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The T-Maze is a behavioral assay of cognitive ability. It assesses reference memory, working memory, and spontaneous exploration. It also provides insight into the function of the hippocampus and septum in models of neurodegenerative diseases such as Alzheimer’s disease or other diseases that impact hippocampal function.

The apparatus has three arms of equal length at an angle of 90º from one another. There are guillotine doors and a small partition at the top of the maze separating the left and right “GOAL” arms.

Each arm has a small hole that can hold food within a well. The apparatus can have opaque or clear arms and walls for exploration with the following dimensions for each arm: 15in x 6in x 9in (40cm x 15cm x 23cm) (LxWxH).

Materials for T-Maze

  • Animals: Mice, rats
  • Sample Size:  n ≥ 12 animals
  • Lights: Lamps
  • Video: Video camera mounted on our portable camera stand with articulating arm.
  • Cleaning: SporKlenz, 10% Bleach, or 70% Ethanol
  • Optional: Dark box for the ends of the arms is sold separately as an add-on modification of the Y-maze.

Testing Conditions for T-Maze

  • Lights: Confirm the area is well-lit to ensure that alternation is not associated with preference for darker areas of the maze
  • Training: Discrete trial procedure; no fasting required to stimulate motivation to explore or food-seeking behaviors.
  • Privacy: The examiner at least (5 feet/1.5m) away from the testing apparatus.
  • Stimulus: Patterns or objects that the animal must respond to and engage in exploration
  • Analysis: Observation/review of footage; automated video tracking software for behavioral analysis
  • Pairing test with other assays: Two alternation trials per day can be run before the other scheduled tests
  • Automated T-Maze (Coming Soon): Automated mazes are equipped with photoelectric beams and cameras to record successful entry into an arm of the maze.

Software Analysis for T-Maze

  • Automated behavioral tracking software: Our list of recommended software for behavioral analysis.

T-Maze Protocol

As an example of the research applications for the T-maze, review this protocol used to assess spatial memory and cognitive function in 5XFAD mice, an established model for Alzheimer’s disease.

Step 1. Preparation:

  1. Acclimate mice to the housing facility for at least one week before the experiment.
  2. Maintain a controlled environment with a 12-hour light-dark cycle and ad libitum access to food and water.
  3. Ensure ethical compliance with institutional guidelines and regulatory requirements.

Step 2. Mouse Selection:

  1. Use age-matched 5XFAD transgenic mice and wild-type controls.
  2. Randomly assign mice to experimental and control groups.
  3. Record baseline characteristics such as age, weight, and sex.

Step 3. Habituation:

  1. Habituate mice to the testing room for at least 30 minutes before the experiment.
  2. Handle mice gently to minimize stress.
  3. Minimize extraneous odors and noises.

Step 4. T-Maze Setup:

  1. Place the T-maze in a quiet, well-lit room.
  2. Clean the maze thoroughly with a mild disinfectant before each trial.
  3. Ensure that the T-maze is symmetrical, and arms are identical in appearance.
  4. Record relevant maze dimensions and angles.

Step 5. Experimental Procedure:

  1. Perform a pre-training session to familiarize mice with the maze. Allow free exploration for 10 minutes.
  2. Conduct the actual trial: a. Start each trial by placing the mouse in the start arm. b. Allow the mouse to choose between left and right arms. c. Record the time taken to make a choice (latency) and the arm chosen. d. If the mouse chooses the correct arm (reward arm), provide a small food reward. e. If the mouse chooses the incorrect arm, gently guide it to the correct arm and allow exploration without a reward. f. Conduct a predetermined number of trials per session, with inter-trial intervals.

