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Institutional Animal Care and Use Committee
Protocol Application
PI Last Name:
PI First Name:
Protocol Title: New breeding protocol for conditional deletion in mice
Directions: Please answer all questions as completely as possible. Fields will expand to allow for unlimited text to
be answered in all descriptive question fields. You may cut and paste text into fields. Any figures may be appended
in IRBNet as separate documents.
Administrative Information
1. In conducting this project, I agree to have my laboratory and all animals assigned to this protocol available for examination
by IACUC members or Department of Comparative Medicine personnel at all times during the project. I understand that
according to the regulations under which the IACUC is authorized, any time any animal is deemed to be in unacceptable
distress or is receiving inhumane treatment, in the professional opinion of the attending veterinarian, the animal may be
subjected to immediate euthanasia and/or the procedure immediately halted. The IACUC will be informed of any action
taken. The Director of University Biological Resources may suspend a protocol, for cause, until the IACUC has reviewed the
incidents leading to the suspension.
If the animals assigned to this protocol are supported by a grant or contract, I further certify that the procedures described in
this protocol agree with the procedures described in the grant or contract application. I agree to provide the IACUC with one
copy of my grant or contract proposal to allow verification that the description of the animal use portion of the grant or
contract agrees with the protocol. I understand that changes in animal use procedures during the grant or contract period
must be described on the annual monitoring/review form.
I understand that IACUC approval of this protocol does not guarantee availability of resources or services from the
Department of Comparative Medicine.
I certify the information provided herein is correct. I agree to accept responsibility for this project in accordance with all
federal, state and local laws and regulations, NIH/ILAR guidelines, PHS Animal Welfare Assurance, and institutional policies
and procedures.
I certify I have read and agree to the above:
I agree
I disagree
Objectives
Please describe your objectives (255 characters MAX)
The objective is to breed mice that lack expression of CCTalpha, the rate-limiting enzyme in phosphatidylcholine synthesis,
specifically in B cells. These mice will be used to study phosphatidylcholine production in B cells.
Rationale
Please describe your rationale for pursing the following protocol procedures (500 character MAX)
B cells produce antibodies that protect the body against infection. However, B cell cancers and antibody-mediated
autoimmune diseases are major health problems. In activated B cells, there is a large increase in the synthesis of membrane
lipids such as phosphatidylcholine. Therefore, we plan to further investigate the mechanisms that regulate lipid biosynthesis in
activated B cells and explore their potential roles in determining the function and/or fate of activated B cells.
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Description
Please provide a general description of the following procedures.
This protocol concerns the breeding of mice to generate pups in which the CCTalpha gene is conditionally deleted in B cells.
The protocol is composed of 2 parts. In Part 1, the CD19-Cre model will be used to generate mice in which progenitor B cells
undergo deletion of the CCTalpha gene at a very early step in development. In this case, the B cells "grow up" as CCTalphadeficient cells. In Part 2, the Cgamma1-Cre model will be used to generate mice in which mature B cells undergo deletion of
the CCTalpha gene only after activation.
Part 1: CD19-Cre mediated deletion of CCTalpha in progenitor B cells
Expression of the CD19 gene begins at a very early step in B lymphocyte development in the bone marrow. Therefore, in the
CD19-Cre model, floxed genes are deleted in progenitor B cells in the bone marrow. In our previous breeding protocol for
these mice (#09003), we developed CCTalpha flox/flox; CD19 Cre/wt mice. These mice will be maintained as breeder pairs, and
their litters will yield the pups needed for experimental purposes (experimental protocol #09031). We expect approximately 8
pups/month from each breeder pair, usually consisting of 4 CCTalpha flox/flox; CD19 Cre/wt pups (B cells lack CCTalpha), 2
CCTalpha flox/flox; CD19 wt/wt pups (B cells express CCTalpha), and 2 CCTalpha flox/flox; CD19 Cre/Cre (B cells lack both
CCTalpha and CD19, not useful experimentally, these pups will be euthanized by CO2 euthanasia followed by cervical
dislocation). However, this distribution can vary. Pups will be genotyped by PCR analysis of genomic DNA isolated from tail
clippings. Tails will be clipped before 21 days of age, so anesthesia will not be required. Tail clipping will be done using sterile
instruments on each animal. A 1 cm tail snip will be obtained and the pups will be monitored for excessive bleeding. A hole
punch will be used to mark the pups' ears for identification. Tail clipping and ear punching will be performed by trained DCM
personnel according to standard operating procedures.
Part 2: Cgamma1-Cre mediated deletion of CCTalpha in activated B cells
The Cgamma1 promoter becomes transcriptionally active only in activated B cells that have initiated isotype switching.
