (2007).</li><li>Hulme, S. E., Shevkoplyas, S. S., Apfeld, J., Fontana, W., Whitesides, G. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+microfabricated+array+of+clamps+for+immobilizing+and+imaging+C.+elegans.">A microfabricated array of clamps for immobilizing and imaging C. elegans.</a> Lab Chip. 7, 1515-1523 (2007).</li><li>Zeng, F., Rohde, C. B., Yanik, M. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sub-cellular+precision+on-chip+small-animal+immobilization,+multi-photon+imaging+and+femtosecond-laser+manipulation.">Sub-cellular precision on-chip small-animal immobilization, multi-photon imaging and femtosecond-laser manipulation.</a> Lab Chip. 8, 653-656 (2008).</li><li>Chokshi, T. V., Ben-Yakar, A., Chronis, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=CO2+and+compressive+immobilization+of+C.+elegans+on-chip.">CO2 and compressive immobilization of C. elegans on-chip.</a> Lab Chip. 9, 151-157 (2009).</li><li>Krajniak, J., Lu, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Long-term+high-resolution+imaging+and+culture+of+C.+elegans+in+chip-gel+hybrid+microfluidic+device+for+developmental+studies.">Long-term high-resolution imaging and culture of C. elegans in chip-gel hybrid microfluidic device for developmental studies.</a> Lab Chip. 10, 1862-1868 (2010).</li><li>Goodman, M. B., Hall, D. H., Avery, L., Lockery, S. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Active+currents+regulate+sensitivity+and+dynamic+range+in+C.+elegans+neurons.">Active currents regulate sensitivity and dynamic range in C. elegans neurons.</a> Neuron. 20, 763-772 (1998).</li><li>Snow, J. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Two+anterograde+intraflagellar+transport+motors+cooperate+to+build+sensory+cilia+on+C.+elegans+neurons.">Two anterograde intraflagellar transport motors cooperate to build sensory cilia on C. elegans neurons.</a> Nat. Cell Biol. 6, 1109-1113 (2004).</li><li>Lewis, J. A., Wu, C. H., Berg, H., Levine, J. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+genetics+of+levamisole+resistance+in+the+nematode+Caenorhabditis+elegans.">The genetics of levamisole resistance in the nematode Caenorhabditis elegans.</a> Genetics. 95, 905-928 (1980).</li><li>Richmond, J. E., Jorgensen, E. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=One+GABA+and+two+acetylcholine+receptors+function+at+the+C.+elegans+neuromuscular+junction.">One GABA and two acetylcholine receptors function at the C. elegans neuromuscular junction.</a> Nat. Neurosci. 2, 791-797 (1999).</li><li>Badre, N. H., Martin, M. E., Cooper, R. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+physiological+and+behavioral+effects+of+carbon+dioxide+on+Drosophila+melanogaster+larvae.">The physiological and behavioral effects of carbon dioxide on Drosophila melanogaster larvae.</a> Comp. Biochem. Physiol. A. Mol. Integr. Physiol. 140, 363-376 (2005).</li><li>Stiernagle, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Maintenance+of+C.+elegans.">Maintenance of C. elegans.</a> WormBook. , 1-11 (2006).</li><li>Westerfield, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+zebrafish+book.">The zebrafish book.</a> A guide for the laboratory use of zebrafish (Danio rerio). , (2000).</li><li>Henn, K., Braunbeck, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dechorionation+as+a+tool+to+improve+the+fish+embryo+toxicity+test+(FET)+with+the+zebrafish+(Danio+rerio).">Dechorionation as a tool to improve the fish embryo toxicity test (FET) with the zebrafish (Danio rerio).</a> Comp. Biochem. Physiol. C. Toxicol. Pharmacol. 153, 91-98 (2011).</li><li>Fatouros, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Inhibition+of+tau+aggregation+in+a+novel+Caenorhabditis+elegans+model+of+tauopathy+mitigates+proteotoxicity.">Inhibition of tau aggregation in a novel Caenorhabditis elegans model of tauopathy mitigates proteotoxicity.