EthoVision XT - The Sociability Cage Test
Last updated: Jul 25, 2026
The Sociability Cage Test
Introduction
the sociability test (three-chamber paradigm)
The three-chamber paradigm test known as Crawley's sociability and preference for social novelty protocol has been successfully employed to study social affiliation and social memory in several inbred and mutant mouse lines. Sociability testing is a more specific study of social behavior and only focuses on the behavior of one animal towards others. There are several variances to this test, all conducted in a three- chambered test apparatus. In the first variant, the animal's interest in a social stimulus (a conspecific in a wired cage placed in one of the outer chambers) versus a neutral stimulus (an empty wired cage placed in the other outer chamber) is studied. In a preference test for social novelty, the two wired cages contain a familiar and an unfamiliar conspecific. A third variation involves testing the social preference for two different, but both unfamiliar conspecifics.
Figure 6.1 The Noldus Sociability test cage with three chambers and two wired
cages for mice. In most cases the interest is measured by assessing the time spent in the same chamber or in close proximity to the familiar or unfamiliar other mouse.
Representative results
Typically, a wild type animal will spend significantly more time in the compartment with a strange mouse compared to the compartment with empty cage, indicating normal sociability, social motivation and affiliation. In the social novelty test, the subject has a free choice between the first, already-investigated, now-familiar mouse, and a novel unfamiliar mouse. Usually, a wild-type animal recalls its previous contact with the familiar mouse, and, in this session, tends to spend more time with the newly encountered mouse, indicating intact social memory and predilection for novel experiences.
references
Nadler, J.J., Moy, S.S., Dold, G., Perez, A., Young, N.B., Barbaro, R.P., Piven, J., Magnuson, T.R., Crawley, J.N. (2004). Automated apparatus for rapid quantification of social approach behaviors in mice. Genes Brain Behav., 3:303 - 314. Moy, S,S, Nadler, J.J., Perez, A., Barbaro, R.P., Johns, J.J., Magnuson, T.R., Piven, J., Crawley, J.N. (2004) Sociability and preference for social novelty in five inbred strains: an approach to assess autistic-like behaviors in mice. Genes Brain Behav 3, 287 - 302.
The Sociability test and EthoVision XT
Bechtstein, P., Rehbach, N.-J., Yuhasingham, G., Schürmann, C., Göpfert, M., Kössl, M., Maronde, E. (2014). The clock gene Period1 regulates
innate routine behavior in mice. Proceedings of the Royal B Society, 281,
- DOI 10.1098/rspb.2014.0034. Desbonnet, L., O'Tuathaigh, C., Clarke, G., O'Leary, C., Petit E., Clarke N., Tighe, O., Lai, D., Harvey, R., Cryan, J.F. Dinan, T.G., Waddington, J.L. (2012). Phenotypic effects of repeated psychosocial stress during adolescence in mice mutant for the schizophrenia risk gene neuregulin- 1: A putative model of gene × environment interaction. Brain, Behavior, and Immunity 26(4), 660 - 671. Glass, M.J., Robinson, D.C., Waters, E., Pickel, V.M. (2013). Deletion of the NMDA-NR1 receptor subunit gene in the mouse nucleus accumbens attenuates apomorphine-induced dopamine D1 receptor trafficking and acoustic startle behavior. Synapse, 67(6), 265-279. Kim J-W, Seung H, Kwon KJ, Ko MJ, Lee EJ, et al. (2014) Subchronic Treatment of Donepezil Rescues Impaired Social, Hyperactive, and Stereotypic Behavior in Valproic Acid-Induced Animal Model of Autism.
PLoS ONE 9(8): e104927. DOI: 10.1371/journal.pone.0104927
Ter Horst, J. P., van der Mark, M., Kentrop, J., Arp, M., van der Veen, R., de Kloet, E. R., & Oitzl, M. S. (2014). Deletion of the forebrain mineralocorticoid receptor impairs social discrimination and decision- making in male, but not in female mice. Frontiers in Behavioral
Neuroscience, 8, 26. DOI: 10.3389/fnbeh.2014.00026
The Sociability test in EthoVision XT
experiment settings
Choose Setup > Experiment Settings.
Under Video Source, and Tracked Features, make sure that the options
selected corresponds to your needs. If you want to analyze exploration in detail, choose Center-point, Nose-point and Tail-base detection. If you want to use Deep learning technique to track the subject's nose,
under Body point detection technique choose Deep learning. Note that
in order to use this technique there are additional requirements.
Among other things, you need a powerful graphics card. See Deep
learning: Requirements and Limitations in the EthoVision XT Help.
manual scoring settings
If your EthoVision XT license does not include the Behavior Recognition module, EthoVision XT cannot detect sniffing or rearing automatically. However, you can still score those and other behaviors with keystrokes. Choose Setup > Manual Scoring Settings. If you used the template for an open field experiment, four behaviors have been defined: Sniffing R and Sniffing O, to record manually sniffing at the receiver (unfamiliar mouse) and object (either familiar mouse or empty cage, depending on the test), respectively. Rearing R and Rearing O, to record manually sniffing at the receiver (unfamiliar mouse) the object cage (either familiar mouse or empty cage, depending on the test), respectively. You can also rename the behaviors if you like.
arena settings
Choose Setup > Arena Settings. If you work with a Sociability test cage with backlight unit, you can reduce the intensity of the backlight before adjusting the arena settings. This makes the edges of the floor plate and walls more visible. If the separation walls show reflections of the animal, this may hinder tracking. In the Arena Settings, click the Arena layer, and draw rectangles around the separation walls, like in the example below.
detection settings
Choose Setup > Detection Settings.
