Name: the capillary feeder assay measures food intake in drosophila melanogaster
Uploaded: 2017-03-17T13:00:00
Description: Watch this Scientific Journal Video about The CApillary FEeder Assay Measures Food Intake in Drosophila melanogaster at JoVE.com

We thank the past and present members of the Scholz lab for discussion and Helga D&#246;ring for excellent technical support. We especially thank the members of the Biocenter workshop of the University of Cologne for their support and creativity. The work is supported by SFB 1340, SysMedAlc, and DAAD-Siemens.

1. The CAFE Assay
NOTE: The assay consists of three components: an experimental vial, a specific lid and micro- capillaries. A plastic box with cover is used to transport the prepared vials and to control the humidity more efficiently.
<ol>
	<li>Use a Drosophila melanogaster culture plastic vial (optional 8 cm height, 3.3 cm diameter) as a tube for the assay.</li>
	<li>Seal the vial with a manufactured Plexiglas lid containing an O-ring (Figures 1A, 1B). Load flies ...

<ol>
	<li>Krauth, C., Buser, J., Vogel, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=How+high+are+the+costs+of+eating+disorders+-+anorexia+nervosa+and+bulimia+nervosa+-+for+German+society.">How high are the costs of eating disorders - anorexia nervosa and bulimia nervosa - for German society.</a> Eur. J. Health Econ. 3 (4), 244-250 (2002).</li><li>Cawley, J., Meyerhoefer, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+medical+costs+of+obesity+and+instrumental+variables+approach.">The medical costs of obesity and instrumental variables approach.</a> J. Health Econ. 31, 219-230 (2012).</li><li>The costs of eating disorders: Social, health and economic impacts. Assessing the impact of eating disorders across the UK on behalf of BEAT. PwC Available from: <a target="_blank" href="https://www.beat.co.uk/assets/000/000/302/The_costs_of_eating_disorders_Final_original.pdf">https://www.beat.co.uk/assets/000/000/302/The_costs_of_eating_disorders_Final_original.pdf</a> (2015)</li><li>Lenard, N. R., Berthoud, H. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Central+and+peripheral+regulation+of+food+intake+and+physical+activity:+pathways+and+genes.">Central and peripheral regulation of food intake and physical activity: pathways and genes.</a> Obesity. 16, S11-S22 (2008).</li><li>Magni, P., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Feeding+behavior+in+mammals+including+humans.">Feeding behavior in mammals including humans.</a> Trends in Comp. Endocrinology and Neurobiology. 1163, 221-232 (2009).</li><li>Morton, G. J., Meek, T. H., Schwartz, M. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neurobiology+of+food+intake+in+health+and+disease.">Neurobiology of food intake in health and disease.</a> Nature Reviews Neuroscience. 15, 367-378 (2014).</li><li>Bharuchka, K. N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+epicurean+fly:+using+Drosophila+melanogaster+to+study+metabolism.">The epicurean fly: using Drosophila melanogaster to study metabolism.</a> Pediatr. Res. 65 (2), 132-137 (2009).</li><li>Smith, W. W., Thomas, J., Liu, J., Li, T., Moran, T. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=From+fat+fruit+fly+to+human+obesity.">From fat fruit fly to human obesity.</a> Physiol. Behav. 136, 15-21 (2014).</li><li>Rajan, A., Perrimon, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Of+flies+and+men:+insights+on+organismal+metabolism+from+fruit+flies.">Of flies and men: insights on organismal metabolism from fruit flies.</a> BMC Biology. 11, (2013).</li><li>Ja, W. W., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Prandiology+of+Drosophila+and+the+CAFE+assay.">Prandiology of Drosophila and the CAFE assay.</a> Proc. Natl. Acad. Sci. USA. 104 (20), 8253-8256 (2007).</li><li>Dethier, V. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> The Hungry Fly: A Physiological Study of the Behavior Associated with Feeding. , (1976).</li><li>Albin, S. D., Kaun, K. R., Knapp, J., Chung, P., Heberlein, U., Simpson, 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=A+subset+of+serotonergic+neurons+evokes+hunger+in+adult+Drosophila.">A subset of serotonergic neurons evokes hunger in adult Drosophila.</a> Curr. Biol. 25, 2435-2440 (2015).</li><li>Deshpande, S. A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Quantifying+Drosophila+food+intake:+comparative+analysis+of+current+methodology.">Quantifying Drosophila food intake: comparative analysis of current methodology.</a> Nat. Methods. 11 (5), 535-540 (2014).</li><li>Geer, B. W., Olander, R. M., Sharp, P. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Quantification+of+dietary+choline+utilization+in+adult+Drosophila+melanogaster+by+radioisotope+methods.">Quantification of dietary choline utilization in adult Drosophila melanogaster by radioisotope methods.</a> J. Insect Physiol. 