</a> Hum. Mol. Genet. , (2012).</li><li>Barkus, R. V., Klyachko, O., Horiuchi, D., Dickson, B. J., Saxton, W. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Identification+of+an+axonal+kinesin-3+motor+for+fast+anterograde+vesicle+transport+that+facilitates+retrograde+transport+of+neuropeptides.">Identification of an axonal kinesin-3 motor for fast anterograde vesicle transport that facilitates retrograde transport of neuropeptides.</a> Mol. Biol. Cell. 19, 274-283 (2008).</li><li>Craig, M. P., Gilday, S. D., Hove, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dose-dependent+effects+of+chemical+immobilization+on+the+heart+rate+of+embryonic+zebrafish.">Dose-dependent effects of chemical immobilization on the heart rate of embryonic zebrafish.</a> Lab. Anim. (NY). 35, 41-47 (2006).</li><li>Pardo-Martin, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=High-throughput+in+vivo+vertebrate+screening.">High-throughput in vivo vertebrate screening.</a> Nat. Methods. 7, 634-636 (2010).</li><li>Stainier, D. Y., Lee, R. K., Fishman, M. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cardiovascular+development+in+the+zebrafish.+I.+Myocardial+fate+map+and+heart+tube+formation.">Cardiovascular development in the zebrafish. I. Myocardial fate map and heart tube formation.</a> Development. 119, 31-40 (1993).</li><li>Hall, D. H., Hedgecock, E. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Kinesin-related+gene+unc-104+is+required+for+axonal+transport+of+synaptic+vesicles+in+C.+elegans.">Kinesin-related gene unc-104 is required for axonal transport of synaptic vesicles in C. elegans.</a> Cell. 65, 837-847 (1991).</li><li>Kumar, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Caenorhabditis+elegans+Kinesin-3+motor+UNC-104/KIF1A+is+degraded+upon+loss+of+specific+binding+to+cargo.">The Caenorhabditis elegans Kinesin-3 motor UNC-104/KIF1A is degraded upon loss of specific binding to cargo.</a> PLoS Genet. 6, e1001200 (2010).</li><li>Ou, G., Vale, R. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Molecular+signatures+of+cell+migration+in+C.+elegans+Q+neuroblasts.">Molecular signatures of cell migration in C. elegans Q neuroblasts.</a> J. Cell. Biol. 185, 77-85 (2009).</li><li>Levitan, E. S., Lanni, F., Shakiryanova, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=In+vivo+imaging+of+vesicle+motion+and+release+at+the+Drosophila+neuromuscular+junction.">In vivo imaging of vesicle motion and release at the Drosophila neuromuscular junction.</a> Nat. Protoc. 2, 1117-1125 (2007).</li><li>Morris, R. L., Hollenbeck, P. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+regulation+of+bidirectional+mitochondrial+transport+is+coordinated+with+axonal+outgrowth.">The regulation of bidirectional mitochondrial transport is coordinated with axonal outgrowth.</a> J. Cell. Sci. 104, 917-927 (1993).</li><li>Louie, K., Russo, G. J., Salkoff, D. B., Wellington, A., Zinsmaier, K. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+imaging+conditions+on+mitochondrial+transport+and+length+in+larval+motor+axons+of+Drosophila.">Effects of imaging conditions on mitochondrial transport and length in larval motor axons of Drosophila.</a> Comp. Biochem. Physiol. A. Mol. Integr. Physiol. 151, 159-172 (2008).</li><li>Pilling, A. D., Horiuchi, D., Lively, C. M., Saxton, W. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Kinesin-1+and+Dynein+are+the+primary+motors+for+fast+transport+of+mitochondria+in+Drosophila+motor+axons.">Kinesin-1 and Dynein are the primary motors for fast transport of mitochondria in Drosophila motor axons.</a> Mol. Biol. Cell. 17, 2057-2068 (2006).</li></ol>

