Application for Protocol Approval Involving Laboratory Animal Use

UNIVERSIDAD CENTRAL DEL CARIBE APPLICATION FOR PROTOCOL APPROVAL INVOLVING LABORATORY ANIMAL USE Institutional Animal Care and Use Committee

Instructions for Filling out this Form

1. This application should be completed and submitted to the Institutional Animal Care and Use Committee (IACUC) in order to obtain authorization:

 for all use of vertebrate animals and/ or  to obtain tissues and cells from vertebrate animals, procured and sacrificed solely and specifically for research and teaching purposes.

2. The National Institutes of Health (NIH) require that funded grant applications including the use of vertebrates animals must have an approved IACUC protocol before funds are released to the Principal Investigator (PI) and affiliated institution.

3. All animal use protocols will require an annual update and must be re-submitted for re- evaluation every three years.

4. The electronic form of this application is available at (under construction) and has to be submitted via e-mail to at least one month before our scheduled meetings of IACUC. IACUC approval needs to be obtained prior to beginning the research project. Hardcopy applications will not be processed.

5. Applications submitted by postdoctoral fellows, graduate and undergraduate students should be accompanied by the name and contact information of his/her academic advisor on the certification section of this application.

6. All personnel included in the Application are required to have current animal handling procedure training.

7. Protocols involving an animal’s exposure to radioisotopes, biological and/or, chemical hazardous agents and tumor lines/body, and use of the rodent breeding must be accompanied with the appropriate attachments #1, 2, 3, 4 and 5.

1 Form: UCC/APAILAU-2014-06-02

1UNIVERSIDAD CENTRAL DEL CARIBE

APPLICATION FOR PROTOCOL APPROVAL INVOLVING LABORATORY ANIMAL USE

INSTITUTIONAL ANIMAL CARE AND USE COMMITTEE

FOR OFFICIAL USE

Identification Number:

A. INVESTIGATOR/PERSONNEL INFORMATION

Principal Investigator

1 Principal Investigator/PI Name: (Last name, first name)

2 Department extensio n

3 E-mail

4 Emergency Phone

Research Staff (List all individuals handling live/dead animals or animal tissue, including PI). ROLE IN ANIMAL TRAINING THE E-MAIL NAME Date General PROJECT Specie (s) Month/year Content

2 B. PROJECT INFORMATION

5 Title of Project

6 Type of Application

Research Training Pilot Other (specify)

Primary ( ) ( ) ( )

Additiona ( ) ( ) ( ) l

7 Protocol From (mm/dd/yy): ______To (mm/dd/yy):______Period

C. FUNDING SOURCES

Source 1 Source 2 Source 3 Institutional

8 Name of Department Director source Signature

9 Title grant ______1 Grant # 0

1 Start date 1

3 E. ANIMAL MODEL 12. Description of Animal to be used:

Animals must be obtained from Animal Resources Center approved sources/vendors only. Quarantine before entering UCC Animal House is required for certain animals based on their origin. Refer to the Handbook for the Use of Laboratory Animal or contact the Animal Resources Center administrative office 787-798-3001, ext. 2096 or [email protected] for further information. Note that protocol approval does not guarantee that the desired space for housing animals will be available.

Sources Species Strain Age Weight Sex Projected numbers ♀ ♂ # # Animals Animal /month /week

If rodent breeding is included in the study, Investigators who plan to breed rodents refer to Mice Breeding Protocol, as part of their study must complete the Rodent Breeding and Weaning Supplemental (Attachment #5).

E. CAGING AND HOUSING

13. Housing/ Identification of animals Sex Weight * Housing Maximum # of If an identification Strain (age) Location Animals Housed at method in addition (Animal One Time to a cage card is Facility) used (tattoo, ear

4 1 2 3 clip, punch method, year year year etc.). Describe

*For Satellite Animal Housing (holding animal’s outside for more than 12 hours) Request Policy on Satellite Animal Facilities.

General Specifications (mark with an X)

14 Is special caging required for holding animals more than 12 hours? ( )YES ( ) NO

15 Does caging deviate from the standards specified in either the ( )YES ( ) NO Guide for the Care and Use of Laboratory Animals (GUIDE) or the Animal Welfare Act?

