We thank Colby Carpenter and Spencer Mathias for their assistance in 3D printing design. We thank Ellen Klinger for assistance in producing the photographic figures and Jonathan B. Koch for providing assistance with revisions. Funding was provided by the USDA-ARS-Pollinating Insect Biology, Management, and Systematics Research Unit.

1. Print pollen trap structures
<ol>
	<li>Download the appropriate STL file for the nest box that bumble bees are nesting in (e.g., Biobest or Koppert style hives, https://www.ars.usda.gov/pacific-west-area/logan-ut/pollinating-insect-biology-management-systematics-research/docs/pollen-traps/). The files are available to the public, free for download and modification by the end user.</li>
	<li>Open the STL file in the printer program. Follow printer manufacturer directions to build the four trap components. 
	NOTE: Allow approximately 3 h for the trap body to print, 2 h for catch basin to print and 30 min each for the filter and trap closure insert to print. Trap body size is 6 cm x 3.8 cm x 7 cm.</li>
</ol>
2. Pollen trap assembly
<ol>
	<li>Remove the support structures printed with the trap body and catch basin including those of the sieve structure of the trap body (Figure 1).</li>
	<li>Use a 3/16 in. (0.476 cm) drill bit mounted in a hand drill to clear any plastic strands crossing the raised edges of the pollen filter that might hinder a bee from moving through the filter holes. Use a box cutting razor blade and sandpaper to even out any bumps or raised edges on the flat side of the pollen filter.</li>
	<li>Place the pollen filter in the trap body by gently pushing the plastic filter through one side of the trap body. The filter will only fit in one way, as the left side of the trap has a larger opening to accommodate passage of the raised filter cones.
	<ol>
		<li>If the side slits are too small for the pollen filter to slide through smoothly, scrape enough plastic away from the slit in the trap body with a razor or other tool that can remove small portions of plastic at a time. Ensure that the pollen filter fits securely in place with no more than a 2 mm gap between the pollen filter and trap body.</li>
	</ol>
	</li>
	<li>Attach the catch basin to the trap body by sliding the raised edges on the bottom of the trap body into the groove on the top of the catch basin. The catch basin should be positioned directly underneath the sieve region of the trap body (Figure 1A&#8210;E).</li>
	<li>Make appropriate modifications by cutting or sanding the plastic to allow smooth placement and removal of the catch basin from the trap body. Placement of the catch basin will secure the pollen filter into place and it cannot be removed until the catch basin is removed, nor can the pollen filter be inserted while the catch basin is in place. 
	NOTE: If multiple hives are placed close to one another, providing each hive with a unique color combination of the trap body, catch basin and pollen filter structures in addition to deploying hives with varied orientation will help returning workers find their nests.</li>
</ol>
3. Bumble bee colony preparation
<ol>
	<li>Stop pollen feeding 24&#8210;48 h before deploying colonies. This will cause workers to use up any stored pollen and stimulate them to leave the nest in search of pollen.</li>
	<li>Prepare the hive trap body for installation by inserting a trap closure insert into the filter slot to prevent bees from escaping while installing the trap.</li>
	<li>Working under red light to prevent the bees from flying, lift the plastic nest box out of the cardboard outer box.</li>
	<li>Locate the plastic nest entrance at the front of the nest. There are two styles of entrance depending on the nest supplier: Koppert-style boxes (step 3.5) and Biobest-style boxes (step 3.6).</li>
	<li>For Koppert-style hive entrances, mount the pollen trap onto the nest entrance by pulling up on the entrance tab until both entrance holes are open.
	<ol>
		<li>Insert the two tubes of the pollen trap into the entrance holes, ensuring that the sieve of the pollen trap is on the bottom. Gently push down on the plastic entrance tab to secure the pollen trap in place.</li>
	</ol>
	</li>
	<li>To install the pollen trap into Biobest-style hive entrances, use a flat head screwdriver to gently pry the plastic entrance device from the nest box. Insert the pollen trap into the nest entrance holes until the pollen trap is firmly against the nest (Figure 1E).
	<ol>
		<li>Secure the trap to the nest using tape or quick dry glue where the trap contacts the nest box if needed.</li>
	</ol>
	</li>
	<li>Return the plastic nest box to the cardboard box. Cardboard may need to be cut away to accommodate the pollen trap.</li>
</ol>
4. Deployment of nests
<ol>
	<li>Place nest boxes into the study area. Provide cover to protect from precipitation and anchorage for wind as these can adversely affect the quality and quantity of pollen that is collected. Provide hives placed in greenhouses with adequate sun cover to reduce overheating.</li>
	<li>Remove the trap closure insert from the trap body to allow bees to forage freely so that foragers can orient themselves with the surrounding area and the location of their nest. Orientation flight time should be complete in 24 h under normal conditions.</li>
</ol>
5. Pollen collection
<ol>
	<li>To engage the trap, slide the pollen filter into the filter slot, ensuring that it is securely in place.</li>
	<li>Install the catch basin by sliding it onto the trap body from the front until it is fully closed. If the catch basin is excessively loose or falls off the trap body, use a rubber band to secure it to the trap body.</li>
	<li>Observe bees entering and exiting the pollen trap at first deployment to ensure the pollen filter holes are large enough to accommodate the bees.
	<ol>
		<li>If workers are unable to pass through the pollen filter, remove the filter and then use drill bits larger than 3/16 in. (0.476 cm) to increase the holes sizes. Do so in a sequential manner, increasing hole diameter 1/32 in. (0.079 cm) each time, as holes that are too large will not collect any pollen.</li>
		<li>Once bees are able to pass through the filter, continue to observe the entrance to ensure pollen is being removed upon re-entry. 
		NOTE: Bees should have some difficulty moving though the pollen filter holes, especially the first few times they pass through. If they pass through too easily, the pollen may not be dislodged from the corbiculae.</li>
	</ol>
	</li>
	<li>After the designated period of pollen collection, slide and remove catch basin from the trap body.</li>
	<li>Process the pollen loads according to your experimental design.</li>
	<li>Remove the pollen filter to allow workers to forage freely until the next period of pollen collection. The trap body may remain attached to the hive for the duration of the experiment. 
	NOTE: Pollen traps may be engaged for as long as the researcher desires to collect pollen from a colony. However, deployment of pollen traps for over 24 h in a week may result in starvation of brood in a hive and retard colony development.</li>
</ol>