PDMS microfluidic devices are optically transparent therefore can be used for high resolution in vivo imaging of any transparent/translucent model organism. Our design is suitable for high magnification spatio-temporal imaging of cellular and sub-cellular events in intact live animals. Microfabrication using soft lithography techniques allows easy manipulation of device dimensions for various sizes of model organisms. Devices of various sizes are fabricated for different stages of C. elegans, Drosop...

<table border="1">
<tbody>
 <tr>
 <td>Name of the reagent</td>
 <td>Company</td>
 <td>Catalogue number</td>
 <td>Comments (optional)</td>
 </tr>
 <tr>
 <td>Silicon wafers</td>
 <td>University wafer</td>
 <td>150 mm (100) Mech Grade SSP Si</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>Clewin Software</td>
 <td>WieWeb software</td>
 <td>Version 2.90</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>Laser plotter</td>
 <td>Fine Line Imaging</td>
 <td>65,024 DPI</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>HMDS</td>
 <td>Sigma-Aldrich</td>
 <td>440191-100ML</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>SU8</td>
 <td>Microchem</td>
 <td>SU8-2025, SU8-2050</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>Developer</td>
 <td>Microchem</td>
 <td>SU8 Developer</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>Silane</td>
 <td>Sigma-Aldrich</td>
 <td>448931-10G</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>PDMS</td>
 <td>Dow corning</td>
 <td>Sylgard 184</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>UV lamp</td>
 <td>Oriel</td>
 <td>66943</td>
 <td>200W Hg Oriel Light</td>
 </tr>
 <tr>
 <td>Hot air oven</td>
 <td>Ultra Instruments</td>
 <td>Custom made</td>
 <td>Set at 50 &deg;C</td>
 </tr>
 <tr>
 <td>Hot plate</td>
 <td>IKA Laboratory Equipment</td>
 <td>3810000</td>
 <td><a href="http://www.ika.com" target="_blank">http://www.ika.com</a></td>
 </tr>
 <tr>
 <td>Plasma cleaner</td>
 <td>Harrick Plasma</td>
 <td>PDC-32G</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>Spinner</td>
 <td>Semiconductor Production Systems</td>
 <td>SPIN150-NPP</td>
 <td><a href="http://www.SPS-Europe.com" target="_blank">www.SPS-Europe.com</a></td>
 </tr>
 <tr>
 <td>Glass cover slip</td>
 <td>Gold Seal</td>
 <td>22 X 22 mm, No. 1 thickness</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>C. elegans</td>
 <td>Caenorhabditis Genetics Center (CGC)</td>
 <td>e1265, ayIs4</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>Drosophila</td>
 <td>Bloomington</td>
 <td>P{chaGAL4}/cyo, UAS-syt.eGFP</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>Zebrafish</td>
 <td>Indian wild type</td>
 <td>Wild type</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>Tygon tube</td>
 <td>Sigma</td>
 <td>Z279803</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>Micro needle</td>
 <td>Sigma</td>
 <td>Z118044</td>
 <td>Cut into 1 cm pieces</td>
 </tr>
 <tr>
 <td>3-way stopcock</td>
 <td>Sigma</td>
 <td>S7521</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>Harris puncher</td>
 <td>Sigma</td>
 <td>Z708631</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>Compressed nitrogen gas</td>
 <td>Local Gas supplier</td>
 <td>&nbsp;</td>
 <td>Use a regulator to control the pressure</td>
 </tr>
 <tr>
 <td>Stereo microscope</td>
 <td>Nikon</td>
 <td>SMZ645</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>Confocal microscope</td>
 <td>Andor &amp; Olympus</td>
 <td>Yokogawa spinning disc confocal microscope</td>
 <td>&nbsp;</td>
 </tr>
 <tr>
 <td>ImageJ</td>
 <td>National Institutes of Health</td>
 <td><a href="http://www.rsbweb.nih.gov/ij" target="_blank">www.rsbweb.nih.gov/ij</a> </td>
 <td>Java based image processing program</td>
 </tr>
 </tbody>
</table>

simple microfluidic devices for in vivo imaging of c. elegans, drosophila and zebrafish

Micro fabricated fluidic devices provide an accessible micro-environment for in vivo studies on small organisms. Simple fabrication processes are available for microfluidic devices using soft lithography techniques 1-3. Microfluidic devices have been used for sub-cellular imaging 4,5, in vivo laser microsurgery 2,6 and cellular imaging 4,7. In vivo imaging requires immobilization of organisms. This has been achieved using suction 5,8, tapered channels 6,7,9, deformable membranes 2-4,10, suction with additional cooling 5, anesthetic gas 11, temperature sensitive gels 12, cyanoacrylate glue 13 and anesthetics such as levamisole 14,15. Commonly used anesthetics influence synaptic transmission 16,17 and are known to have detrimental effects on sub-cellular neuronal transport 4. In this study we demonstrate a membrane based poly-dimethyl-siloxane (PDMS) device that allows anesthetic free immobilization of intact genetic model organisms such as Caenorhabditis elegans (C. elegans), Drosophila larvae and zebrafish larvae. These model organisms are suitable for in vivo studies in microfluidic devices because of their small diameters and optically transparent or translucent bodies. Body diameters range from ~10 &mu;m to ~800 &mu;m for early larval stages of C. elegans and zebrafish larvae and require microfluidic devices of different sizes to achieve complete immobilization for high resolution time-lapse imaging. These organisms are immobilized using pressure applied by compressed nitrogen gas through a liquid column and imaged using an inverted microscope. Animals released from the trap return to normal locomotion within 10 min.
We demonstrate four applications of time-lapse imaging in C. elegans namely, imaging mitochondrial transport in neurons, pre-synaptic vesicle transport in a transport-defective mutant, glutamate receptor transport and Q neuroblast cell division. Data obtained from such movies show that microfluidic immobilization is a useful and accurate means of acquiring in vivo data of cellular and sub-cellular events when compared to anesthetized animals (Figure 1J and 3C-F 4).
Device dimensions were altered to allow time-lapse imaging of different stages of C. elegans, first instar Drosophila larvae and zebrafish larvae. Transport of vesicles marked with synaptotagmin tagged with GFP (syt.eGFP) in sensory neurons shows directed motion of synaptic vesicle markers expressed in cholinergic sensory neurons in intact first instar Drosophila larvae. A similar device has been used to carry out time-lapse imaging of heartbeat in ~30 hr post fertilization (hpf) zebrafish larvae. These data show that the simple devices we have developed can be applied to a variety of model systems to study several cell biological and developmental phenomena in vivo.