16 Will special husbandry requirements be needed to accomplish this protocol (e.g. individual housing, special diets, water or feed ( )YES ( ) NO restrictions, treatments in feed or water, etc.)? If yes, provide the following: please describe details and scientific justifications below:

Specifications for Rodents The standard caging (microisolator caging is standard in this ( )YES ( ) NO 17 facilities) Special microisolators are needed since animals are 18 immunocompromised and/or would be exposed to biohazards ( )YES ( ) NO

Wire bottoms cages are required for this protocol. 19 (Rat weight should not exceed 500 grams and cannot be ( )YES ( ) NO housed for more 9 months)

5 20 Indicate any other special cages that are required in addition ( )YES ( ) NO to the ones mention above. (Provide justification below)

F. ANIMAL ENRICHMENT

The research requires species specific environmental cage ( )YES ( ) NO 21 enrichment devices and techniques? If you answered “yes, please describe and scientifically justify below:

Behavioral Testing Facilities support required.

Coordinator must sign.

G. USE SITES AND TRANSPORTATION METHODS

Will animals be transported out from the Animal Resources 22 Center? ( )YES ( ) NO (If you marked YES, provide justification, below)

Justification

Description of animal transportation route(s) and mechanism

23 Will live animals be returned after handling procedures outside the Animal Resources Center? ( )YES ( ) NO

Will vertebrate animals be housed in these alternate ( )YES ( ) NO 24 locations for more than 12 hours? (If yes, fill and attach special section, Satellite Animal Facilities to this application)

Indicate laboratory sites where hazardous tissue (e.g. animal tissues from studies involving radioisotopes, hazardous chemicals, or BSL-2 or higher) will be collected and 25 stored. Note that if the tissue (s) being transferred will be handled by investigators not listed on this protocol, you are required to inform that person (s) of any inherent danger associated with the tissues.

6 H. PROCEDURE NARRATIVE

26 Concisely describe in non-technical language the procedures to which animals will be exposed (in their sequential order) for duration of the project.

Procedure Procedure Name # of animals Description of the Procedure # required

27 Provide, whenever possible, flow charts/tables, illustrating experimental design and numbers of animals required for each procedure.

7 I. OFFICE OF LABORATORY ANIMAL WELFARE (OLAW) REQUIRED INFORMATION

If this protocol is duplicated, by LAW (PUBLIC LAW 99-158 ANIMAL RESEARCH SEC. 495 (http://grants.nih.gov/grants/olaw/olaw.htm) you must consider alternatives. In the following section, check which of the following databases and other sources have been searched to assure that the proposed experiments do not unnecessarily duplicate, that previous experiments do not cause undue distress or pain, and that alternative species or alternative methods have been considered and are not available. The search should include “refining” by using less stressful procedures, “replacing” the species selected, with one lower on the phylogenetic scale, and “reducing” the number of animals you are requesting. The search should have been conducted within the last 3 months.

Does this research duplicate any previous work? 28 ( )YES ( ) NO (If you marked YES, please explain below)

What is the scientific justification for the number of animals to be used (i.e. statistical validity, previous experience, etc.)? Please be aware of the following when planning your experiments:  Limit animal involvement by using the minimum number required to obtain reliable results.  Use non-animal methods, such as mathematical models, computer simulation, or in vitro biological systems if possible.  Use appropriate sedation, analgesia, or anesthesia when your procedures will cause more than momentary pain or distress. Do not perform surgical or other painful procedures on

8 non-anesthetized animals.  In addition, include information such as the number of control and experimental groups, number of animals per group or other reasoning.

29 Justify the rationale for animal use, including why non-animal models cannot be used.

30 Justify the selected model and why phylogenetically lower species were not selected.

31 Justification for total numbers of animal model.

No prior information is available to ( ) determine the numbers of animals required for completing the experiments. (The IACUC accepts requests to perform pilot studies to collect preliminary data to 32 Approach for Sample size fulfill this purpose, <10 animals total) determination ( ) Previous experience by this PI