We assume that you followed the procedure in detection settings on
page 17.
Check in the Video Section that the sample rate is set to:
For center point tracking: 5-8 samples/second (for rats) and 12.5-15 samples/second (for mice). For Nose-tail tracking (rats and mice). 25-30 samples/second.
Notes
If detection of the subject is not optimal after using the Automated
setup function, see Configure Detection Settings in the EthoVision XT
Help for details on the advanced detection settings. If you use hooded animals, use the detection method Differencing. If the animal moves from one compartment to the other, it may not be detected well, like in this example.
To improve detection, open the Advanced section In the Detection
Settings pane. Under Subject Contour, select one or more pixels for the
Dilation filter and the second Erosion filter. Leave the first Erosion filter
to zero. Select more pixels for Dilation than for Erosion, for example 3
vs. 1, until the entire animal's contour is detected.
trial list
Choose Setup > Trial List. Click Add Variables and enter your
independent variables such as the experimental phases and the ID code of your test subject.
Acquiring data
protocol
Sociability test
For each test animal, three trials are carried out: 1. The first trial (for example 5 min) aims at habituating the test individual to the chamber. Tracking is required when one wishes to know whether the animal explores one chamber more than the other. 2. A second trial (5 min) is carried out with the empty tubes in the test chambers. The aim of this trial is to quantify the time spent in the two side chambers and near the tubes. 3. In the third trial (next 5 min), an empty tube is removed and replaced by a female animal restrained in an identical tube. This session is designed to observe differences between the time spent sniffing the social versus non-social object.
For each test animal, three trials are carried out: 1. The first trial (10 min) is aimed at habituating the test individual to the chamber and empty tubes. 2. In the sociability test (next 10 min), an empty tube is removed and replaced by a female animal restrained in an identical tube. This session is designed to observe differences between the time spent sniffing the social versus non-social object. 3. In the social discrimination test (10 min), the other empty tube is removed and replaced by a novel female. This session is designed to test the ability of the test animal to distinguish between the two females. See also acquiring tracks on page 21.
score behaviors manually
To score sniffing and other behaviors defined under Manual Scoring Settings, open the Manual Scoring tab on the Acquisition screen. There you find the key codes for the behaviors. To score an instance of the behavior or its end, either press the key or click the corresponding button on the screen.
Notes
important If you do tracking from video, make sure you de-select
DDS in the Playback Control window.
You can also score behaviors after tracking. See Acquire Data >
Score behaviors manually in the EthoVision XT Help.
Data analysis
data preparation
Data editing
Choose Acquisition > Edit Tracks. You can fix tracking errors and swap
back nose-points and tail-points that have been swapped in tracking. Normally you will not need to edit your data.
Smoothing data
Choose Acquisition > Track Smoothing Profile and open MDM Filter 0.2
cm. In this profile, the Minimal Distance Moved filter is used with a
value of 2 mm and option Direct.
Review video and behaviors
If you have scored behaviors manually, you can review the video and if
necessary edit the data. Choose Acquisition > Score behaviors
manually. For details, see Acquire Data > Score behaviors manually in
the EthoVision XT Help.
analysis profiles
The template experiment contains four analysis profiles: Distance and velocity. For quantifying activity and exploration. Combine this with a Data profile that specifies Results per zone.
In chambers. To calculate the time spent in each of the three
chambers.
Behaviors. To analyze behaviors scored manually (rearing and
sniffing) or the time that the nose point was detected in the area around each cage (sniffing zone).
On cage. To calculate how often and how long the center point of
the subject was found in the zone "cage" (left and right). This correlates with the time the mouse spent on top of the cages. Add other analysis profiles according to your research questions. Click
Statistics tab to select which statistic to calculate.
Example
The researcher wants to know the time from the moment the animal is entirely in the right compartment to the first moment the animal's nose-point is close to the cage in the same compartment. To calculate this time, in the Analysis profile choose Free Interval. Select an interval that goes from the time when the Current value of In zone, for zone Cage right, and for all body points, is "true" (Start criterion), to when the Current value of In zone for the zone "Interest zone right", and for the nose point, is true (Stop criterion). In the Statistics tab, select Cumulative duration. For more information, see the EthoVision XT Help.
visualizing data
Choose Analysis > Results > ...
Plot tracks. You can view your tracks on a still image of the
background. Tracks can be shown in different colors according to the values of independent variables (for example, blue for animals in Phase 2 and red for animals in Phase 3). Sample points can be shown in different colors according to the values of dependent variables, for example red when the animal was moving fast.
Plot integrated data. You can look at a track with the video file in
the background. When you plot integrated data you can also view Time Event plots of your independent variables. Just like with plotting tracks, you can show tracks or sample points in different colors.
Plot Heatmaps. Make heatmaps of the location of your animals
during the different trials. In the example below: Left heatmap: Cages empty. Right: mouse restrained in the Left cage (right cage empty).
Source: EthoVision XT 17.5 Application Manual, The Sociability Cage Test