The immobilization device is a bilayer PDMS block fabricated by bonding two layers: a flow layer (Layer 1) and a control layer (Layer 2) as shown in Figure 1. The main trap is connected to a nitrogen gas cylinder through a regulator and a 3-way stop cock to apply necessary (3-14 psi) pressure onto the membrane through a liquid column (Figure 1A). The deflected membrane immobilizes C. elegans, Drosophila or zebrafish larvae in the flow channel designed with different dim...

Watch this Scientific Journal Video about Simple Microfluidic Devices for in vivo Imaging of C. elegans, Drosophila and Zebrafish at JoVE.com

Simple Microfluidic Devices for in vivo Imaging of C. elegans, Drosophila and Zebrafish

A simple microfluidic device has been developed to perform anesthetic free in vivo imaging of C. elegans, intact Drosophila larvae and zebrafish larvae. The device utilizes a deformable PDMS membrane to immobilize these model organisms in order to perform time lapse imaging of numerous processes such as heart beat, cell division and sub-cellular neuronal transport. We demonstrate the use of this device and show examples of different types of data collected from different model systems.

Simple Microfluidic Devices for in vivo Imaging of C. elegans, Drosophila and Zebrafish

Micro fabricated fluidic devices provide an accessible micro-environment for in vivo studies on small organisms. Simple fabrication processes are ...

simple-microfluidic-devices-for-vivo-imaging-c-elegans-drosophila

Research

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Bioengineering

17.0K ビュー.  NCBS-TIFR. シンプルなマイクロ流体デバイスは、麻酔薬を無料で行うために開発されましたイメージング Cエレガンス無傷ショウジョウバエ幼虫とゼブラフィッシュの幼虫。デバイスは、心拍、細胞分裂や細胞内輸送などの多数の神経プロセスのタイムラプスイメージングを実行するために、これらのモデル生物を固定化...

ビデオ: Simple Microfluidic Devices for in vivo Imaging of C. elegans, Drosophila and Zebrafish

A simple microfluidic device has been developed to perform anesthetic free in vivo imaging of C. elegans, intact Drosophila larvae and zebrafish larvae. The device utilizes a deformable PDMS membrane to immobilize these model organisms in order to perform time lapse imaging of numerous processes such as heart beat, cell division and sub-cellular neuronal transport.  We demonstrate the use of this device and show examples of different types of data collected from different model...

3d particle tracking for noninvasive in vivo analysis of synaptic microtubule dynamics in dendrites and neuromuscular junctions of drosophila

3D Particle Tracking for Noninvasive In Vivo Analysis of Synaptic Microtubule Dynamics in Dendrites and Neuromuscular Junctions of Drosophila

This study presents a noninvasive intravital neuronal imaging strategy combined with a new software strategy to achieve automated, unbiased tracking and analysis of in vivo microtubule (MT) plus-end dynamics in the sensory dendrites and the neuromuscular junctions of Drosophila.

immobilization of caenorhabditis elegans to analyze intracellular transport in neurons

Immobilization of Caenorhabditis elegans to Analyze Intracellular Transport in Neurons

Caenorhabditis elegans (C. elegans) is a good model to study axonal and intracellular transport. Here, I describe a protocol for in vivo recording and analysis of axonal and intraflagellar transport in C.

Immobilization of Caenorhabditis elegans to Analyze Intracellular Transport in Neurons

quantitative approaches for studying cellular structures and organelle morphology in caenorhabditis elegans

Quantitative Approaches for Studying Cellular Structures and Organelle Morphology in Caenorhabditis elegans

This study outlines quantitative measurements of synaptic size and localization, muscle morphology, and mitochondrial shape in C. elegans using freely available image processing tools. This approach allows future studies in&#160;C. elegans&#160;to quantitatively compare the extent of tissue and organelle structural changes as a result of genetic...

Quantitative Approaches for Studying Cellular Structures and Organelle Morphology in Caenorhabditis elegans

using microfluidics chips for live imaging and study of injury responses in drosophila larvae

Using Microfluidics Chips for Live Imaging and Study of Injury Responses in Drosophila Larvae

Drosophila larvae are an attractive model system for live imaging due to their translucent cuticle and powerful genetics. This protocol describes how to utilize a single-layer PDMS device, called the &#39;larva chip&#39; for live imaging of cellular processes within neurons of 3rd instar Drosophila...

Using Microfluidics Chips for Live Imaging and Study of Injury Responses in Drosophila Larvae

a simple microfluidic chip for long-term growth and imaging of caenorhabditis elegans

A Simple Microfluidic Chip for Long-Term Growth and Imaging of Caenorhabditis elegans

The protocol describes a simple microfluidic chip design and microfabrication methodology used to grow C. elegans in presence of a continuous food supply for up to 36 h. The growth and imaging device also enables intermittent long-term high-resolution imaging of cellular and sub-cellular processes during development for several days.