( ) Previous literature

33 Describe power analysis to determine sample size

J. DRUG ADMINISTRATION

9 For all drugs used please note:

 Animals must be treated exactly as indicated in this protocol with regards to drug dose and duration of the treatment.  Changes in type of drug, dose, etc. must be approved via protocol amendment. If a veterinary recommendation results in a permanent change to the protocol, the investigator should submit an amendment of protocol to obtain IACUC approval.  Include clinical, laboratory or other parameters used to determine length of treatment.  Investigators are required to maintain treatment records on all animals. (Refer to Post Procedure Care of Mice and Rats in Research: Reducing and Distress)  For anesthesia, analgesia, tranquilizing and paralytic drugs specify dosage in mg/kg (provide justifications below and/or provide information and/or data that demonstrate the proposed)

34 NON-SURGICAL

Administration Species Drug Dose Route Frequency Strain (I.V., I.P, I.D.)

35 Duration and Monitoring Supply duration and method of monitoring treatment efficacy for drug listed above. Drug Duration Drug Efficacy Monitoring

36 Drugs administered for therapeutic purposes (excluding anesthetics, analgesics, and tranquilizers). SPECIES AGENT DOSE ROUTE EFFECT

10 (I.V., I.P, Strain I.D.)

Describe any adverse effects associated with the administration of these agents and a detailed plan for monitoring and alleviating these effects if applicable:

If you are using aquatics (e.g., frogs or fish), rats, and mice.

Are *non-pharmaceutical agents to be introduced into Animal Welfare Act (AWA) non- regulated species (rats, mice, birds, 37 ( ) YES ( ) NO or aquatics) for the purpose of veterinary medical care or to relieve pain and distress (e.g. anesthetics, analgesics, sedatives, antibiotics, or paralytics)?

Please note: As per Universidad Central del Caribe IACUC approved policy all compounds used in aquatics, rats, or mice for veterinary medical care or to relieve pain and distress (e.g. anesthetics, analgesics, sedatives, antibiotics or paralytics) must be pharmaceutical grade (when available) unless scientifically justified. Cost savings alone are not considered by the IACUC an adequate justification for using non-pharmaceutical grade drug for veterinary medical care or to relieve pain and distress in these species.

*Non- pharmaceutical agent: An agent that was not specifically prepared for injection into animal or human in its purchased form. If Yes, provide the scientific rationale for their use, source, proposed sterilization technique, sterile packaging, labeling and expiration information along with proposed methods to determine efficacy.

K. PROCEDURE DETAILS

11 Non-Surgical

Animal Handling Rationale

38 Will blood sampling be conducted? ( ) YES ( ) NO If yes provide rationale, method, site, volume frequency. Rationale Method Site Volume Frequency (ml)

39 Will food scheduling or restriction (other than standard pre- ( ) YES ( ) NO operative fasting) be conducted? If yes provide rationale, frequency and duration. Rationale Frequency Duration

40 Will water scheduling or restriction (other than standard pre- ( ) YES ( ) NO operative fasting) be conducted? If yes provide rationale, frequency and duration. Rationale Frequency Duration

41 Will restraint methods be utilized? ( ) YES ( ) NO If yes provide rationale, frequency and duration. Rationale Frequency Duration

42 SURGICAL

12 Route Administration Frequency Species Drug Dose (I.V., I.P, Strain I.D.)

43 Duration and Monitoring Supply duration and method of monitoring treatment efficacy for drug listed above. Drug Duration Drug Efficacy Monitoring

Postoperative Plan (Methods to assess/alleviate pain/distress, recovery criteria, monitoring criteria.

Will two or more of these survival surgeries involve major 44 procedures (defined as a surgical intervention that penetrates ( ) YES ( ) NO and exposes a body cavity or any procedures that produce substantial or permanent impairment of physical or physiological function?

Federal guidelines specify that no animals are to be used in more than one major survival operative procedure except in cases of scientific necessity or to provide adequate veterinary care.

J. ANIMAL EXPOSURE: These sections will be compared to applicable safety committee (s) regulations and should be consistent between documents.

45 Radioisotopes (Prior to any study involving the administration of radioisotopes to animals, the Radiation Safety Office must review and approve the use). 13 Will radioisotopes substance be administered in live animal? ( ) YES ( ) NO If you marked YES, complete attachment # 1 and provide the next information. Species Isotope Activity Dose Location of Use Strain

46 Biological Agents (Introduction of any genetically modified tissues, cells, viruses, or vectors into animals requires approval from the Institutional Biosafety Committee).