Therefore, in the Cgamma1-Cre model, floxed genes are deleted only in mature B cells that have been activated. In our
previous breeding protocol for these mice (#09003), we developed CCTalpha flox/flox; Cgamma1 Cre/wt mice. These mice will
be maintained as breeder pairs, and their litters will yield the pups needed for experimental purposes (experimental protocol
#11027). We expect approximately 8 pups/month from these breeder pairs, usually consisting of 4 CCTalpha flox/flox;
Cgamma1 Cre/wt pups (activated B cells lack CCTalpha), 2 CCTalpha flox/flox; Cgamma1 Cre/Cre (activated B cells lack
CCTalpha) and 2 CCTalpha flox/flox; Cgamma1 wt/wt pups (activated B cells express CCTalpha). However, this distribution can
vary. Genotyping, tail clipping and ear punching will be done as described in Part 1 above.
Breeder Replacement
We will replace breeder pairs every 6 months. Retired breeders will be euthanized by CO2 euthanasia followed by cervical
dislocation. Based on the previous 3 years of experience with these mouse models, we expect these mice to be healthy and to
thrive under normal conditions. Thus, we do not expect health problems to arise in these animals. However, if at any time prior
to the 6 month retirement, a breeder animal appears distressed as evidenced by loss of appetite, weight loss or lethargy, we
will euthanize the animal by CO2 euthanasia followed by cervical dislocation. To replace retired breeders, we will use CCTalpha
flox/flox; CD19 Cre/wt mice and CCTalpha flox/flox; Cgamma1 Cre/wt mice that have been set aside for this purpose from
previous litters.
2. Describe the methods used to determine the number of animals needed for this project. List statistical tests or other methods
used to determine these numbers.
To maintain the colonies and generate the pups needed for experimental purposes, we anticipate needing 4 CCTalpha flox/flox
CD19 Cre/wt breeder pairs (Part 1) and 4 CCTalpha flox/flox; Cgamma1 Cre/wt breeder pairs (Part 2) at a time. This will total 8
breeder pairs/year for Part 1 and 8 breeder pairs/year for Part 2. Assuming that a 30% failure rate is possible, we would like to
plan for 11 breeder pairs/year for each Part (11 breeder pairs/yr x 3 yrs = 33 pairs x 2 parts = 66 pairs). Therefore, for this 3
year protocol, we request a total of up to 66 breeder pairs (132 mice).
Confidential- USA Internal Use Only
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Alternative Methodologies
3. Describe what alternatives to the use of animals were considered (e.g., computer modeling, tissue/cell culture, etc.) and
explain why they cannot be used in place of this animal use protocol.
Our goal is to further investigate the roles CCTalpha and phosphatidylcholine synthesis in the development and function of B
cells in vivo. Therefore, mice with B cells that either express or lack CCTalpha are required. The complex processes that
regulate B cell development, activation and fate are only partly understood and cannot be fully modeled using cell lines in vitro
and/or computer modeling at this time.
4. State why a lower order would not be used in this study. Ex. mice instead of swine.
The mouse remains the lowest order species appropriate for studies of immune cell development and function.
5. Describe considerations given to the use of alternatives to painful procedures which will be performed on the animal(s) in this
project. If such alternative procedures cannot be used, provide justification for the use of painful procedures.
There are no painful procedures in this breeding protocol.
Non-Duplication of Research
6. Has the desired information been previously published?
No
7. Regarding this proposed project, indicate which databases were searched, the keywords used for the search, and the date(s) of
those searches.
PubMed, January 9, 2012
Education, Training, and Assurances
8. I certify that each person associated with this project has completed the IACUC-mandated training in the appropriate
laboratory animal research techniques and procedures outlined in this submission. I further assure that documentation in
support of this training is on file in the IACUC office (with a copy maintained in the animal use laboratory).
Please indicate Yes training is complete and records on file or No, read 8a.
Yes
8a. If 'No', then the individual(s) cannot be involved in the animal-related portions of this project until
such time as the appropriate training has been completed and documentation forwarded to the IACUC office.
9. Are all personnel associated with the animal-related portions of this project enrolled in the College of Medicine Occupational
Health Program (OHP)?
Yes
9a. If 'No", identify personnel NOT participating in the Occupational Health Program (OHP). NOTE: A waiver must
be signed by each individual not participating in the OHP. The signed waiver must be on file in the Research
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Compliance office. Individuals not participating in the OHP may be prohibited from working with certain species.
Please list the names of those with waivers.
Survival Surgery
10. Will any animal be subjected to a survival surgical procedure ( a surgical procedure from which the animal is EXPECTED TO
RECOVER from anesthesia)? Please indicate Yes or No.
No
10a. If 'Yes', Survival Surgery will be performed under which of the following conditions?
Aseptic - REQUIRED FOR cats, dogs, rabbits, non-human primates, and livestock: requires a dedicated surgical environment, sterile
instruments, and sterile garments
Non-Aseptic
- Acceptable for rodent surgery; requires clean surgical environment using sterile surgical instruments.
10b. If 'Yes', At what locations will survival surgery be performed.
Please indicate building and Room numbers for all sites.
11. Provide a detailed description of the survival surgical procedure. Please provide the appropriate information under each of the
italicized headers below (boxes will expand to fit content).