A Simple Microfluidic Chip for Long-Term Growth and Imaging of Caenorhabditis elegans

in vivo neuronal calcium imaging in c. elegans

In vivo Neuronal Calcium Imaging in C. elegans

With its small transparent body, well-documented neuroanatomy and a host of amenable genetic techniques and reagents, C. elegans makes an ideal model organism for in vivo neuronal imaging using relatively simple, low-cost techniques. Here we describe single neuron imaging within intact adult animals using genetically encoded fluorescent calcium...

In vivo Neuronal Calcium Imaging in C. elegans

in vivo calcium imaging in c. elegans body wall muscles

In Vivo Calcium Imaging in C. elegans Body Wall Muscles

This method provides a way to couple optogenetics and genetically encoded calcium sensors to image baseline cytosolic calcium levels and changes in evoked calcium transients in the body wall muscles of the model organism C.

In Vivo Calcium Imaging in C. elegans Body Wall Muscles

setting up a simple light sheet microscope for in toto imaging of c. elegans development

Setting Up a Simple Light Sheet Microscope for In Toto Imaging of C. elegans Development

This protocol describes the setup of a light sheet microscope and its implementation for in vivo imaging of C. elegans...

Setting Up a Simple Light Sheet Microscope for In Toto Imaging of C. elegans Development

in vivo laser axotomy in c. elegans

In vivo Laser Axotomy in C. elegans

A protocol to cut neurons in C. elegans with a MicroPoint  pulsed laser is presented. We describe setting up the system, immobilizing worms, and severing labeled neurons. Advantages include a relatively low-cost system and the ability to sever neuronal processes or ablate cells in...

In vivo Laser Axotomy in C. elegans

in vivo imaging of dauer-specific neuronal remodeling in c. elegans

In Vivo Imaging of Dauer-specific Neuronal Remodeling in C. elegans

Following exposure to specific environmental stressors, the nematode Caenorhabditis elegans undergoes extensive phenotypic plasticity to enter into a stress-resistant &#8216;dauer&#8217; juvenile stage.  We present methods for the controlled induction and imaging of neuroplasticity during dauer.

In Vivo Imaging of Dauer-specific Neuronal Remodeling in C. elegans

microstructured devices for optimized microinjection and imaging of zebrafish larvae

Microstructured Devices for Optimized Microinjection and Imaging of Zebrafish Larvae

Microinjection of zebrafish embryos and larvae is a crucial but challenging technique used in many zebrafish models. Here, we present a range of microscale tools to aid in the stabilization and orientation of zebrafish for both microinjection and...

a microfluidic platform for longitudinal imaging in caenorhabditis elegans

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...

A Microfluidic Platform for Longitudinal Imaging in Caenorhabditis elegans

using a microfluidics device for mechanical stimulation and high resolution imaging of c. elegans

Using a Microfluidics Device for Mechanical Stimulation and High Resolution Imaging of C. elegans

New tools for mechanobiology research are needed to understand how mechanical stress activates biochemical pathways and elicits biological responses. Here, we showcase a new method for selective mechanical stimulation of immobilized animals with a microfluidic trap allowing high-resolution imaging of cellular...

Using a Microfluidics Device for Mechanical Stimulation and High Resolution Imaging of C. elegans

using an adapted microfluidic olfactory chip for the imaging of neuronal activity in response to pheromones in male c. elegans head neurons

Using an Adapted Microfluidic Olfactory Chip for the Imaging of Neuronal Activity in Response to Pheromones in Male C. Elegans Head Neurons

The use of an adapted &#34;olfactory chip&#34; for the efficient calcium imaging of C. elegans males is described here. Studies of male exposure to glycerol and a pheromone are also...

Using an Adapted Microfluidic Olfactory Chip for the Imaging of Neuronal Activity in Response to Pheromones in Male C. Elegans Head Neurons

a smartphone-based imaging method for c. elegans lawn avoidance assay

Author Spotlight: A Smartphone-Based Imaging Method for C. elegans Lawn Avoidance Assay  

This article describes a simple, low-cost method of recording the lawn avoidance behavior of Caenorhabditis elegans, using readily available items such as a smartphone and a light emitting diode (LED) light box. We also provide a Python script to process the video file into a format more amenable for counting.

Smartphone-Based Imaging for &lt;em&gt;C. elegans&lt;/em&gt; Lawn Avoidance Assay

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imaging and quantifying mitochondrial morphology in c. elegans during aging

Author Spotlight: Advancing Imaging and Quantification Methods to Study Mitochondrial Homeostasis in C. elegans

This protocol provides a standardized approach for imaging mitochondrial morphology in multiple tissues of C. elegans during...