Will biological agents be used in living hold or animal tissue? ( ) YES ( ) NO

If you marked YES, complete attachment #2 and provide the next information. Species Biological Dose Route Effect Strain agent

Describe any adverse effect associated with the administration of these agents and a detailed plan for monitoring and alleviating these effects if applicable:

47 Chemical Hazardous Agents (Do not include drugs used for therapeutic purpose) (Prior to any study involving the administration of chemical agents to animals, approval from the Institutional Chemical Committee is required).

Will chemical agents be introduced to the living animals? ( ) YES ( ) NO

If you marked YES, complete attachment #3 and provide the next information.

Species Chemical Dose Route Effect agent

Describe any adverse effect associated with the administration of these chemicals and a detailed plan for monitoring and alleviating these effects if applicable:

14 48 Tumor Lines/Body (Introduction of any genetically modified tissues, cells, viruses, or vectors into animals require approval from the Institutional Biosafety Committee).

Will tumor lines or body fluids be introduced to the live animals? ( ) YES ( ) NO

If you marked YES, complete attachment #4 and provide the next information. Species Agent Dose Route Effect

K. EUTHANASIA

( ) YES ( ) NO 49 Will animals be euthanized

50 If YES, provide details of euthanasia procedure.

See http://www.avma.org/issues/animal_welfare/euthanasia.pdf for approved euthanasia methods AVMA Guidelines 2007

L. LITERATURE SEARCH Instructions: Provide at least 2 keywords and delimiters used to search the literature for reference on the background and experimental procedures addressed in this protocol. Check (X) keywords and Database Date Search Years Covered delimiters Performed Medline Highwire.org Schoolar.google.com Current Research Information (CRIS) EMBASE Other

M. PRINCIPAL INVESTIGATOR CERTIFICATION. Your electronic signature or e-mail response on this protocol form certifies your agreement

15 to the following terms.

 I certify that the research proposed herein does not unnecessarily duplicate any previous investigations.

 I confirm that the IACUC will be notified before initiating any changes in this protocol.

 I certify that IACUC will be informed of any unexpected results that impact animals in use. Any unanticipated pain, distress, morbidity or mortality will be reported to the attending veterinarian.

 I understand that in the event that I cannot be contacted, any animal that shows evidence of distress, illness or pain, emergency care, including euthanasia if necessary, will be taken care by the veterinary medical staff.

 I ensure that I will inform any recipient (not listed on section A) of any hazard associated with the use animal body fluids or tissue, derived from the studies described in this protocol.

 I certify that I will hand copy of the approved protocol to personnel identified in the section A and will assure that they become familiar with details contained in this protocol.

Today’s Date (mm/dd/yy)

Signature

N. CHECKLIST (Please respond by placing an X in the appropriate box).

Attachments General Sections (required) X Include X

Investigator/Personnel Information Radioisotopes Use #1

Project Information Biological Hazard #2

Funding Sources Chemical Agents #3

Animal Model Tumor Lines/Body #4

Caging and Housing Rodent Breeding and Weaning #5 Supplement

16 Animal Enrichment

Use Sites and Transportation Methods

Procedure Narrative

Animal Welfare Act Required Information

Drug Administration

Procedure Details

Animal Exposure

Euthanasia

Literature Search

Certification

FOR OFFICIAL USE ONLY

COMMITTEE MEMBER COMMITTEE COMMENTS NAME MEMBER SIGNATURE

1. Dr. Pedro Ferchmin

2. Dr. Krishna Baksi

3. Dr. Priscilla Sanabria

4. Dr. Legier Rojas

5. Dr. Juan Amieiro Veterinarian Consulting

17 6. Dr. Misty Eaton

7. Dr. Thomas Schikorski

8. Mrs. Vivian Valiente, Community Member

9- Dr. Luis Cubano

10- Lic. Betzaida Torres, Supervisor Animal Resource Center 11. Gerardo Torres Student Representative (Class 2016)

ACTION Approved with Approved ( ) modifications ( ) Disapproved ( )

Prof. Zilka Ríos Date (mm/dd/yy) President of UCC/IACUC

IACUC Attachment 1 Form: UCC/IACUC/URAAS-2012

USE OF RADIOISOTOPES ACTIVITY IN ANIMAL STUDIES

Approval must be obtained from the IACUC before this study may be performed. Prior to any study involving the administration of radioisotopes to animals, the Radiation Safety Office must review and approve the use. This form will be forwarded to the Radiation Safety office by the IACUC.

