May 25, 2026
Vibration Latency at the Knee in Wistar Rodents
- Ashleigh Abbott1,
- Robert Dalton1,
- Kaitlin Southern1,
- Folly Patterson1,
- Kyle D. Allen1
- 1University of Florida
Protocol Citation: Ashleigh Abbott, Robert Dalton, Kaitlin Southern, Folly Patterson, Kyle D. Allen 2026. Vibration Latency at the Knee in Wistar Rodents. protocols.io https://dx.doi.org/10.17504/protocols.io.ewov1r6zolr2/v1
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it's working
Created: April 28, 2026
Last Modified: May 25, 2026
Protocol Integer ID: 315893
Keywords: Vibration, Nociception, Pain-like behavior, Rodent, Mechanical Sensitivity, Knee, wistar rodents vibration nociception threshold in rodent, wistar rodents vibration nociception threshold, vibration nociception threshold, variability in vibration nociception, vibration latency at the knee, vibration nociception, nociception at the knee, withdrawal threshold of vibration, nociception, vibration, different pain model, vibration latency, unilateral models such as medial collateral ligament transection, knee, medial collateral ligament transection, medial meniscus transection, rodent, withdrawal threshold
Funders Acknowledgements:
National Institute of Arthritis and Musculoskeletal and Skin Diseases
Grant ID: UC2AR082196
Abstract
Vibration nociception threshold in rodents is assessing using a 128 hz tuning fork to stimulate nociception at the knee in rodents. Withdrawal threshold of vibration is determined when the rodent either physically withdrawals their leg and if they vocalize. Vibration nociception threshold is beneficial to measure the variability in vibration nociception across rodents as well as across different pain models or unilateral models such as medial collateral ligament transection and medial meniscus transection (MCLT+MMT).
Materials
- Tuning fork (128 hz)
- Timer (phone timer works)
Personal Protective Equipment
- Disposable gloves
- Face mask (optional)
- Disposable gown
Safety warnings
Protocols and behavioral collection requires prior approval by the users' Institutional Animal Care and Use Committee (IACUC) or equivalent ethics committee.
Ethics statement
Protocols and behavioral collection described here were approved by the local Institutional Animal Care and Use Committee (IACUC) and university ethics committee.
Before start
Acclimation (1-2 weeks)
Before data is collected, animals must be acclimated to experimenter handling and the tuning forks. The acclimation period occurs before baseline data is collected. There is no data collection during acclimation. On the first week, each rat should undergo acclimation to handling for periods of 5-10 minutes (routine handling acclimation). The second week of acclimation will consist of handling in the c-hold and acclimation to palpation of the knee. It is important the animals are comfortable through testing to not skew results, so increase acclimation time as needed. During acclimation, the conditions in the behavior room should mimic those present during the testing period.
Materials and Personal Protective Equipment
Materials:
Tuning fork (128 hz)
Timer (phone timer can be used)
Figure 1: Tuning fork at 128 hz used in testing. When holding the tuning fork for testing, don't touch the prongs to prevent vibration damping. To activate vibration hit the top of the prongs near the weights on the palm of your hand. Use roughly the same force and distance of hitting it on your palm to keep consistent throughout the experiment.
Personal Protective Equipment
Disposable gloves
Face mask (optional)
Disposable gown
Acclimation (~2 Weeks)
Before data is collected, animals must be acclimated to experimenter handling. The acclimation period occurs before baseline data is collected. There is no data collection during acclimation. The animals are acclimated to the room for 30 minutes prior to starting handling. On the first week, each rat should undergo acclimation to handling for periods of 5-10 minutes (routine handling acclimation). Routine handling acclimation includes being touched, picked up, and general handling. The second week of acclimation will consist of handling in the c-hold and acclimation to palpation of the knee. It is important the animals are comfortable through testing to not skew results, so increase acclimation time as needed. During acclimation, the conditions in the behavior room should mimic those present during the testing period.
Note: the acclimation experience and time can depend on the age and sex of the rats and handlers. Younger rats (3-12 months of age) were more active and did not remain still. However, the older rats (15+ months of age) needed longer acclimation times. The older rats were less active, but were more reactive. Also, females were generally more reactive to handlers and male handlers over females.
Data Collection
Frequency: The anterior aspect on the knee will be tested three times, resulting in 3 trials collected/rodent. Only one aspect of the knee is tested because the vibrational signal is conducted through bone.
When ready to test, take a few rodents into the room at a time to reduce the effect of the light cycle (if on reverse light cycle).
Wait 30 minutes for animals to acclimate to the testing space (in their cage) before beginning testing. Then when ready remove animals from their cage one rat at a time for testing.
When testing, the handler will be in a seated position holding the rodent against their chest in a C-hold. A second experimenter will be necessary to start the timer and apply the tuning fork. A third person to do the timer is ideal when possible.
The handler will hold the rodent in the C-hold against their chest with their dorsal side towards the handlers chest and the knee facing out.
Once the animal is settled and not squirming, the second experimenter will palpate the knee lightly to locate the knee joint space.
Note: to locate the knee joint space, feel the knee to find where there is a space or where you can feel the BB like structure on the lateral side of the knee (See Figure 2A). Then, apply the tuning fork near the anterior aspect of the knee inferior to patella (See Figure 2B). [Figure created using biorender.com]
Figure 2: A) Apply the tuning fork inferiro to the patella in the medial aspect of the knee joints. To located the joint space, palpate the knee gently to locate the "BB like structure" (fibular head). B) Apply the tuning fork to the medial aspect of the knee. Note: As the rat can vary in size and anatomy, check for the joint space as it may be more cranial than expedcted.
Hit the tuning fork to activate it and place the bottom of the tuning fork on the anterior aspect of the joint space.
Note: Hit the tuning fork approximately 5-7 inches away from the palm of your hand and use approx. the same force every time to normalize across trials. Try to avoid hitting the tuning fork near the rodent before testing to prevent fear response.
Without touching any portion of the tuning fork except the base, place the bottom surface of the tuning fork on the knee near the patella.
Hold the tuning fork on this aspect of the knee for about 10 seconds or until a response is recorded.
Record the type of response and time to response(s):
A positive response is an audible vocalization to the ear, a visual withdrawal from the stimulus, and/or a reflexive withdrawal felt by the animal handler. Audible response is only considered a positive response if accompanied by a visual or reflexive withdrawal.
An ambiguous response to the stimulus is if during or after the ten seconds count the rat moves/slightly raises or shakes its leg from the tuning fork without vocalizing.
If an animal has an ambiguous response repeat the measurement or application of the tuning fork until you get a positive (X) or a negative (O) response.
A negative or non-response to the stimulus, is if after the ten seconds count rat neither moves/takes away their leg (an ambiguous response) or physically withdraws their leg. (a positive response).
Note: Record the type of withdrawal (vocalization, visual withdrawal, reflexive withdrawal)
Proceed to testing once on each animal for one trial.
Once all animals have been tested once, repeat steps 7-14 two times resulting in 3 trials for each rodent.
Note: make sure there is a 30-minute separation between trials on the same rodent.