Imaging and Quantifying Mitochondrial Morphology in C. elegans During Aging

rapid fabrication of custom microfluidic devices for research and educational applications

Rapid Fabrication of Custom Microfluidic Devices for Research and Educational Applications

Here we present a protocol to design and fabricate custom microfluidic devices with minimal financial and time investment. The aim is to facilitate the adoption of microfluidic technologies in biomedical research laboratories and educational...

in vivo visualization of calcium transients during fertilization and early development in c. elegans

Author Spotlight: Deciphering the Cellular Signals During Coordinated Oocyte to Embryo Transition in &lt;i&gt;C. elegans&lt;/i&gt;

Here, we present protocols and tools for visualizing calcium during fertilization and early embryogenesis using a genetically encoded calcium reporter expressed in the germline of the model nematode C.

In Vivo Visualization of Calcium Transients during Fertilization and Early Development in C. elegans

thermal measurement techniques in analytical microfluidic devices

Thermal Measurement Techniques in Analytical Microfluidic Devices

Here, we present three protocols for thermal measurements in microfluidic devices.

Separating Beads and Cells in Multi-channel Microfluidic Devices Using Dielectrophoresis and Laminar Flow

Microfluidic devices have advanced cell studies by providing a dynamic fluidic environment on the scale of the cell for studying, manipulating, sorting and counting cells. However, manipulating the cell within the fluidic domain remains a challenge and requires complicated fabrication protocols for forming valves and electrodes, or demands specialty equipment like optical tweezers. Here, we demonstrate that conventional printed circuit boards (PCB) can be used for the non-contact manipulation of cells by employing dielectrophoresis (DEP) for bead and cell manipulation in laminar flow fields for bioactuation, and for cell and bead separation in multichannel microfluidic devices. First, we present the protocol for assembling the DEP electrodes and microfluidic devices, and preparing the cells for DEP. Then, we characterize the DEP operation with polystyrene beads. Lastly, we show representative results of bead and cell separation in a multichannel microfluidic device. In summary, DEP is an effective method for manipulating particles (beads or cells) within microfluidic devices.

Dielectrophoresis (DEP) is an effective method to manipulate cells. Printed circuit boards (PCB) can provide inexpensive, reusable and effective electrodes for contact-free cell manipulation within microfluidic devices. By combining PDMS-based microfluidic channels with coverslips on PCBs, we demonstrate bead and cell manipulation and separation within multichannel microfluidic devices.

Microfluidic devices have advanced cell studies by providing a dynamic fluidic environment on the scale of the cell for studying, manipulating, sorting ...

A Microfluidic Technique to Probe Cell Deformability

Here we detail the design, fabrication, and use of a microfluidic device to evaluate the deformability of a large number of individual cells in an efficient manner. Typically, data for ~102 cells can be acquired within a 1 hr experiment. An automated image analysis program enables efficient post-experiment analysis of image data, enabling processing to be complete within a few hours. Our device geometry is unique in that cells must deform through a series of micron-scale constrictions, thereby enabling the initial deformation and time-dependent relaxation of individual cells to be assayed. The applicability of this method to human promyelocytic leukemia (HL-60) cells is demonstrated. Driving cells to deform through micron-scale constrictions using pressure-driven flow, we observe that human promyelocytic (HL-60) cells momentarily occlude the first constriction for a median time of 9.3 msec before passaging more quickly through the subsequent constrictions with a median transit time of 4.0 msec per constriction. By contrast, all-trans retinoic acid-treated (neutrophil-type) HL-60 cells occlude the first constriction for only 4.3 msec before passaging through the subsequent constrictions with a median transit time of 3.3 msec. This method can provide insight into the viscoelastic nature of cells, and ultimately reveal the molecular origins of this behavior.

We demonstrate a microfluidics-based assay to measure the timescale for cells to transit through a sequence of micron-scale constrictions.

Here we detail the design, fabrication, and use of a microfluidic device to evaluate the deformability of a large number of individual cells in an ...

Layer-by-layer Collagen Deposition in Microfluidic Devices for Microtissue Stabilization

Although microfluidics provides exquisite control of the cellular microenvironment, culturing cells within microfluidic devices can be challenging. 3D culture of cells in collagen type I gels helps to stabilize cell morphology and function, which is necessary for creating microfluidic tissue models in microdevices. Translating traditional 3D culture techniques for tissue culture plates to microfluidic devices is often difficult because of the limited channel dimensions. In this method, we describe a technique for modifying native type I collagen to generate polycationic and polyanionic collagen solutions that can be used with layer-by-layer deposition to create ultrathin collagen assemblies on top of cells cultured in microfluidic devices. These thin collagen layers stabilize cell morphology and function, as shown using primary hepatocytes as an example cell, allowing for the long term culture of microtissues in microfluidic devices.

The creation of functional microtissues within microfluidic devices requires the stabilization of cell phenotypes by adapting traditional cell culture techniques to the limited spatial dimensions in microdevices. Modification of collagen allows the layer-by-layer deposition of ultrathin collagen assemblies that can stabilize primary cells, such as hepatocytes, as microfluidic tissue models.

