Until now, there has been no easy method available to allow the investigator and Quality Assurance Manager to check the validity of their mechanical threshold measurements on mice or to compare their overall measurement technique with other institutions. The buckling force of individual filaments may be checked against laboratory scales but this takes no account of poor application technique or errors in the use, after testing, of the up-down or % response algorithms. The same is true if an Electronic Von Frey (EVF) system is used.
MouseCal is a novel validation tool which allows organisations to check the accuracy of the tester’s technique when measuring mechanical thresholds. It is also a valuable training aid for new operators in the use of filaments (or EVF) before they start work with animals. The tool may be used with all traditional von Frey filament sets and with EVF systems (such as MouseMet).
For filaments, the possible sources of error include:
- Bent or damaged filaments that buckle at the wrong force
- Poor operator technique when using the filaments
- Incorrect use of the up-down or % response algorithms, resulting in the wrong threshold value from the data.
For EVF systems, the possible sources of error include:
- Incorrect force calibration of the system
- Poor operator technique
- Use of an inappropriate system for the measurement (usually with too large a force range for mice).
Please watch the video and then read on for the principle of operation and an example of how it may be used to validate your mechanical threshold measurements.
MouseCal is the latest product from Topcat Metrology. As can be seen in the video, the see-saw arm has an over-centre action which mimics a mouse’s foot withdrawal. Practise is therefore required (as with an animal) to ensure that the filament (or EVF) is applied correctly. The force required to tip the arm also has a random component, to simulate the biological variation that always occurs with live subjects (a variation that is in fact necessary for up-down filament algorithms to work). The random variation in MouseCal is generated in software, applied electronically and has been carefully chosen to give a standard deviation similar to that from data on real mice. The tool therefore provides two validity, or quality, measures from each data point.
MouseCal will be available worldwide, initially for short term rent, with the aim of building up a database of information about the accuracy of mechanical threshold tests on mice. This information, showing the spread of measurements around each data point, will be available to all contributors, together with (in confidence) an indication of where the measurement made by each organisation sits within the spread. Four data points will initially be available, spanning the range of 1-4 grams force.
Participation will involve the following steps:
- PI (or QA manager) receives MouseCal with a set of test weights.
- The PI gives the equipment to the operator who, following the instructions given, measures the threshold force required to tip the arm for each of the masses in turn, and for the bare arm without any weight, using whatever method the organisation normally uses for mice. If using filaments and the up-down method then the algorithm (and method of applying it) for deriving the threshold force from the up-down sequence is also checked. If using an EVF system, then the usual number of readings at each time point are taken and averaged.
- The operator gives the PI their estimate of the mean and standard deviation for the threshold force of each weight.
- The PI returns MouseCal to Topcat. The four mean forces are entered into Topcat’s evolving database and compared with the values from the other contributors. For each force, the variation from the database mean is then reported to the PI.