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Use of MS-222 (Tricaine Methanesulfonate) for Fish Anesthesia and Euthanasia

Number: IACUC-POL-020 

Responsible Office: Office of Research and Creative Scholarship (ORCS) 

Applies to: Principal Investigators Conducting Animal Research or All animal research and teaching protocols involving live vertebrate animals  

1. Purpose 

This policy outlines institutional requirements for the humane use of MS-222 as an anesthetic and euthanasia agent in fish, and specifies proper waste disposal procedures for laboratory and field activities. 

2. Scope 

This policy applies to all research, teaching, and testing activities involving fish under IACUC oversight, regardless of location. 

3. Requirements and Training 

MS-222 is an IACUC-approved anesthetic and euthanasia agent for most fish species and may only be used when it is prepared at the correct concentration, buffered to physiological pH, and administered with appropriate monitoring and—when used for euthanasia—confirmation of death. Individuals using MS-222 must be listed on an approved IACUC protocol, have documented training in its preparation, buffering, species-specific considerations, monitoring, recovery, and confirmation of death, and must demonstrate competency before performing procedures independently. No alternative methods may be used without prior IACUC approval.  

4. Field Sedation  

MS-222 is the only FDA-approved drug for the sedation of fish in field settings within the United States. The commercially available product (Tricaine-S™, Western Chemical, Inc., Ferndale, WA) contains tricaine methanesulfonate as the active ingredient. However, field use is limited due to FDA requirements that fish intended for human consumption—including wild or feral fish that may later be caught and eaten—must undergo a 21-day withdrawal period before release or harvest (Anderson et al. 1997; Trushenski et al. 2012a, 2012b).  

5. Procedures 

Formulation of MS-222 

• The IACUC requires the use of pharmaceutical-grade MS-222 product (Tricaine-S, Syncaine®); however, if other MS-222 products are to be utilized, justification for the use of a non-pharmaceutical grade compound must be provided in the IACUC protocol. 

Safety Precautions 

• MS-222 is considered an irritant to the eyes, respiratory system, and skin. Safety precautions should be employed when used.
• To prevent exposure, the powder should be weighed in a fume hood or while wearing an N95 respirator. To avoid skin and eye contact, goggles, gloves and lab coat should be worn. Employees who are required to wear respirators must be fit tested through UM Environmental Health and Safety to ensure proper mask fitting and testing. 
• When handling liquid containing MS-222, PPE should include gloves, lab coat and eye goggles to prevent contact with eyes or skin. 

Preparation 

• MS-222 comes in a powdered form and should be stored at room temperature, in a cool dry place protected from light. 
• Ideally MS-222 stock solutions are utilized the same day as preparation per vendor recommendation. When necessary, stock solutions of MS-222 may be kept for up to 30 days. Stock solution must be refrigerated and stored in tinted (amber) or opaque bottles. 

Use as an Anesthetic 

• Solutions for bath immersion are acidic and irritating and must be buffered with sodium bicarbonate to a normal pH (7.0-7.4) before use. 
• The action of MS-222 as an anesthetic varies widely between species and is affected by water temperature, hardness, and size of the individual animal. Pilot tests should be conducted to determine concentration and exposure time for each application to ensure sufficient anesthetic depth and safe recovery. Doses given in the following section give a rough indication of the dose, but the particular regimen should be tailored to the experimental conditions through pilot experiments. 
  
  Dose:
  15-50 mg/L Sedation
  50-200 mg/L Induction
  50-100 mg/L Maintenance
  1 g/L Spray – large fish on gills directly 
  
  
• After procedures are completed, place animals in well-oxygenated/aerated, un-medicated water. 
• Closely monitor fish recovering from anesthesia until they are swimming normally, and completely regained their righting response. 

Use for Euthanasia 

• MS-222 can be used to euthanize fish. It is considered an acceptable method per the AVMA Guidelines on Euthanasia. 
• Immersion in buffered MS 222 for at least 30 minutes; it is recommended that this is followed by an adjunctive method (decapitation, pithing, exsanguination, freezing). 
• A concentration of 250-500 mg/L or 5 to 10 times the anesthetic dose is effective for most species. 

6. Disposal   

Laboratory Use 

• Used MS-222 should be collected in an acid-resistant hazardous waste container. The container must be properly labeled with the contents, accumulation start date, and a hazardous waste label, and stored with appropriate secondary containment. 
• When the waste container is full or the study has concluded, contact UM Environmental Health and Safety (EHS) at 406-243-4503 to arrange to pick it up for disposal. 
• UM EHS will coordinate disposal through CleanEarth Labpack system. 
• At the conclusion of the study, any remaining solid MS-222 should be collected and UM 
• EHS should be contacted for proper disposal. 

Field Use  

For MS-222 (2g/L) solution disposal in the field, the remaining anesthetic will be diluted at least 4:1 with clean water and dispersed at least 100 meters away from the stream, ensuring that no concentrated chemical enters surface waters.  

Disposal will follow all state and local environmental guidelines. 

7. Supporting Documents  

SOP Tricaine Methanesulfonate (MS-222) Use in Fish 

8. References 

1. The Guide for the Care and Use of Laboratory Animals, 8th Edition. National Research Council Committee for the Update of the Guide for the Care and Use of Laboratory Animals. Washington (DC): National Academies Press (US). 
2. AVMA Guidelines for the Euthanasia of Animals: 2020 Edition 
3. Guidelines for the Use of Fish in Research, Americal Fisheries Society: 2014.  

9. Review, Approval and Version History 

Review and approval version history for this policy

Version	Date	Description of Changes	Approved By
1.0	January 13, 2026	Initial policy creation	IACUC Committee