16, 33-43 (1970).</li><li>Thompson, E. D., Reeder, B. A., Bruce, 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=Characterization+of+a+method+for+quantitating+food+consumption+for+mutation+assays+in+Drosophila.">Characterization of a method for quantitating food consumption for mutation assays in Drosophila.</a> Environ. Mol. Mutagen. 18, 14-21 (1991).</li><li>Wong, R., Piper, M. D., Wertheim, B., Partridge, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Quantification+of+food+intake+in+Drosophila.">Quantification of food intake in Drosophila.</a> PLoS One. 4 (6), e6063 (2009).</li><li>Scheiner, R., Steinbach, A., Classen, G., Strudthoff, N., Scholz, H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Octopamine+indirectly+affects+proboscis+extension+response+habituation+in+Drosophila+melanogaster+by+controlling+sucrose+responsiveness.">Octopamine indirectly affects proboscis extension response habituation in Drosophila melanogaster by controlling sucrose responsiveness.</a> J. Insect Physiol. 69, 107-117 (2014).</li><li>Liu, Y., Luo, J., Carlsson, M. K., N&#228;ssel, D. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Serotonin+and+insulin-like+peptides+modulate+leucokinin-producing+neurons+that+affect+feeding+and+water+homeostasis+in+Drosophila.">Serotonin and insulin-like peptides modulate leucokinin-producing neurons that affect feeding and water homeostasis in Drosophila.</a> J. Comp. Neurol. 523, 1840-1863 (2015).</li><li>Ro, J., Harvanek, Z. M., Pletcher, S. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=FLIC:+high-throughput,+continuous+analysis+of+feeding+behaviors+in+Drosophila.">FLIC: high-throughput, continuous analysis of feeding behaviors in Drosophila.</a> PLoS One. 9 (6), e101107 (2014).</li><li>Itskov, P. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Automated+monitoring+and+quantitative+analysis+of+feeding+behavior+in+Drosophila.">Automated monitoring and quantitative analysis of feeding behavior in Drosophila.</a> Nat. Commun. 5, 4560 (2014).</li><li>Qi, W., Yang, Z., Lin, Z., Park, J. Y., Suh, G. S. B., Wang, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+quantitative+feeding+assay+in+adult+Drosophila+reveals+rapid+modulation+of+food+ingestion+by+its+nutritional+value.">A quantitative feeding assay in adult Drosophila reveals rapid modulation of food ingestion by its nutritional value.</a> Mol. Brain. 8, 87 (2015).</li><li>Marx, V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Metabolism:+feeding+fruit+flies.">Metabolism: feeding fruit flies.</a> Nat. Methods. 12 (7), 609-612 (2015).</li><li>Spieth, H. T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Courtship+behavior+in+Drosophila.">Courtship behavior in Drosophila.</a> Annu. Rev. Entomol. 19, 385-405 (1974).</li><li>Devineni, A. V., Heberlein, U. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Preferential+ethanol+consumption+in+Drosophila+models+features+of+addiction.">Preferential ethanol consumption in Drosophila models features of addiction.</a> Curr. Biol. 19 (24), 2126-2132 (2009).</li><li>Lee, K. P., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Lifespan+and+reproduction+in+Drosophila:+New+insights+from+nutritional+geometry.">Lifespan and reproduction in Drosophila: New insights from nutritional geometry.</a> Proc. Natl. Acad. Sci. USA. 105 (7), 2498-2503 (2008).</li><li>Pohl, J. B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ethanol+preference+in+Drosophila+melanogaster+is+driven+by+its+caloric+value.">Ethanol preference in Drosophila melanogaster is driven by its caloric value.</a> Alcohol Clin. Exp. Res. 36 (11), 1903-1912 (2012).</li><li>Vargas, M. A., Luo, N., Yamaguchi, A., Kapahi, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+role+for+S6+kinase+and+serotonin+in+postmating+dietary+switch+and+balance+of+nutrients+in+D.+melanogaster.">A role for S6 kinase and serotonin in postmating dietary switch and balance of nutrients in D. melanogaster.</a> Curr. Biol. 20 (11), 1006-1011 (2010).</li><li>Masek, P., Scott, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Limited+taste+discrimination+in+Drosophila.">Limited taste discrimination in Drosophila.</a> Proc. Natl. Acad. Sci. 107 (33), 14833-14838 (2010).</li><li>Pool, A. H., Scott, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Feeding+regulation+in+Drosophila.">Feeding regulation in Drosophila.</a> Curr. Opin. Neurobiol. 29, 57-63 (2014).</li><li>Luo, J. N., Lushchak, O. V., Goergen, P., Williams, M. J., N&#228;ssel, D. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Drosophila+insulin-producing+cells+are+differentially+modulated+by+serotonin+and+octopamine+receptors+and+affect+social+behavior.">Drosophila insulin-producing cells are differentially modulated by serotonin and octopamine receptors and affect social behavior.</a> Plos One. 9 (6), e99732 (2014).</li></ol>