Although microfluidics provides exquisite control of the cellular microenvironment, culturing cells within microfluidic devices can be challenging. 3D ...

Electrotaxis Studies of Lung Cancer Cells using a Multichannel Dual-electric-field Microfluidic Chip

The behavior of directional cell migration under a direct current electric-field (dcEF) is referred to as electrotaxis. The significant role of physiological dcEF in guiding cell movement during embryo development, cell differentiation, and wound healing has been demonstrated in many studies. By applying microfluidic chips to an electrotaxis assay, the investigation process is shortened and experimental errors are minimized. In recent years, microfluidic devices made of polymeric substances (e.g., polymethylmethacrylate, PMMA, or acrylic) or polydimethylsiloxane (PDMS) have been widely used in studying the responses of cells to electrical stimulation. However, unlike the numerous steps required to fabricate a PDMS device, the simple and rapid construction of the acrylic micro&#64258;uidic chip makes it suitable for both device prototyping and production. Yet none of the reported devices facilitate the efficient study of the simultaneous chemical and dcEF effects on cells. In this report, we describe our design and fabrication of an acrylic-based multichannel dual-electric-field (MDF) chip to investigate the concurrent effect of chemical and electrical stimulation on lung cancer cells. The MDF chip provides eight combinations of electrical/chemical stimulations in a single test. The chip not only greatly shortens the required experimental time but also increases accuracy in electrotaxis studies.

Many microfluidic devices have been developed for use in the study of electrotaxis. Yet, none of these chips allows the efficient study of the simultaneous chemical and electric-field (EF) effects on cells. We developed a polymethylmethacrylate-based device that offers better-controlled coexisting EF and chemical stimulation for use in electrotaxis research.

The behavior of directional cell migration under a direct current electric-field (dcEF) is referred to as electrotaxis. The significant role of ...

Using Adhesive Patterning to Construct 3D Paper Microfluidic Devices

We demonstrate the use of patterned aerosol adhesives to construct both planar and nonplanar 3D paper microfluidic devices. By spraying an aerosol adhesive through a metal stencil, the overall amount of adhesive used in assembling paper microfluidic devices can be significantly reduced. We show on a simple 4-layer planar paper microfluidic device that the optimal adhesive application technique and device construction style depends heavily on desired performance characteristics. By moderately increasing the overall area of a device, it is possible to dramatically decrease the wicking time and increase device success rates while also reducing the amount of adhesive required to keep the device together. Such adhesive application also causes the adhesive to form semi-permanent bonds instead of permanent bonds between paper layers, enabling single-use devices to be non-destructively disassembled after use. Nonplanar 3D origami devices also benefit from the semi-permanent bonds during folding, as it reduces the likelihood that unrelated faces may accidently stick together. Like planar devices, nonplanar structures see reduced wicking times with patterned adhesive application vs uniformly applied adhesive.

We demonstrate the use of patterned aerosol adhesives to construct 3D paper microfluidic devices. This method of adhesive application forms semi-permanent bonds between layers, enabling single-use devices to be non-destructively disassembled after use and to ease folding complex nonplanar structures.

We demonstrate the use of patterned aerosol adhesives to construct both planar and nonplanar 3D paper microfluidic devices. By spraying an aerosol ...

Multi-step Variable Height Photolithography for Valved Multilayer Microfluidic Devices

Microfluidic systems have enabled powerful new approaches to high-throughput biochemical and biological analysis. However, there remains a barrier to entry for non-specialists who would benefit greatly from the ability to develop their own microfluidic devices to address research questions. Particularly lacking has been the open dissemination of protocols related to photolithography, a key step in the development of a replica mold for the manufacture of polydimethylsiloxane (PDMS) devices. While the fabrication of single height silicon masters has been explored extensively in literature, fabrication steps for more complicated photolithography features necessary for many interesting device functionalities (such as feature rounding to make valve structures, multi-height single-mold patterning, or high aspect ratio definition) are often not explicitly outlined. 
Here, we provide a complete protocol for making multilayer microfluidic devices with valves and complex multi-height geometries, tunable for any application. These fabrication procedures are presented in the context of a microfluidic hydrogel bead synthesizer and demonstrate the production of droplets containing polyethylene glycol (PEG diacrylate) and a photoinitiator that can be polymerized into solid beads. This protocol and accompanying discussion provide a foundation of design principles and fabrication methods that enables development of a wide variety of microfluidic devices. The details included here should allow non-specialists to design and fabricate novel devices, thereby bringing a host of recently developed technologies to their most exciting applications in biological laboratories.

Multilayer microfluidic devices often involve the fabrication of master molds with complex geometries for functionality. This article presents a complete protocol for multi-step photolithography with valves and variable height features tunable to any application. As a demonstration, we fabricate a microfluidic droplet generator capable of producing hydrogel beads.