1 Principal Investigator e-mail 2 Title of Project

3 Strain Radionuclide/ # of animals # of animals Dose Route compound /study experiment/stud y

4 Project start date

18 5 Period of time over which experiments will be performed 6 Location where Room# Building administration will occur: 7 Location where radio- Laborator Animal facility room labeled animals will be y housed 8 Period of time before animal will be sacrificed 9 Are pathogenic/infectious agents to be administered also? ( ) YES ( ) NO REQUIREMENTS:

UNLESS AN EXEMPTION IS GRANTED BY THE RADIATION SAFETY OFFICE, THE FOLLOWING REQUIREMENTS ARE MANDATORY: 1. In the Animal Resource Center, all cages with animals must be labeled with a “Caution Radioactive Material” label stating the isotope and amount per animal, the date of administration, and the authorized user name. 2. Animal cages must be properly surveyed for contamination before being returned to the Animal Resource Center washer. 3. Animal carcasses that are disposed of as radioactive waste must be packaged separately. Do not include any other material such as pads, tubing, needle, instruments, etc. with the carcass. Signature of Investigator Date(mm/dd/yy)

Reviewed by Date(mm/dd/yy)

Special Conditions and Requirements

IACUC Attachment #2 Form: UCC/IACUC/ UBAA -2012

USE OF BIOLOGICAL ACTIVITY IN ANIMAL STUDIES

Approval must be obtained from the IACUC before this study may be performed. Prior to any study involving the administration of biological to animals, the Biohazard Committee must review and approve the use. This form will be forwarded to the Biohazard Safety representative by the IACUC.

1 Principal Investigator e-mail

2 Title of Project Species Pathogen # of animals # of animals Dose Route 3 Strain /study experiment/study

19 4 Project start date 5 Period of time over which experiments will be performed 6 Location where administration will Room# Building occur: 7 Location where pathogen-labeled Laboratory Animal animals will be housed facility room 8 Period of time before animal will be sacrificed

REQUIREMENTS:

UNLESS AN EXEMPTION IS GRANTED BY THE BIOSAFETY SAFETY COMMITTEE, THE FOLLOWING REQUIREMENTS ARE MANDATORY: 1. In the Animal Resource Center, all cages with animals must be labeled with a “Caution Biosafety Material” label stating the pathogen and amount per animal, the date of administration, and the authorized user name. 2. Animal cages must be properly surveyed for contamination before being returned to the Animal Resource Center washer. 3. Animal carcasses that are disposed of as radioactive waste must be packaged separately. Do not include any other material such as pads, tubing, needle, instruments, etc. with the carcass.

Signature of Date(mm/dd/yy) Investigator

IACUC Attachment # 3 Form: UCC/IACUC/UCAAS-2013-12-12

USE OF HAZARDOUS CHEMICAL AGENTS IN ANIMAL STUDIES

Approval must be obtained from the IACUC before this study may be performed. Prior to any study involving the administration of hazardous chemical hazardous to animals, the Chemical Committee must review and approve the use. This form will be forwarded to the Chemical Committee representative by the IACUC. 1 Authorized User Name e-mail 2 Title of Project

3 Strain Hazardous # of # of animal Dose Route Chemical animals/study experiments/study Agent/compound

4 Project start date 5 Period of time over which experiments will be performed

20 6 Location where Room# Building administration will occur: 7 Location where chemical Lab.: Animal facility room agent animals will be housed 8 Period of time before animal will be sacrificed 9 Are pathogenic/infectious agents to be administered also? ( ) YES ( ) NO REQUIREMENTS:

UNLESS AN EXEMPTION IS GRANTED BY THE CHEMICAL COMMITTEE, THE FOLLOWING REQUIREMENTS ARE MANDATORY: 1. In the Animal Resource Center, all cages with animals must be labeled with a “Caution Chemical Agent” label stating the chemical and the date of administration, and the authorized user name. 2. Animal cages must be properly surveyed for contamination before being returned to the Animal Resource Center washer. 3. Animal carcasses that are disposed of as chemical agent should be packaged separately in special containers. Do not include any other material such as pads, tubing, needles, instruments, etc. with the carcass.