The report describes the CAFE assay in a step-by-step fashion, focusing on the technical setup and its successful performance in the laboratory. Due to its simplicity, this assay could also be used educationally as a school experiment. The examples show that the assay allows investigation of food sensing, preference and consumption in Drosophila melanogaster over short and longer time periods (hours to days). The CAFE assay has been used widely in the field to investigate subjects including food and drug consump...

Eating is essential; however, deregulation of food intake resulting in eating disorders such as bulimia, anorexia or the general tendency to overeat imposes costs on individuals and society1,2,3. The goal of the present research is to uncover regulatory mechanisms of food intake and to provide a strategy for circumventing disorder formation. Numerous studies using mammalian model organisms have provided new insights of the circuitry and the role of signaling systems in eating disorders4,5,6. Nevertheless, our knowledge of the neuronal and molecular bases underlying these disorders remains far from complete. In recent years, the fruit fly Drosophila melanogaster has become a valuable model system for unraveling basic mechanistic insights into the regulation of metabolism7,8,9. The CApillary FEeder (CAFE) assay for Drosophila melanogaster was established in the lab of Seymour Benzer in 2007 inspired by an earlier work by Dethier in blowfly10, 11. The CAFE assay made it possible to directly measure food intake in Drosophila melanogaster. In this behavioral test system, flies feed on liquid food provided in graded glass capillaries placed inside a vial. The decline of the capillary meniscus indicates loss of food solution via evaporation and food consumption. Determining the evaporation rate by vials without flies allows the accurate quantification of food intake.
The CAFE assay is one of several behavioral paradigms used to measure feeding in Drosophila melanogaster and researchers have to choose the most appropriate one for their specific question. The decision to use a certain assay should consider the following points: the nature of the food provided; the feeding condition; the measurement of intake or uptake of nutrients and investigation food consumption or response to food.
The CAFE assay as described in this report is ideal for following food intake of a liquid food source under an upright feeding condition. Alternatively the food intake can be measured for a fly group on a colored food source in a vial or on a plate. Flies are normally killed or anesthetized after feeding and the amount of ingested dye is determined by spectrometry or visual inspection of the stained abdomen. Flies start to excrete the ingested food only 30 min after intake, therefore this approach is difficult to use for the analysis of continuous longer feeding behaviors12, 13.
In contrast flies are kept intact when absorbable dyes with radioactive tracers are used and their consumption of radioisotope is scored in a scintillation counter14, 15. Absorption of the radiolabel by the fly digestive system makes long-term food uptake measurement possible, but might lead to underestimation of consumption because of non-absorbed and excreted tracer molecules. Another approach to measure response to food in Drosophila melanogaster is the proboscis extension response (PER), which normally occurs for food intake16. This elegant method measures the initial response to a food stimulus but does not record the quantity of intake. Food intake is dynamically adjusted during feeding using several post-digestive feedback signals that are critical for the regulation of feeding17, 18. Several attempts have been made in recent years to semi-automate data collection in the PER assay19, 20. The PER is detected by an electric pad or a combination of electrodes and counted via computer. Combining the PER assay with radioisotope uptake revealed that this assay is limited by low sensitivity to detecting quantity feeding differences18. The manual feeding assay (MAFE)21, in which a fly is fed manually with a glass capillary, was recently developed to measure food uptake in a single immobilized fly. The MAFE assay eliminates the interferences of foraging and feeding initiation and has a time resolution of seconds, and initiation of PER and food consumption can be monitored independently in the assay. However, the way in which immobilization of the fly affects certain aspects of feeding behavior (e.g. locomotion, motivation) still needs to be investigated. For excellent comparative reviews of different assays for measuring food consumption in Drosophila melanogaster and to help researchers finding the most appropriate one, see reports by Deshpande and Marx 13, 22.
The CAFE assay avoids some of the disadvantages of other assays described above and combines simplicity of use with reliable measurement of food intake. Here, a detailed description of the CAFE assay is provided and we show a simple setup modification to reduce evaporation. Representative results including a two food choice assay (short and long term) and the sucrose uptake of flies is demonstrated. In the discussion we compare our described method with alternative ways to perform the CAFE assay, and highlight potential limitations.