Microfluidic systems have enabled powerful new approaches to high-throughput biochemical and biological analysis. However, there remains a barrier to ...

Fabrication of Three-dimensional Paper-based Microfluidic Devices for Immunoassays

Paper wicks fluids autonomously due to capillary action. By patterning paper with hydrophobic barriers, the transport of fluids can be controlled and directed within a layer of paper. Moreover, stacking multiple layers of patterned paper creates sophisticated three-dimensional microfluidic networks that can support the development of analytical and bioanalytical assays. Paper-based microfluidic devices are inexpensive, portable, easy to use, and require no external equipment to operate. As a result, they hold great promise as a platform for point-of-care diagnostics. In order to properly evaluate the utility and analytical performance of paper-based devices, suitable methods must be developed to ensure their manufacture is reproducible and at a scale that is appropriate for laboratory settings. In this manuscript, a method to fabricate a general device architecture that can be used for paper-based immunoassays is described. We use a form of additive manufacturing (multi-layer lamination) to prepare devices that comprise multiple layers of patterned paper and patterned adhesive. In addition to demonstrating the proper use of these three-dimensional paper-based microfluidic devices with an immunoassay for human chorionic gonadotropin (hCG), errors in the manufacturing process that may result in device failures are discussed. We expect this approach to manufacturing paper-based devices will find broad utility in the development of analytical applications designed specifically for limited-resource settings.

We detail a method to fabricate three-dimensional paper-based microfluidic devices for use in the development of immunoassays. Our approach to device assembly is a type of multilayer, additive manufacturing. We demonstrate a sandwich immunoassay to provide representative results for these types of paper-based devices.

Paper wicks fluids autonomously due to capillary action. By patterning paper with hydrophobic barriers, the transport of fluids can be controlled and ...

Polydimethylsiloxane-polycarbonate Microfluidic Devices for Cell Migration Studies Under Perpendicular Chemical and Oxygen Gradients

This paper reports a microfluidic device made of polydimethylsiloxane (PDMS) with an embedded polycarbonate (PC) thin film to study cell migration under combinations of chemical and oxygen gradients. Both chemical and oxygen gradients can greatly affect cell migration in vivo; however, due to technical limitations, very little research has been performed to investigate their effects in vitro. The device developed in this research takes advantage of a series of serpentine-shaped channels to generate the desired chemical gradients and exploits a spatially confined chemical reaction method for oxygen gradient generation. The directions of the chemical and oxygen gradients are perpendicular to each other to enable straightforward migration result interpretation. In order to efficiently generate the oxygen gradients with minimal chemical consumption, the embedded PC thin film is utilized as a gas diffusion barrier. The developed microfluidic device can be actuated by syringe pumps and placed into a conventional cell incubator during cell migration experiments to allow for setup simplification and optimized cell culture conditions. In cell experiments, we used the device to study migrations of adenocarcinomic human alveolar basal epithelial cells, A549, under combinations of chemokine (stromal cell-derived factor, SDF-1&#945;) and oxygen gradients. The experimental results show that the device can stably generate perpendicular chemokine and oxygen gradients and is compatible with cells. The migration study results indicate that oxygen gradients may play an essential role in guiding cell migration, and cellular behavior under combinations of gradients cannot be predicted from those under single gradients. The device provides a powerful and practical tool for researchers to study interactions between chemical and oxygen gradients in cell culture, which can promote better cell migration studies in more in vivo-like microenvironments.

The control of chemical and oxygen gradients is essential for cell cultures. This paper reports a polydimethylsiloxane-polycarbonate (PDMS-PC) microfluidic device capable of reliably generating combinations of chemical and oxygen gradients for cell migration studies, which can be practically utilized in biological labs without sophisticated instrumentation.

This paper reports a microfluidic device made of polydimethylsiloxane (PDMS) with an embedded polycarbonate (PC) thin film to study cell migration under ...

A Simple and Scalable Fabrication Method for Organic Electronic Devices on Textiles

Today, wearable electronics devices combine a large variety of functional, stretchable, and flexible technologies. However, in many cases, these devices cannot be worn under everyday conditions. Therefore, textiles are commonly considered the best substrate to accommodate electronic devices in wearable use. In this paper, we describe how to selectively pattern organic electroactive materials on textiles from a solution in an easy and scalable manner. This versatile deposition technique enables the fabrication of wearable organic electronic devices on clothes.

In this paper, we present a protocol to selectively deposit organic materials on textiles, which allows for the direct integration of organic electronic devices with wearables. The fabricated devices can be fully integrated in textiles, respecting their mechanical appearance and enabling sensing capabilities.

Today, wearable electronics devices combine a large variety of functional, stretchable, and flexible technologies. However, in many cases, these devices ...

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.

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.

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