Submitted by Date(mm/dd/yy)

IACUC Attachment # 4 Form: UCC/IACUC/UCTAS-2012

REQUEST TO INTRODUCE CELL LINES, TRANSPLANTABLE TUMORS, BODY FLUIDS, SERUM, TISSUES, AND ANTIBODIES INTO ANIMALS.

21  Due to the risk of human or animal cell lines, transplantable tumors, body fluids, serum, tissues, and antibodies being contaminated with rodent viruses, which can cause disease outbreaks within the rodent’s colony or adversely affect research, the IACUC requires testing of all such material that is deemed to be at risk.

 Required testing is for pathogens to which the colony of recipient species is Specific Pathogen Free (SPF). See the “Cell Line Usage and Rodents Biological” policy on the IACUC web site for more details. Failure to complete any required testing prior to use in animal is considered a noncompliance issue and puts the whole program at risk.

 An investigator wishing to introduce any of the above biological into animal, must have an approved protocol that describes the use of these biological and receive written permission from the university consulting veterinarian or their designee before any animals may be ordered.

 To ensure rodent biosecurity, previously tested materials may be required to be retested depending on how the materials have been handled and stored since testing and if the original testing still meets the current policy standards. The need to retest previously tested biological materials will be reviewed as part of the standard three year IACUC protocol renewal process and upon submission of IACUC protocol modification.

 Introduction of any genetically modified tissues, cells, viruses, or vectors into animals require approval from the Institutional Biosafety Committee.

1 Principal Investigator e-mail

2 Title of Project List the category of the biological (cell line, transplantable tumor, body fluid, serum, 3 tissue or antibody):

4 Designation (i.e. ID#, name, description) Species cell lines, # of animals # animals Dose Route 5 Strain etc. /study experiment/stu dy

Description of Biological Indicate the category of the biological (cell line, 6 transplantable tumor, body fluid, serum, tissue or antibody):

22 7 Designation (i.e. ID#, name, description)

8 Histological type of material introduced

9 Species of origin of material introduced

10 Provide the name and contact information of the source (institution and/or individual) providing the biological.

11 Has this material been PCR, MAP or RAP tested for viral pathogens? Is yes, send test result to IACUC

Please describe how the biological is stored or propagated in you laboratory. In the case of a cell line please include the following information in your description: 12  Is culture medium containing rodent serum or rodent-derived feeder cells used during in vitro propagation? ( ) YES ( ) NO  Is the cell line incubated in the same incubator with other rodent or human cell lines? ( ) YES ( ) NO  Is the cell line ever manipulated at the same time in the same biosafety cabinet as other rodent cell lines? ( ) YES ( ) NO  Is the cell line frozen as a stock and used as needed or is it propagated in vivo or in vitro? ( ) YES ( ) NO REQUIREMENTS:

UNLESS AN EXEMPTION IS GRANTED BY THE BIOSAFETY SAFETY COMMITTEE, THE FOLLOWING REQUIREMENTS ARE MANDATORY: 1. In the Animal Resource Center, all cages with animals must be labeled with a “Caution Biosafety Material” label stating the pathogen and amount per animal, the date of administration, and the authorized user name. 2. Animal cages must be properly surveyed for contamination before being returned to the Animal Resource Center washer. Animal carcasses that are disposed of as radioactive waste must be packaged separately. Do not include any other material such as pads, tubing, needle, instruments, etc. with the carcass.

Signature of Investigator Date(mm/dd/yy)

IACUC Attachment # 5 Form: UCC/RBWS-2013-12-12

RODENT BREEDING AND WEANING SUPPLEMENT 23 A. Individual(s) Responsible for Colony Maintenance

1 P.I Name Department Emergency 2 Work Phone # Phone (after hours) #

3 E-Mail

4 Other Individuals Involved in Colony Maintenance (Must Also be Listed in Application for Protocol Approval Involving Laboratory Animal Use, section A)

5 Describe training and experience of these individuals as it relates to colony maintenance & breeding of animals:

B. Breeding Colony Justification

6 What strains will be bred?

Provide a justification for establishing and maintaining a breeding colony of animals at the Universidad Central del Caribe.