<table border="0" cellpadding="0" cellspacing="0" width="582">
	<tbody>
		<tr height="31">
			<td height="31" style="height:31px;width:161px;">Vials (breeding)</td>
			<td style="width:117px;">Greiner Bio-One</td>
			<td style="width:123px;">960177</td>
			<td style="width:183px;">www.greinerbioone.com</td>
		</tr>
		<tr height="31">
			<td height="31" style="height:31px;">Vials (CAFE assay)</td>
			<td>Greiner Bio-One</td>
			<td>217101</td>
			<td>www.greinerbioone.com</td>
		</tr>
		<tr height="31">
			<td height="31" style="height:31px;">Lid-CAFE assay</td>
			<td>Workshop</td>
			<td>&#8211;</td>
			<td>&#8211;</td>
		</tr>
		<tr height="31">
			<td height="31" style="height:31px;">Plastic box, low wall</td>
			<td>Plastime</td>
			<td>353</td>
			<td>www.plastime.it</td>
		</tr>
		<tr height="31">
			<td height="31" style="height:31px;">Cover for the plastic box</td>
			<td>Workshop</td>
			<td>&#8211;</td>
			<td>&#8211;</td>
		</tr>
		<tr height="31">
			<td height="31" style="height:31px;">Capillaries</td>
			<td>BLAUBRAND&#160;</td>
			<td>REF 7087 07</td>
			<td>www.brand.de</td>
		</tr>
		<tr height="31">
			<td height="31" style="height:31px;">Pipette tips</td>
			<td>Greiner Bio-One</td>
			<td>771290</td>
			<td>www.greinerbioone.com</td>
		</tr>
		<tr height="31">
			<td height="31" style="height:31px;">Filter paper circles</td>
			<td>Whatman</td>
			<td>10 311 804</td>
			<td>www.sigmaaldrich.com</td>
		</tr>
		<tr height="31">
			<td height="31" style="height:31px;">D(+)-Sucrose</td>
			<td>AppliChem</td>
			<td>57-50-1</td>
			<td>www.applichem.com</td>
		</tr>
		<tr height="31">
			<td height="31" style="height:31px;">Ethanol absolute</td>
			<td>VWR Chemicals</td>
			<td>20,821,330</td>
			<td>www.vwr.com</td>
		</tr>
		<tr height="31">
			<td height="31" style="height:31px;">Food color (red, E124)</td>
			<td>Backfun</td>
			<td>10027</td>
			<td>www.backfun.de</td>
		</tr>
		<tr height="31">
			<td height="31" style="height:31px;">Food color (blue, E133)</td>
			<td>Backfun</td>
			<td>10030</td>
			<td>www.backfun.de</td>
		</tr>
		<tr height="31">
			<td height="31" style="height:31px;">Soap solution (CVK 8)</td>
			<td>CVH</td>
			<td>103220</td>
			<td>www.cvh.de</td>
		</tr>
		<tr height="31">
			<td height="31" style="height:31px;">Digital caliper</td>
			<td>GARANT</td>
			<td>412,616</td>
			<td>www.hoffmann-group.com</td>
		</tr>
		<tr height="31">
			<td height="31" style="height:31px;">Vials (breeding)</td>
			<td></td>
			<td></td>
			<td>Height 9.8 cm, diameter 4.8 cm&#160;</td>
		</tr>
		<tr height="31">
			<td height="31" style="height:31px;">Vials (CAFE assay)</td>
			<td></td>
			<td></td>
			<td>Height 8 cm, diameter 3.3 cm</td>
		</tr>
		<tr height="31">
			<td height="31" style="height:31px;">Lid-CAFE assay</td>