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The authors have nothing to disclose.

Collection of pollen from bumble bee colony entrances can allow for a variety of ecological and agricultural studies. Identifying the floral sources from which bumble bees collect pollen provides valuable information and insight into the diversity of plants that contribute to a colony&#8217;s overall diet19. Identifying the pollen source has implications for both agricultural production and studies of ecosystem services in wild lands12,20....

Bumble bees (Bombus spp.) are large robust insects that are found across the temperate, alpine, and arctic regions of the world1. They are important to&#160;plant communities&#160;and provide important pollination service for the agricultural crops that they visit2. Recent declines in the&#160;abundance and distribution of several species has brought their importance as pollinators to the forefront of&#160;public&#160;awareness3. Researchers have identified several stressors that are likely contributing to population&#160;declines including a lack of diverse and abundant floral resources on which bumble bees forage4. Identifying which&#160;plant species bumble bees forage from allows researchers and land managers to understand how bumble bees may be responding to changes in resource availability, competition, and&#160;anthropogenic disturbances5,6.
Studies investigating the pollen foraging preferences of bumble bees are often conducted by researchers catching individual bees foraging at flowers, and then removing the corbicular pollen loads from specimens for further processing and identification7,8,9,10. While this method provides insight into how a species or an assemblage of bumble bee species utilizes the resources in an area7, it is time intensive and potential differences in preferences among hives cannot be discerned without additional molecular analyses to identify colony of origin of the foraging bee11.
For some studies of foraging dynamics, it is desired to conduct the studies at individual colonies; however, wild bumble bee nests are generally located underground or at ground level making them difficult to locate12. Commercially produced bumble bee hives provide researchers greater access and better experimental control and the removal of pollen off workers is still primarily conducted by capturing foragers as they return to the hive and manually removing their corbicular pollen loads13,14. The removal of pollen by hand from the corbicula of a bee is time intensive with a low hourly yield of pollen especially at hive entrances where the rate of returning pollen foragers may be low. Additionally, manually removing pollen from bees can result in stings from disturbed workers.
Pollen traps have been used for experimental removal of pollen from honey bees for decades15; yet, a passive method for removing pollen from bumble bees has not been developed. The primary obstacle in developing a mechanism to remove pollen from returning forager bumble bees is the large variation of worker sizes that exist in a bumble bee colony16. Honey bee pollen traps are effective largely because honey bee worker size does not vary much. Additionally, these traps require only minor manipulations after installation and don&#8217;t require bees to be sacrificed17. This is achieved using screens or plastic surfaces that dislodge the pollen off of the hind legs of workers as they return to the hive. These traps remove only a portion of the pollen loads from returning foragers and the various designs of those result in varied efficiencies at pollen collection. As the pollen is removed from the bee legs, it falls through a screen and into a collection basin to which the bees have no access, so that the researcher can remove it with only minor disturbance to the hive.
The purpose of the present study is to adapt the techniques used for collecting pollen from honey bee hives and apply them to bumble bee nests using 3D printed structures and test the trap designs on colonies of Bombus huntii. The design process followed the assumptions that the traps should be inexpensive to produce, adaptable to a variety of bumble bee species, cause minimal harm or disturbance to the bees, and that the rate of pollen removal should exceed hand collection of pollen. Three-dimensional printing technology is versatile, easily accessible, and a cost-effective tool allowing researchers to replicate and modify objects for specific purposes18. The technique presented here instructs the user to build pollen traps and attach them onto commercially available bumble bee colonies. The traps are not designed to be use with wild colonies. These traps passively remove the corbicular pollen loads from the hind legs of pollen carrying bumble bees as they return to their nest boxes.

