Protocol to document Animal Care Issues

Training information for personnel in the Division of Advanced Medical Imaging Research

(A) Purpose

The purpose of this overview document is to summarize laboratory policies for animal care and handling, especially as they pertain to animal health and use of hazardous agents for animal imaging procedures.  The detailed protocols for specific procedures are available, and included by reference in this document.

(B) Animal Health

1. Animals generally arrive at UAB from vendors that maintain colonies that are free of all diseases that would adversely affect our animal experiments.  Our goal is to insure that the animals remain free of disease.  In our laboratory we have the extra challenge of making certain that animals coming from one area of campus do not transfer disease agents to our facility.  We maintain a logbook (next to the IVIS system in VH B021 and WTI 630D) of all animal arrivals to our facility.  This includes the date, investigator’s name, and UAB location from where the animals came.  Please record this information.  This information will be valuable in the event of an unexpected disease outbreak.
2. If an animal appears ill, or dies unexpectedly, please inform Sharon Samuel or Dr. Lapi by e-mail or phone.  If they are un reachable, please inform an Animal Resource Program (ARP) staff member.  Store dead mice at 4^oC if necropsy is indicated, or in the ARP animal morgue.  Follow the UAB Carcass Disposal policy.
3. For all tumor studies, mice will be terminated if they become ill or suffering (lack normal grooming and avoidance behaviors), are unable to eat or drink, the tumor becomes overly large as to hinder normal body movement or ulcerates.  Tumor burden will not be allowed to exceed 10% of the normal body weight.  With ascitic volumes, the fluid accumulation will not be allowed to cause gross abdominal distension and solid deposits and cachexia will not be allowed to become clinically significant.  The ascitic volumes will not exceed 20% of the normal body weight.  Weight loss in tumor bearing animals will not be allowed to exceed 20% of the normal body weight.
4. We often routinely use extra animals as sentinels for disease control.  A program is utilized in the Animal Resources program to screen for common rodent diseases.   Check with Sharon if appropriate to send an extra animal for screening.

(C) Procedures to reduce the potential for spreading disease

1. Always wear appropriate attire, including lab coat, gloves, and if desired a hair net and mask.  Short pants and uncovered shoes (sandals) are not allowed.
2. Start with clean bench paper, and change paper after each experiment.
3. Clean all imaging equipment, anesthesia chamber, nose cones, etc. with disinfectant before handling the animals.
4. If you notice that any person working in our facilities is not following the established procedures, please inform Sharon Samuel for corrective actions.

(D) Regulatory Requirements

1. Only trained individuals are allowed to conduct procedures with the animals.
2. The IACUC has approved for mice to be housed in WTI 650D and VH B54K.  Animals cannot be outside of room 650D or B54M for longer than 12 hours.  Animals can be kept in VH B21 or WTI 650D for </=12 hours only.
3. All cages must be marked with a cage card that includes the investigator, IACUC protocol number, and account number.
4. Please call Sharon Samuel or an Animal Resources staff member if you notice any problems with the animal room ventilation, temperature, isolator motors, etc. If after hours, then have the UAB operator (934-4011) page the veterinarian or animal care staff member that is on-call.
5. Any hazardous materials (viruses, radiation, chemicals, etc.) require special review and approval. Each AUSI form is specific for a particular IACUC approval number, so make certain the correct form is used.  All forms are kept in the black AUSI notebook on the spare shelving in B54K.
6. Additional training documentation is needed for imaging mice injected with infectious bacteria.
7. Cages containing mice dosed with biohazardous agents must be labeled with a “Biohazard” sticker.
8. Inform the ARP building supervisor and Earle Durboraw when mice will be dosed with a radioactive agent.  They would like to have at least a 2 days notice for this in case certain changes may need to be made to the room or additional materials need to be placed there for subsequent changes.
9. Cages containing mice dosed with radioactive agents must be labeled with a radioactive sticker that includes the date, radioisotope given, and amount in millicuries.  Radioactive stickers must be eventually removed from the cages before return to ARP staff.  The stickers be disposed in the radioactive trash, and not placed in regular trash.
10. Cages containing dosed mice are check by our lab daily until the decay time has passed (3d for Tc-99m, 4d for Pb-212, and 1d for F-18).  ARP supplies us with the daily care sheet that we initial and check.  Additionally, additional signage is required on any shielding for the cages.
11. Cages with radioactive mice are not changed by ARP staff.  It is our responsibility to change cages.  With Tc-99m, it is possible to wait 3 days for decay and then the cages can be changed by ARP staff. 
12. All cages containing radioactive agents are surveyed prior to release back to ARP staff.  When you survey a cage, record this information on the Radioactive Survey form.  There is a space on the form for this information.
13. Animals can be transported between our laboratory and other UAB facilities, but only following the UAB Animal Transportation Policy.
14. Animal euthanasia must be conducted according to the UAB Animal Euthanasia policy.
15. Animal carcasses are handled according to the UAB Carcass Disposal policy.  Please note that in our laboratory we dispose of all animals as if they contained radioactivity, whether they have been given radioactivity or not.  In this manner, we insure that all mice are incinerated and the ash is monitored.  Store dead mice in the –20 C freezer in VH B021 in marked plastic bags.
16. Never allow strangers access to the animal facilities, and report any suspicious activities immediately to the UAB police.
17. Do not take any pictures or videos of the mice unless approved to do so by Sharon Samuel, Dr. Lapi or ARP administration.