			<td></td>
			<td></td>
			<td>Produced in university workshop, technical drawing supplied</td>
		</tr>
		<tr height="31">
			<td height="31" style="height:31px;">Plastic box, low wall</td>
			<td></td>
			<td></td>
			<td>A plastic grid inlay was custom-made for 8 x 10 vial positions&#160;</td>
		</tr>
		<tr height="31">
			<td height="31" style="height:31px;">Cover for the plastic box</td>
			<td></td>
			<td></td>
			<td>Dimensions (37 x29 x18 cm)</td>
		</tr>
		<tr height="31">
			<td height="31" style="height:31px;">Capillaries</td>
			<td></td>
			<td></td>
			<td>DIN ISO 7550 norm,&#160; IVD-guideline 98/79 EG, ends polished</td>
		</tr>
		<tr height="31">
			<td height="31" style="height:31px;">Pipette tips</td>
			<td></td>
			<td></td>
			<td>Pipettes for the outer circle are cut according to the lid</td>
		</tr>
		<tr height="31">
			<td height="31" style="height:31px;">Filter paper circles</td>
			<td></td>
			<td></td>
			<td>45 mm diameter works nicely if folded for the vials used</td>
		</tr>
		<tr height="31">
			<td height="31" style="height:31px;">D(+)-Sucrose</td>
			<td></td>
			<td></td>
			<td>Not harmful</td>
		</tr>
		<tr height="31">
			<td height="31" style="height:31px;">Ethanol absolute</td>
			<td></td>
			<td></td>
			<td>Highly flammable liquid and vapor</td>
		</tr>
		<tr height="31">
			<td height="31" style="height:31px;">Food color (red, E124)</td>
			<td></td>
			<td></td>
			<td>Not stated</td>
		</tr>
		<tr height="31">
			<td height="31" style="height:31px;">Food color (blue, E133)</td>
			<td></td>
			<td></td>
			<td>Not stated</td>
		</tr>
		<tr height="31">
			<td height="31" style="height:31px;">Soap solution (CVK 8)</td>
			<td></td>
			<td></td>
			<td>Odor neutral soap</td>
		</tr>
		<tr height="31">
			<td height="31" style="height:31px;">Digital caliper</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr height="31">
			<td height="31" style="height:31px;">Standard fly food</td>
			<td></td>
			<td></td>
			<td>(for 20 L)</td>
		</tr>
		<tr height="31">
			<td height="31" style="height:31px;">Agar</td>
			<td></td>
			<td></td>
			<td>160 g</td>
		</tr>
		<tr height="31">
			<td height="31" style="height:31px;">Brewer`s Yeast</td>
			<td></td>
			<td></td>
			<td>299.33 g</td>
		</tr>
		<tr height="31">
			<td height="31" style="height:31px;">Cornmeal</td>
			<td></td>
			<td></td>
			<td>1200g</td>
		</tr>
		<tr height="31">
			<td height="31" style="height:31px;">Molasses</td>
			<td></td>
			<td></td>
			<td>1.6 L</td>
		</tr>
		<tr height="31">
			<td height="31" style="height:31px;">Propionic acid</td>
			<td></td>
			<td></td>
			<td>57.3 mL</td>
		</tr>
		<tr height="31">
			<td height="31" style="height:31px;">Nipagin 30%</td>
			<td></td>
			<td></td>
			<td>160 mL</td>
		</tr>
	</tbody>
</table>