<table border="0" cellpadding="0" cellspacing="0" style="width:461px;" width="462">
	<colgroup>
		<col />
		<col span="2" />
		<col />
	</colgroup>
	<tbody>
		<tr height="35">
			<td height="35" style="height:35px;width:71px;">MakerBot Replicator+</td>
			<td style="width:87px;">MakerBot</td>
			<td style="width:87px;">Model PABH65</td>
			<td style="width:217px;"></td>
		</tr>
		<tr height="52">
			<td height="52" style="height:52px;width:71px;">MakerBot Tough Material</td>
			<td style="width:87px;">PLA Filament</td>
			<td style="width:87px;"></td>
			<td>various colors</td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:71px;">Nest Box</td>
			<td style="width:87px;">Biobest</td>
			<td style="width:87px;"></td>
			<td>Not sold publicly without bee purchase</td>
		</tr>
	</tbody>
</table>

a 3d printed pollen trap for bumble bee (bombus) hive entrances

To verify the plant sources from which bumble bees forage for pollen, individuals must be collected to remove their corbicular pollen loads for analysis. This has traditionally been done by netting foragers at nest entrances or on flowers, chilling the bees on ice, and then removing the pollen loads from the corbiculae with forceps or a brush. This method is time and labor intensive, may alter normal foraging behavior, and can result in stinging incidents for the worker performing the task. Pollen traps, such as those used on honey bee hives, collect pollen by dislodging corbicular pollen loads from the legs of workers as they pass through screens at the nest entrance. Traps can remove a large quantity of pollen from returning forager bees with minimal labor, yet to date no such trap is available for use with bumble bee colonies. Workers within a bumble bee colony can vary in size making size selection of entrances difficult to adapt this mechanism to commercially reared bumble bee hives. Using 3D printing design programs, we created a pollen trap that successfully removes the corbicular pollen loads from the legs of returning bumble bee foragers. This method significantly reduces the amount of time required by researchers to collect pollen from bumble bee foragers returning to the colony. We present the design, results of pollen removal efficiency tests, and suggest areas of modifications for investigators to adapt traps to a variety of bumble bee species or nest box designs.

Eight different pollen filter designs were tested to determine their efficacy and efficiency at removing corbicular pollen loads from returning bumble bee workers. All designs were successful at removing at least of one corbicular pollen load from a returning forager. However, some were found to slow workers from leaving or entering the hive or failed to remove pollen loads (Table 1). Pollen traps with various filters were tested sequentially on 4 laboratory reared colonies of B. huntii Greene f...

Watch this Scientific Journal Video about A 3D Printed Pollen Trap for Bumble Bee Bombus Hive Entrances at JoVE.com

A 3D Printed Pollen Trap for Bumble Bee Bombus Hive Entrances

We present a non-lethal and automated mechanism to collect pollen from bumble bee (Bombus) workers returning to a hive. Instructions for producing, preparing, installing and using the devices are included. By using 3D-printed objects, modification to the design was timely, efficient and allowed for quick turnaround for testing.

A 3D Printed Pollen Trap for Bumble Bee (Bombus) Hive Entrances

To verify the plant sources from which bumble bees forage for pollen, individuals must be collected to remove their corbicular pollen loads for analysis.  ...

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Research

JoVE Journal

Biology

5.0K Views.  Utah State University. We present a non-lethal and automated mechanism to collect pollen from bumble bee (Bombus) workers returning to a hive. Instructions for producing, preparing, installing and using the devices are included. By using 3D-printed objects, modification to the design was timely, efficient and allowed for quick turnaround for testing.