C. Housing and Husbandry Issues

7 Building

8 Are there any special husbandry requirements needed for ( )Yes ( ) No the maintenance of the colony?

24 If you answered yes, please describe the needs (e.g., special diets, special housing, immunocompromized strains, etc.).

D. Colony Management Information (Please provide the following information below. Request the Mice Breeding Protocol for additional instruction) 9 Breeding Scheme

( ) Harem Mating (polygamous)

Pair Breeding (monogamous) ( ) If this method is selected, describe how you will ensure that multiple litters do not occur within once cage?

( ) Timed (hand) Mating

( ) Other- Describe and provide a justification:

10 Weaning Plan (no greater than one litter is to be kept in a cage. The Principal Investigator is responsible for weaning unless otherwise contracted). ( ) Animal will be weaned at 21-28 days.

The breeding requires additional time for weaning (beyond 28 days). ( ) Please describe and justify below. You must include in your response how you will ensure that the density of animals does not exceed the Guidelines for Animal Care and Use.

11 Phenotype Information

Are there any health concerns associated with the development ( ) Yes ( ) No of the phenotypes for the strains described?

If YES, please describe the health concern (e.g., behavioral, anatomical and/or physiological) and describe how the health of these animals will be managed.

25 E. Genotyping and Other Colony Management Procedures The goal of this section is to identify techniques and procedures used to manage your colony.

12 Genetic Identification

Tail Clip- If clipping is done on animals over 28 days of age, local or general ( ) anesthesia is required. If this is the case what anesthetic method (s) will be used? (Specify to the right)

( ) Blood sample-describe the collection procedure:

( ) Anesthesia method (if applicable):

( ) Other, (e.g., ear notch, buccal swab). Specify:

13 Age of animals for genotyping

( ) 0-21 days (anesthesia is recommended)

( ) 21-23 days (anesthesia is highly recommended)

( ) 21-28 days (anesthesia is strongly recommended)

( ) >28 days and older (adult post-weaning-anesthesia is mandatory)

14 What method of animal identification will be used?

( ) Ear notch

( ) Ear tag

( ) Tattoo

( ) Microchip implant

26 ( ) Cage card identification only

15 Euthanasia

Will animals that cannot be utilized be euthanized in the same ( )Yes ( ) No manner as described in the IACUC Protocol Application?

IF no, please describe the alternate euthanasia method planned

F. Estimated Number of Animals to Establish and Maintain the Colony

A mechanism for tracking colony management should be employed to allow review during semiannual IACUC inspections. If you need assistance in estimating numbers, please refer to ILAR Guidelines for the Care and Use of Mammals in Neuroscience and Behavioral Research (2003): http://www.nap.edu/books/0309089034/html

17 Enter Estimates

Estimated number of weaned and adult animals to be subject to experimental manipulations. (Please specify number of animals by: week, month or per year).

Estimated number of suckling animals to be subject to experimental manipulations. (Please specify number of animals by: week, month or per year).

Estimated number of breeder held but not subject to experimental manipulations. (Please specify number of animals by: week, month or per year).

Estimated number of suckling animals to be euthanized at or prior to weaning, and not subject to experimental manipulation. (Please specify number of animals by: week, month or per year).

TOTAL ESTIMATED (Please specify number of animals by: week, month or per year):

FOR OFFICIAL USE ONLY COMMITTEE MEMBER COMMITTEE MEMBER COMMENTS NAME SIGNATURE

27 1. Dr. Pedro Ferchmin

2. Dr. Krishna Baksi

3. Dr. Priscilla Sanabria

4. Dr. Legier Rojas

5. Dr. Juan Amieiro Veterinarian Consulting

6. Dr. Misty Eaton

7. Thomas Schikorski

8. Mrs. Vivian Valiente , Community Member

9- Dr. Luis Cubano

10- Lic. Betzaida Torres, Supervisor Animal Resource Center 11- Gerardo Torres Student Representative (Class 2016)

ACTION Approved with Approved ( ) modifications ( ) Disapproved ( )

Prof. Zilka Ríos Date President of (mm/dd/yy) UCC/IACUC