Nov 27, 2024
Intraarticular injection of fast blue
- 1King's College London
Protocol Citation: George Goodwin, Franziska Denk 2024. Intraarticular injection of fast blue. protocols.io https://dx.doi.org/10.17504/protocols.io.kxygxwexzv8j/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: November 15, 2024
Last Modified: November 27, 2024
Protocol Integer ID: 112180
Keywords: intraarticular injection, fast blue, retrograde labelling, joint afferents
Funders Acknowledgements:
UKRI
Grant ID: MR/W027518/1
Abstract
Protocol for injection of fast blue into the knee joint cavity for retrograde labelling of intraarticular afferents.
Materials
Equipment/drugs:
Heating blanket
Ketamine (20mg/ml)
Medetomidine (0.2mg/ml)
Atipamezole (0.5mg/ml)
Fast blue (2% in sterile water; Polysciences, cat.#: 105190-94-1)
Electric shaver
70% Ethanol swab
10ul Hamilton syringe
30g Hamilton syringe removable needle
1ml Syringes
26 G needle
Warm recovery chamber
0.9% saline
Preparing animal and induction of anaesthesia
Preparing animal and induction of anaesthesia
Weigh mouse.
Prepare heat mat to keep animal warm during anaesthesia.
Using 1ml syringe with 26g needle attached, perform intraperitoneal (i.p.) injection of ketamine (75mg/kg) and medetomidine (1mg/kg).
Once mouse has lost consciousness, use a shaver to remove the fur from around the knee joint you intend to inject.
Wipe skin around knee joint with 70% ethanol swab.
Intraarticular injection of Fast blue
Intraarticular injection of Fast blue
Place the mouse belly up and hold the leg at the ankle with the knee in a slightly bent position.
Run the side of the needle up and down the front of the leg until it rests in the ‘groove’, running from the lateral to medial meniscus. Press with the side of the needle to indent the skin slightly, so that you can visualise the location of the ridge.
Using a 30g removable needle attached to a 10ul Hamilton syringe, insert the needle from the front (with the needle perpendicular to the front of the knee) about 2-2.5mm depth. You will know that the needle has entered the articular space as you will feel a slight ‘give’ as the needle passes into the space rather than hitting bone.
Inject 2l of Fast blue into the knee joint. Note that this is reported to be the maximum volume that can be injected without it leaking out of the knee joint1.
Take care not to move the mouse around too much to ensure that the knee is not flexed/extended. Leave mouse on heat pad for 1-1.5 hours - the idea behind this waiting period is that it gives the Fast blue more time to be absorbed, and there is reduced chance that it leaks out. However, we have never directly tested whether shorter periods would lead to more leakage.
Animal recovery
Animal recovery
Ensure mouse is hydrated during recovery by administering 0.2ml 0.9% sterile saline subcutaneously.
Using a 1ml syringe with 26g needle attached, reverse medetomidine anaesthesia by giving an i.p. injection of atipamezole (1mg/kg).
Place mouse in warm heated chamber to recover for ~1.5-2 hours.
Once mice have fully recovered from the ketamine anaesthesia, place back into home cage.
Protocol references
1 Xu Yang, V. C., Alberto Carli, Matthew B. Shirley, & Olufunmilayo Adebayo, M. C. H. v. d. M., F. Patrick Ross, Mathias P.G. Bostrom. 2016 Annual Meeting of the Orthopaedic Research
Society Orlando, FL March 5-8, 2016 -
Refining Intra-articular and Intramedullary Injection Techniques to Establish
Joint, Bone and Periprosthetic Joint Infection in Mice. J Orthop Res 34 Suppl 1,
S1, doi:10.1002/jor.23247 (2016).