Video: A 3D Printed Pollen Trap for Bumble Bee Bombus Hive Entrances

Measuring the Bending Stiffness of Bacterial Cells Using an Optical Trap

We developed a protocol to measure the bending rigidity of filamentous rod-shaped bacteria. Forces are applied with an optical trap, a microscopic three-dimensional spring made of light that is formed when a high-intensity laser beam is focused to a very small spot by a microscope's objective lens. To bend a cell, we first bind live bacteria to a chemically-treated coverslip. As these cells grow, the middle of the cells remains bound to the coverslip but the growing ends are free of this restraint. By inducing filamentous growth with the drug cephalexin, we are able to identify cells in which one end of the cell was stuck to the surface while the other end remained unattached and susceptible to bending forces. A bending force is then applied with an optical trap by binding a polylysine-coated bead to the tip of a growing cell. Both the force and the displacement of the bead are recorded and the bending stiffness of the cell is the slope of this relationship.

We present a protocol for bending filamentous bacterial cells attached to a cover-slip surface with an optical trap to measure the cellular bending stiffness.

We developed a protocol to measure the bending rigidity of filamentous rod-shaped bacteria. Forces are applied with an optical trap, a microscopic ...

Associated Chromosome Trap for Identifying Long-range DNA Interactions

Genetic information encoded by DNA is organized in a complex and highly regulated chromatin structure. Each chromosome occupies a specific territory, that may change according to stage of development or cell cycle. Gene expression can occur in specialized transcriptional factories where chromatin segments may loop out from various chromosome territories, leading to co-localization of DNA segments which may exist on different chromosomes or far apart on the same chromosome. The Associated Chromosome Trap (ACT) assay provides an effective methodology to identify these long-range DNA associations in an unbiased fashion by extending and modifying the chromosome conformation capture technique. The ACT assay makes it possible for us to investigate mechanisms of transcriptional regulation in trans, and can help explain the relationship of nuclear architecture to gene expression in normal physiology and during disease states.

The associated chromosome trap (ACT) assay is a novel unbiased method for identifying long-range DNA interactions. The characterization of long range DNA interactions will allow us to determine the relationship of nuclear architecture to gene expression in both normal physiology and in diseased states.

Genetic information encoded by DNA is organized in a complex and highly regulated chromatin structure.  Each chromosome occupies a specific territory, ...

Methods for Comparing Nutrients in Beebread Made by Africanized and European Honey Bees and the Effects on Hemolymph Protein Titers

Honey bees obtain nutrients from pollen they collect and store in the hive as beebread. We developed methods to control the pollen source that bees collect and convert to beebread by placing colonies in a specially constructed enclosed flight area. Methods were developed to analyze the protein and amino acid composition of the pollen and beebread. We also describe how consumption of the beebread was measured and methods used to determine adult worker bee hemolymph protein titers after feeding on beebread for 4, 7 and 11 days after emergence. Methods were applied to determine if genotype affects the conversion of pollen to beebread and the rate that bees consume and acquire protein from it. Two subspecies (European and Africanized honey bees; EHB and AHB respectively) were provided with the same pollen source. Based on the developed methods, beebread made by both subspecies had lower protein concentrations and pH values than the pollen. In general, amino acid concentrations in beebread made by either EHB or AHB were similar and occurred at higher levels in beebread than in pollen. Both AHB and EHB consumed significantly more of the beebread made by AHB than by EHB. Though EHB and AHB consumed similar amounts of each type of beebread, hemolymph protein concentrations in AHB were higher than in EHB. Differences in protein acquisition between AHB and EHB might reflect environmental adaptations related to the geographic region where each subspecies evolved. These differences could contribute to the successful establishment of AHB populations in the New World because of the effects on brood rearing and colony growth.

Here are methods to quantify nutrients in pollen before and after its conversion to beebread by two subspecies of honeybees. We describe techniques to measure beebread consumption and resulting protein titers in both subspecies.

Honey bees obtain nutrients from pollen they collect and store in the hive as beebread. We developed methods to control the pollen source that bees ...

Monitoring Colony-level Effects of Sublethal Pesticide Exposure on Honey Bees

The effects of sublethal pesticide exposure to honey bee colonies may be significant but difficult to detect in the field using standard visual assessment methods. Here we describe methods to measure the quantities of adult bees, brood, and food resources by weighing hives and hive parts, by photographing frames, and by installing hives on scales and with internal sensors. Data from these periodic evaluations are then combined with running average and daily detrended data on hive weight and internal hive temperature. The resulting datasets have been used to detect colony-level effects of imidacloprid applied in a sugar syrup as low as 5 parts per billion. The methods are objective, require little training, and provide permanent records in the form of sensor output and photographs.