Step 6. Data Analysis:

  1. Perform statistical analysis using appropriate tests (e.g., t-test, ANOVA) to compare 5XFAD and control groups.
  2. Analyze latency to make a choice as a measure of spatial memory.
  3. Record the number of correct and incorrect choices as well as the following parameters for more thorough analysis:
    • Latency to Choice:The time taken by the mouse to make a choice between the arms, indicating spatial memory and decision-making speed. (Seconds)
    • Correct Choices: The number of times the mouse correctly chooses the arm associated with the reward, reflecting spatial learning. (counts)
    • Incorrect Choices: The number of times the mouse selects the arm without a reward, indicating errors in spatial memory or decision-making. (counts)
    • Total Arm Entries: The total number of entries into both arms, providing insights into general exploratory behavior. All four limbs must cross into the arm. (counts)
    • Choice Accuracy: The ratio of correct choices to the total number of choices, offering a comprehensive measure of spatial memory performance. (percentage)
    • Return Visits: The number of times a mouse re-enters the same arm during a trial, reflecting potential confusion or revisiting previous choices. (counts)
    • Inter-Trial Intervals: The time elapsed between the completion of one trial and the initiation of the next, providing insights into the mouse’s ability to retain spatial information across trials. (time; seconds or minutes)
    • Number (#) of alterations: Alternation is the motivation of the animal to explore an environment and to locate food, water, mates or shelter.
    • Side preference (%): calculate the number of entries within either the right or left side of the maze versus the total number of entries in each trial.
    • Percent (%) alteration (each day of testing): Normal animals have around 75% alternation. Anything ≤ 50% is considered a chance alternation or may imply hippocampal damage–depending on experimental groups.
    • Preference Index: A quantitative measure indicating the strength and direction of the mouse’s preference for one arm over the other, calculated as (Correct Choices – Incorrect Choices) / (Total Choices). [Index rage: -1 to 1]

These behavioral parameters collectively offer a detailed analysis of the spatial memory, learning, and decision-making abilities of 5XFAD mice in the T-maze assay. They contribute to a nuanced understanding of cognitive function and potential alterations associated with Alzheimer’s disease.

Alternation Result Interpretation

  • Latency to Choice: Longer latency may indicate impaired spatial memory or slower decision-making, potentially linked to hippocampal dysfunction.
  • Correct Choices: Increased correct choices suggest intact spatial learning and memory processes, while a decline may indicate cognitive deficits associated with Alzheimer’s disease pathology.
  • Incorrect Choices: Elevated incorrect choices may signify errors in spatial memory or decision-making, reflecting cognitive impairment and potential hippocampal dysfunction.
  • Total Arm Entries: Increased total arm entries may suggest heightened exploratory behavior, while a decline might indicate reduced overall activity, potentially associated with changes in motivation or cognitive function.
  • Choice Accuracy: Higher accuracy percentage reflects better spatial memory performance, while decreased accuracy may indicate cognitive deficits, particularly in the context of Alzheimer’s disease.
  • Return Visits: Increased return visits suggest potential confusion or difficulty in maintaining spatial information, providing insights into working memory and cognitive flexibility.
  • Inter-Trial Intervals: Shortened intervals may indicate rapid forgetting or impaired retention of spatial information, reflecting potential hippocampal dysfunction or memory consolidation issues.
  • Number of Alterations: Higher alteration rates suggest increased motivation for exploration and intact hippocampal function, while reduced alterations may indicate decreased motivation or hippocampal dysfunction.
  • Side Preference: Deviation from a balanced side preference may indicate altered spatial bias, potentially associated with changes in brain regions involved in spatial processing.
  • Percent Alteration (Each Day of Testing): A decline in percent alteration over consecutive testing days may suggest habituation or impaired learning, providing insights into cognitive adaptation or dysfunction.
  • Preference Index: Positive values indicate a preference for correct choices, while negative values suggest a bias towards incorrect choices. The index provides a nuanced measure of the strength and direction of cognitive biases in spatial decision-making.


  1. Deacon, R., Rawlins, J. T-maze alternation in the rodent. Nat Protoc 1, 7–12 (2006) doi:10.1038/nprot.2006.2
  2. Wu, C., Lerner, F. M., Couto E Silva, A., Possoit, H. E., Hsieh, T. H., Neumann, J. T., … Lee, R. (2018). Utilizing the Modified T-Maze to Assess Functional Memory Outcomes After Cardiac ArrestJournal of visualized experiments : JoVE, (131), 56694. doi:10.3791/56694.

Additional information

Weight 21.87 lbs
Dimensions 11.8 × 4 × 7.9 in



Honey bee, Mouse, Rat, Zebrafish

Maze Type

Elevated, Enclosed


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