the capillary feeder assay measures food intake in drosophila melanogaster

For most animals, feeding is an essential behavior for securing survival, and it influences development, locomotion, health and reproduction. Ingestion of the right type and quantity of food therefore has a major influence on quality of life. Research on feeding behavior focuses on the underlying processes that ensure actual feeding and unravels the role of factors regulating internal energy homeostasis and the neuronal bases of decision-making. The model organism Drosophila melanogaster, with its great variety of genetically traceable tools for labeling and manipulating single neurons, allows mapping of neuronal networks and identification of molecular signaling cascades involved in the regulation of food intake. This report demonstrates the CApillary FEeder assay (CAFE) and shows how to measure food intake in a group of flies for time spans ranging from hours to days. This easy-to-use assay consists of glass capillaries filled with liquid food that flies can freely access and feed on. Food consumption in the assay is accurately determined using simple measurement tools. Herein we describe step-by-step the method from setup to successful execution of the CAFE assay, and provide practical examples to analyze the food intake of a group of flies under controlled conditions. The reader is guided through possible limitations of the assay, and advantages and disadvantages of the method compared to other feeding assays in D. melanogaster are evaluated.

Flies of the w1118 genotype are used to demonstrate how the assay is performed. The w1118 mutants are commonly used to generate transgenic lines and to control the genetic background of transgenes marked with the white gene. Normally, for behavioral experiments, all transgenic lines are backcrossed for five generations to the same w1118 stock, which is used as an experimental control. We show different experiments: a com...

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The CApillary FEeder Assay Measures Food Intake in Drosophila melanogaster

The CApillary FEeder (CAFE) assay is a simple, budget-friendly, highly reliable method for investigating mechanisms underlying food intake. Used with the highly versatile genetic model organism Drosophila melanogaster, it provides a powerful means of gaining new insights into regulatory mechanisms of food intake.

The CApillary FEeder Assay Measures Food Intake in Drosophila melanogaster

For most animals, feeding is an essential behavior for securing survival, and it influences development, locomotion, health and reproduction. Ingestion of ...