Sublethal doses of pesticides may affect colonies in ways that are difficult to detect using only periodic or visual methods. Methods to measure total adult bee mass, brood, and food resources by weighing hives and hive parts, photographing frames, and installing sensors, are provided. Data analysis is also addressed.

The effects of sublethal pesticide exposure to honey bee colonies may be significant but difficult to detect in the field using standard visual assessment ...

Collection and Identification of Pollen from Honey Bee Colonies

Researchers often collect and analyze corbicular pollen from honey bees to identify the plant sources on which they forage for pollen or to estimate pesticide exposure of bees via pollen. Described herein is an effective pollen-trapping method for collecting corbicular pollen from honey bees returning to their hives. This collection method results in large quantities of corbicular pollen that can be used for research purposes. Honey bees collect pollen from many plant species, but typically visit one species during each collection trip. Therefore, each corbicular pollen pellet predominantly represents one plant species, and each pollen pellet can be described by color. This allows the sorting of samples of corbicular pollen by color to segregate plant sources. Researchers can further classify corbicular pollen by analyzing the morphology of acetolyzed pollen grains for taxonomic identification. These methods are commonly used in studies related to pollinators such as pollination efficiency, pollinator foraging dynamics, diet quality, and diversity. Detailed methodologies are presented for collecting corbicular pollen using pollen traps, sorting pollen by color, and acetolyzing pollen grains. Also presented are results pertaining to the frequency of pellet colors and taxa of corbicular pollen collected from honey bees in five different cropping systems.

We describe methods for collecting corbicular pollen from honey bees as well as protocols for color sorting, acetolysis, and microscope slide preparation of pollen for taxonomic identification. In addition, we present pellet color and taxonomic diversity of corbicular pollen collected from five cropping systems using pollen traps.

Researchers often collect and analyze corbicular pollen from honey bees to identify the plant sources on which they forage for pollen or to estimate ...

Assessing Agrochemical Risk to Mated Honey Bee Queens

Current risk assessment strategies for honey bees rely heavily upon laboratory tests performed on adult or immature worker bees, but these methods may not accurately capture the effects of agrochemical exposure on honey bee queens. As the sole producer of fertilized eggs inside a honeybee colony, the queen is arguably the most important single member of a functioning colony unit. Therefore, understanding how agrochemicals affect queen health and productivity should be considered a critical aspect of pesticide risk assessment. Here, an adapted method is presented to expose honey bee queens and worker queen attendants to agrochemical stressors administered through a worker diet, followed by tracking egg production in the laboratory and assessing first instar eclosion using a specialized cage, referred to as a Queen Monitoring Cage. To illustrate the method&#39;s intended use, results of an experiment in which worker queen attendants were fed diet containing sublethal doses of imidacloprid and effects on queens were monitored are described.

This protocol was developed to enhance the understanding of how agrochemicals affect honey bee (Apis mellifera) reproduction by establishing methods to expose honey bee queens and their worker caretakers to agrochemicals in a controlled, laboratory setting and carefully monitoring their relevant responses.

Current risk assessment strategies for honey bees rely heavily upon laboratory tests performed on adult or immature worker bees, but these methods may not ...

An Innovative 3D-Printed Insert Designed to Enable Straightforward 2D and 3D Cell Cultures

The classical analyses of indirect communication between different cell types necessitate the use of conditioned media. Moreover, the production of conditioned media remains time-consuming and far from physiological and pathological conditions. Although a few models of co-culture are commercially available, they remain restricted to specific assays and are mostly for two types of cells. 
Here, 3D-printed inserts are used that are compatible with numerous functional assays. The insert allows the separation of one well of a 6-well plate into four compartments. A wide range of combinations can be set. Moreover, windows are designed in each wall of the compartments so that potential intercellular communication between every compartment is possible in the culture medium in a volume-dependent manner. For example, paracrine intercellular communication can be studied between four cell types in monolayer, in 3D (spheroids), or by combining both. In addition, a mix of different cell types can be seeded in the same compartment in 2D or 3D (organoids) format. The absence of a bottom in the 3D-printed inserts allows the usual culture conditions on the plate, possible coating on the plate containing the insert, and direct visualization by optical microscopy. The multiple compartments provide the possibility to collect different cell types independently or to use, in each compartment, different reagents for RNA or protein extraction. In this study, a detailed methodology is provided to use the new 3D-printed insert as a co-culture system. To demonstrate several capacities of this flexible and simple model, previously published functional assays of cell communication were performed in the new 3D-printed inserts and were demonstrated to be reproducible. The 3D-printed inserts and the conventional cell culture using conditioned media led to similar results. In conclusion, the 3D-printed insert is a simple device that can be adapted to numerous models of co-cultures with adherent cell types.

