Apr 24, 2026
Alzet Pump Placement and Stereotaxic Injections
Forked from Stereotaxic Injections
- Michael Henderson1,
- Lindsay Meyerdirk1,
- Naman Vatsa1
- 1Van Andel Institute
Protocol Citation: Michael Henderson, Lindsay Meyerdirk, Naman Vatsa 2026. Alzet Pump Placement and Stereotaxic Injections. protocols.io https://dx.doi.org/10.17504/protocols.io.kqdg3mod1l25/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 22, 2026
Last Modified: April 27, 2026
Protocol Integer ID: 315520
Keywords: ASAPCRN, stereotaxic injections this protocol detail, stereotaxic injections for any purpose, stereotaxic injection, injection, misfolded protein, protocol detail, protocol, purpose, protein, Alzet, alzet pump placement, alzet pump
Funders Acknowledgements:
Aligning Science Across Parkinson's
Grant ID: ASAP-020616
Abstract
This protocol details stereotaxic injections for any purpose, but in this case is primarily used for the injection of misfolded proteins, followed by the implantation of a cannula and Alzet pump.
Attachments
Guidelines
*All experiments involving animals must comply with federal and local law. All procedures should be reviewed and approved by the institutional animal care and use committee (IACUC).
Materials
Materials
1. Brilliant Blue (for practice injection)
2. Surgical Tools
- -Fine forceps
- -Needle Holders
- -Small Scissors
- -Curved/Flattened Forceps
- -Scalpel
3. Primed Alzet Pump
4. C&B Metabond Quick Adhesive Cement System, Parkell-(Cat #S380)
5. 10µL Hamilton Syringe (cat #7635-01)
6. 34 g needle custom ordered for: 1 in length, Tip 4, with 45 degree bevel (cat#207434)
7. Sterile Cotton Swabs
8. 6-0 Vicryl sutures (cat #J492G)
9. Syringes(0.5 mL and 1.0 mL)
10. Sterile Saline
11. Betadine Solution (7.5-10%)
12. 70% Ethanol
Anesthesia/Analgesia table:
| A | B | C | D | E | |
| Substance Name | Concentration | Dose | Route | Cat # | |
| Isoflurane | 2% | To effect | Intranasal | Henry Schein 1182097 | |
| Ketamine | 100 mg/mL | 100mg/kg | Intraperitoneal | Covetrus 071173 | |
| Xylazine | 100 mg/mL | 10mg/kg | Intraperitoneal | Covetrus 061035 | |
| Bupivacaine | 5 mg/mL | 4mg/kg | Subcutaneous | Covetrus 054893 | |
| Lidocaine | 2% | 4mg/kg | Subcutaneous | Covetrus 002468 | |
| Tylenol Water | 1 mg/mL | To effect | Oral | Sigma A7085-100g | |
| Buprenorphine | 0.3 mg/mL | 0.1mg/kg | Subcutaneous | Covetrus 059122 | |
| Ethiqa XR | 1.3 mg/mL | 3.25mg/kg | Subcutaneous | Covetrus 072117 |
Priming the Alzet Pump
Prepare sterile 0.9% saline. Add ~30 mL (enough to fully submerge the pump assembly) into a 50 mL conical tube (one tube per pump).
In a biosafety cabinet, remove the ALZET osmotic pump and brain infusion kit from their packaging.
Sterilize the outer surfaces of the pump and brain infusion kit with 70% ethanol before placing them inside the hood.
Weigh the empty pump and record the weight.
Attach the filling tube (provided) or a compatible small-volume syringe (for example, 0.1 mL) to a syringe containing the drug solution at room temperature. Ensure that the syringe and tubing are completely free of air bubbles. Use the smallest practical syringe to improve control.
Hold the pump upright with the outlet pointing vertically.
Insert the filling tube through the opening at the top of the pump until it reaches the bottom of the reservoir.
Slowly depress the syringe plunger to fill the pump. A small amount of backpressure is expected. Continue filling until a droplet appears at the outlet port, indicating the pump is full. Avoid rapid filling to prevent introduction of air bubbles.
Carefully remove the filling tube and wipe off any excess solution.
Weigh the filled pump. Calculate the volume loaded based on the difference between filled and empty weights.
If air bubbles are suspected inside the pump, empty the pump and refill carefully.
Assemble the catheter and cannula. Attach one end of the catheter to the cannula and the other end to the flow moderator.
Fill the catheter-cannula assembly with the drug or vehicle solution using the same method as above, ensuring no air bubbles are introduced.
Attach the filled pump to the catheter-cannula assembly, taking care to avoid trapping air at the connection point.
Place the complete pump-catheter-cannula assembly into the 50 mL conical tube containing sterile saline.
Incubate at 37 °C for 48 hours to allow proper priming of the pump.
After priming, maintain the assembly at 37 °C (for example, in a bead bath) until implantation in the surgery suite.
Head Alignment
With needle directly over Bregma, write down anterior-posterior (A-P), left-right (L-R), dorsal-ventral (D-V) position.
Go to Lambda. Check the height here and level with Bregma to within 0.1 mm tolerance by raising or lowering the ear bars slowly and evenly.
Go 2 mm to the right of Bregma and record the D-V measure. Go 2 mm to the left of Bregma and record the D-V measure. Level the skull within a tolerance of 0.1 mm.
Move to the injection site. Make a V-shaped mark at the injection site.
Injection
Move the needle far from the injection site.
Drill a hole with a 28 g burr (slightly bigger than the injection needle).
Move the injection needle back into place. Carefully move the needle to the top of the dura, without puncturing the tissue.
Calculate how far down to go.
Lower needle to the site of injection.
Inject material at 0.4 μL/min.
Wait 3 minutes after the injection has finished
Raise needle out of the brain.
Alzet Placement
Switch out the syringe holder for the cannula holder
Create a pocket in the skin along the side of the animal for the Alzet pump
Place the pump into the skin pocket and the canula into the cannula holder
Place the cannula holder at your bregma location
Move the cannula to the coordinates of your placement and mark your drilling place
Drill a hole using a 28g burr for the cannula
Using a burr holder and burr rough up the top of skull around the drilled hole
Clean off the area of dust/blood with 70% Ethanol and let it fully dry
Prepare Metabond cement Kit in cold (on ice) mixing block
. Place 4 drops of quick base and 1 drop of catalyst into 2 separate wells
. In the second well mix 1.5 scoops of the violet (clear) powder
Cover the surface you are going to cement with the base/catalyst mixture
Move the cannula down so it is at the skulls edge
Surround the base of the cannula top with the dental cement mixture
Slowly lower the cannula into position
Once in position use the dental cement mixture to cover the top of the cannula edge and skull area
Wait for the cement mixture to be totally dry before moving anything (at least 3 minutes).
Once dry, slowly release the cannula holder and raise it (make sure it is no cemented to the cannula)
Carefully cut off the cannula tab from the cannula using sharp scissors.
Clean the area and skin with sterile PBS to rehydrate the skin before suturing
Suture the area closed (Do NOT use glue)
Post-operative Care
5d
Place the mouse back into a clean cage on a warming pad and watch for the mouse to wake up.
Once awake place a Tylenol water bottle (1 mg/mL) on the cage for the next 5 days.
Monitor the mice once daily for open wounds or adverse side-effects, recording on the surgical card for 120:00:00 .
5d
After day 5 remove the water bottle.