The overall goal of the CApillary FEeder assay is to observe and measure liquid food intake of adult flies. This method can help to answer key questions in metabolic research such as how actual feeding behavior is initiated and how food consumption is terminated. The main advantages of this technique are that it's easy to use and highly reliable.Visual demonstration of the method is critical as several preparatory steps need to be carefully executed for consistent results. To begin the experiment, obtain a drosophila melanogaster culture plastic vial and seal it with a manufactured Plexiglass lid containing an O-ring. Use five microliter microcapillaries with one microliter marks to present the liquid food.Cut off the top of a two to 20 microliter pipette tip and insert the tip into the whole of the lid to position the capillaries in the conical openings. Insert an uncut two to 20 microliter pipette tip into the same opening to prevent flies from escaping. Load eight or 20 flies by trapping or with a blow pipe through the lid's central opening and close the hole with a sponge bung.Next, place the multiple prepared vials into a plastic box with a gridded inlay to handle them safely. To control breeding conditions, introduce 35 virgin females and 15 males for each experimental group into a plastic culture vial containing 50 milliliter fly food. Allow the flies to lay eggs for the first three days, then transfer adult flies to fresh food vials and let them lay eggs for two more days.After two days, repeat the transfer. On day seven, discard the adult flies. Next, anesthetize two to three day old adult flies using a CO2 pad and collect 100 flies sorted by sex in a 1.5 milliliter plastic tube.Measure their weight with a standard laboratory scale. Determine the wet weight of at least four independent fly groups sorted by sex and use the weight to calculate the microliter food consumption per milligram fly. For a three hour assay, use 20 flies in two filled capillaries.For a long term experiment, use a group of eight flies with a supply of four filled capillaries. Fill up to 10 capillaries simultaneously with previously prepared colored food solution. Insert the capillaries into the food solution and then tilt them to a 45 degree angle to accelerate filling.Stop when the liquid reaches the top five microliter mark of the capillary. Remove excess solution on the outside and inside of the capillary with tissue paper. Carefully remove the pipette tip that is closing one of the outer openings and insert a filled glass capillary, bottom end first.Secure the capillary by placing the pipette tip back next to the capillary. Repeat this step for all food solutions tested. Next, place the capillary ends inside all of the vials at the same level three to four centimeters from the lid.Leave some distance between the filter paper to prevent the capillary from leaking. Use a marker pen to label the upper end of the colored liquid. Label them individually using a stripe code to ensure the different capillaries can be identified.Place prepared CAFE assays into a plastic box with a gridded inlay. Use a sponge bung to apply 100 microliters of fresh water every 24 hours to keep the humidity constant inside the assay. Use four separate vials filled with 30 milliliters of double distilled H2O as humidity devices and place them next to the CAFE assays in the plastic box.Cover the plastic box to create a humidity controlled environment during the experiment. Transfer the box to a secure position in a temperature, light, and humidity controlled climate chamber for the experimental period. Before each 24 hour interval, make note of the dead flies and use the number of live flies to calculate the consumption per fly for the following period.At the end of the assay and before replacing the capillary, keep the CAFE assay in the upright position. Do not remove the lid and mark the lower meniscus of the capillary. Discard the data if the mark end is not below the initial mark.Replace the used capillaries with freshly filled capillaries for long term experiments every 24 hours. Measure the decline of the meniscus, then discard the old capillaries. Then carefully remove the capillaries from the assay and store them for data collection.Check if the liquid inside the capillary reached the lower end. If the liquid inside the capillary did not reach the lower end, discard the data. After the experiment, dismantle the setup and wash the vials, lids, and sponge bungs in a soap bath.Then dry them overnight at room temperature for further use. Repeat experiments with the same genotypes on at least three different days. Using a caliper, measure the distance between mark beginning and mark end on the capillary.Finally, discard the capillaries after the measurements. Two food choices were offered, a 1 molar sucrose solution and a 1 molar sucrose solution with 15%ethanol. Visual examination of the abdomen indicates the flies feed on both solutions.However food consumption per fly is nearly two folds greater for the solution containing ethanol. The ethanol preference was also observed in long term experiments where a group of eight flies had access to similar food sources for four days. The CApillary FEeder assay also reveals that male and female flies prefer higher sucrose concentrations over lower sucrose concentrations.And male flies consume significantly more of the high sucrose concentrations than their female counterparts. While attempting this procedure, it's important to remember to perform the assay in a controlled fashion and to repeat the experiment with the same genotypes on at least three different days. In addition, when testing mutant flies, it's important to consider whether the flies can actually perform the task required to obtain the food, such as sending the food source or climbing up and down the capillary.

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