In this paper, a newly designed 3D-printed insert is presented as a model of co-culture and validated through the study of the paracrine intercellular communication between endothelial cells and keratinocytes.

The classical analyses of indirect communication between different cell types necessitate the use of conditioned media. Moreover, the production of ...

An Improved Method to Measure Pollen Hydration Profiles in Arabidopsis thaliana

A High-Resolution, Single-Grain, In Vivo Pollen Hydration Bioassay for Arabidopsis thaliana

Sexual reproduction in flowering plants requires initial interaction between the pollen grain and the stigmatic surface, where a molecular dialog is established between the interacting partners. Studies across a range of species have revealed that a series of molecular checkpoints regulate the pollen-stigma interaction to ensure that only compatible, generally intraspecific pollen is successful in effecting fertilization. In species that possess a 'dry stigma', such as the model plant Arabidopsis thaliana, the first post-pollination, prezygotic compatibility checkpoint is the establishment of pollen hydration. 
This phase of pollination is tightly regulated, whereby signals from the pollen grain elicit the release of water from the stigma, thus permitting pollen hydration. The ability to accurately measure and track pollen hydration over time is key to the design of experiments directed at understanding the regulation of this critical step in reproduction. Published protocols frequently utilize flowers that have been excised from the parent plant, maintained on liquid or solid media, and bulk pollinated. 
This paper describes a noninvasive, in vivo pollination bioassay that permits minute-by-minute hydration tracking of individual A. thaliana pollen grains at high resolution. The assay is highly reproducible, able to detect very subtle variations of pollen hydration profiles, and thus is suitable for the analysis of mutants that affect pathways regulating pollination. Although the protocol is lengthier than those described for bulk pollinations, the precision and reproducibility it provides, along with its in vivo nature, make it ideal for the detailed dissection of pollination phenotypes.

Author Spotlight: A High-Resolution, Single-Grain, In Vivo Pollen Hydration Bioassay for Arabidopsis thaliana  

An improved method to measure pollen hydration profiles in Arabidopsis thaliana is described here. The new method offers higher resolution, is noninvasive, and is highly reproducible. The protocol represents a new tool for a finer dissection of the processes that regulate the early stages of pollination.

Sexual reproduction in flowering plants requires initial interaction between the pollen grain and the stigmatic surface, where a molecular dialog is ...

Applying Compounds to Citrus Plants Using a Novel 3D-Printed Direct Plant Infusion (DPI) Device

Using a 3D Printed Holder for Bulk Electroacupuncture in Small Animals

Bulk Electroacupuncture Operation for Mice or Young Rats

Electroacupuncture (EA) is widely used to treat various health conditions. However, the underlying mechanism of EA treatment remains unclear, hindering its promotion. The mechanistic study requires mouse or rat models to address this issue. However, these animals are not obedient to the experimental process, which is time-consuming. To solve these problems, we designed a 3D-printed small animal body bulk fixator to improve the efficiency of EA's animal experiments. This video shows in detail how to use the fixator to perform bulk EA on mice or young rats. For the selection of acupoints, the anterior oblique line of vertex temporal (MS6 head) and Tianshu point (ST25 belly) were chosen to verify the effect of the fixation device with prone positioning and supine positioning. Using the 3D-printed small animal holder allows multiple rodents to be immobilized and treated simultaneously, reducing the time and resources required for the experiment. This technique could be applied to other animal models by 3D printing different sizes and could potentially be used for various fixing conditions. The device is beneficial for the promotion of experimental scientific research in EA.

Author Spotlight: Using a 3D-Printed Holder for Simultaneous Multiple Electroacupuncture Operation

Here, we present a protocol for efficient and rapid electroacupuncture (EA) in mice or young rats using a three-dimensional (3D) printed holder. This technique allows simultaneous operation on multiple animals, saving time and increasing experimental efficiency.

Electroacupuncture (EA) is widely used to treat various health conditions. However, the underlying mechanism of EA treatment remains unclear, hindering ...