(E)  Imaging Procedures

1. There are separate protocols for each imaging modality.  Use the appropriate protocol to collect imaging data, but only after proper training has been accomplished.

(F)Use of Radioactive Materials in Animals

(Note: This section is used as a supplement for page 12 of the IACUC review form, and details the lab’s commitment to proper use of radioactive materials.  Dr. Lapi’s Radioactive Materials License Number is  #162.)

1.  Animal Housing and radioactive material protocols:

All animal rooms and cages containing radioactive animals are posted with "CAUTION RADIOACTIVE MATERIAL" signs, and rooms are restricted to authorized personnel who have taken the radiation safety course.  Animals are housed at a distance from the radioactivity stock solutions and waste, to minimize exposure.  After each experiment, a radiation survey procedure is followed to check for contamination.  Cages and bedding are handled according to a Radioactive Waste Protocol.  A radioactive storage "safe" with 4-inch thick concrete walls is available to store higher activity samples.  Per the protocol, the cages are either stored until no detectable radioactivity exists, or they are decontaminated.  Results of the cage surveys are recorded on the Radiation Survey Form for the laboratory, per the Radiation Survey Protocol. The animal carcasses are all double bagged, labeled, and frozen in a designated and properly labeled radioactive freezer (Room VH G073).  Any solid waste produced, including absorbent pads and gloves, are stored double bagged, and disposed as radioactive waste.  All liquid waste is stored in plastic or glass container on absorbent paper in a shielded fume hood, or in the "safe".  All radioactive waste is maintained in a safe location, away from well-traveled areas.  All radioactive waste is disposed in accordance with applicable regulatory requirements as stated in the Radiation Safety Procedures Manual.  Protocols related to use of radioactive materials in animals are provided to Animal Resources as necessary to insure compliance with applicable regulations.

2.  Imaging and Biodistribution Studies:

        Syringes for injections of radioactive materials into animals are prepared in 50 - 200 ml volumes, measured in a dose calibrator (AtomLab 100, or Capintec) both before and after injection.  Prior to, and after injections, the syringes are kept in a lead container to minimize dose.  The animals are injected in VH B021A, VH G082G, or WTI 630D3.  Most injections are intravenous, although occasionally other routes are used, as according to the applicable IACUC protocol.  All used syringes are stored in radioactive labeled plastic containers.  Sharps containers are packaged in a labeled cardboard box for radiation waste pickup.  Syringes and vials containing viral vectors are rinsed with Clorox before disposal.  Animal imaging is conduced in VH B021A and WTI 630D3.  For imaging, established protocols for planar gamma camera imaging, microPET/CT, and SPECT/CT imaging are followed.

        A biodistribution protocol is used to collect organs.  The animals are anesthetized, blood taken via cardiac puncture, with cervical dislocation to insure death.   Syringes are put in radioactive labeled sharps containers and disposed of as radioactive waste.  The required tissues are harvested and placed into pre-weighed vials.  All samples are counted in gamma counter.  After counting, the vials with organs and the dead animal carcasses are placed in double bagged radioactive bags and stored in the assigned radioactive freezer until picked up by Radiation Safety waste department.  Cages of animals injected with replication competent viral vectors are held for decay (10 half lives of the isotope used), surveyed, autoclaved, and returned to Animal Resources.  Other cages with radioactive contamination are held for decay, surveyed, and return to Animal Resources without autoclaving.

3.  ALARA Principles are followed:

All procedures are established to minimize dose to personnel.  Numerous lead bricks are used for shielding to minimize radiation dose.  Proper monitoring equipment, including a Geiger counter and Keithley radiation survey meter (Model 3615), is available to accurately measure dose rates.  Body dosimeters are worn by personnel.