Type of survival surgical procedure to be performed:
Name ALL the personnel to be involved in the survival surgery:
Anesthetic Regimen (anesthetic agent, dose, and route of administration):
Method of assuring surgical plane of anesthesia is reached and maintained:
Post-operative care (include anticipated recovery time and location, medications, analgesics, post-op procedures, and names of personnel
who will provide post-op care):
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12. I understand that I am responsible for post survival care of my animals, unless informed otherwise by DCM.
Please indicate Yes to signify your understanding.
13. Will major survival surgery be performed more than once to any single animal?
Please indicate Yes to signify your understanding.
13a. If 'Yes', please explain/Justify the need for multiple survival surgeries on a single animal.
Segment Information
Segment Number: (Ex. 1, 2, 3,...) 1
Segment USDA Category C
Species: Mice
IF other 'other', please indicate:
Genetic Designation: CCTalpha flox/flox; CD19 Cre/wt
Allow Transfer:
No
Strain/Stock/Breed: CCTalpha flox/flox; CD19 Cre/wt (on C57BL/6
If transfer, to what Protocol#ȱȱȱ
Number Authorized: 66
Euthanasia Primary:
CO2
Euthanasia Secondary:cervical dislocation
Segment Description (field will expand to accommodate content):
These mice will be used as breeders to generate pups for conditional deletion of CCTalpha in progenitor B cells.
S1. Specify the Age and/or weight of the animals to be used in this segment of the project:
adult mice
S2.. Specify the sex of the animals to be used in this segment of the project:
Both
S3. Will any of the following requirements or conditions apply to the animals to be procured for this project: lactating (with or
without litter), Neutered/Spayed, Proven or Retired Breeder, Untimed pregnant, Timed Pregnant, Genetically altered,
Surgically altered, or Other similar characteristics.
Please indicate Yes, the animals have at least one of these characteristics or No, none have these characteristics.
Yes
S3a. Fully describe the special conditions, alterations, or modifications for this segment of animals.
(The box will expand to fit content).
Mice hemizygous for the Cre insertion at the CD19 locus (CD19 Cre/wt) retain one functional CD19 allele and have
Confidential- USA Internal Use Only
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phenotypically normal B cells. Several years of work with these mice indicates that these animals experience no
deleterious effects related to Cre-mediated deletion of CCTalpha in B cells.
Animal Care and Husbandry
S4. Will animals be subjected to food or water restriction/deprivation (other than peri-operative fasting) during this project?
No
S4a. Describe the food or water restriction/deprivation regimen. Be sure to include a discussion of how the restriction
will be monitored, what actions will be taken to remedy adverse effects of such restrictions/deprivations, and
what (and where) records will be maintained.
S5. Are there any special husbandry requirements (food, bedding, caging, room temperature, lighting, or water) for the animals
described in this segment of the project?
No
S5a. Fully Describe the special Husbandry requirement(s).
S6. Is this a breeding protocol from which offspring will be transferred to a different protocol after weaning?
Yes
S6a. If ''Yes', please provide a valid protocol number to which offspring from the breeding protocol are to be
transferred. If that experimental protocol is under review concurrently, please indicate.
######
Transportation of Animals
S7. If animals assigned to this project require specific investigator attention or are found dead, should authorized personnel be
contacted outside of regular working hours? If 'Yes', every attempt will be made to contact you; however, DCM is not responsible if you cannot be reached.
No
S8. If animals assigned to this project DIE, should they be saved for your examination?
Yes
S8a. If 'Yes', please select your preferred storage method for the animal prior to your examination:
Refrigerate
Other (Contact DCM):
Segment Number: (Ex. 1, 2, 3,...) 2
Species: Mice
Segment USDA Category C
IF other 'other', please indicate:
Genetic Designation: CCTalpha flox/flox; Cgamma1 Cre/wt
Allow Transfer:
No
Strain/Stock/Breed: CCTalpha flox/flox; Cgamma1 Cre/wt (on C57
If transfer, to what Protocol#
Confidential- USA Internal Use Only
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Number Authorized: 66
Euthanasia Primary:
CO2
Euthanasia Secondary:cervical dislocation
Segment Description (field will expand to accommodate content):
These mice will be used as breeders to generate pups for conditional deletion of CCTalpha in activated B cells.
S1. Specify the Age and/or weight of the animals to be used in this segment of the project:
adult mice
S2.. Specify the sex of the animals to be used in this segment of the project:
Both
S3. Will any of the following requirements or conditions apply to the animals to be procured for this project: lactating (with or
without litter), Neutered/Spayed, Proven or Retired Breeder, Untimed pregnant, Timed Pregnant, Genetically altered,
Surgically altered, or Other similar characteristics.
Please indicate Yes, the animals have at least one of these characteristics or No, none have these characteristics.
Yes
S3a. Fully describe the special conditions, alterations, or modifications for this segment of animals.
(The box will expand to fit content).
The Cre insertion at the Cgamma1 locus (Cgamma1 Cre/wt) does not disrupt a gene and, therefore, has no
deleterious effects on the mice. In addition, several years of work with these mice indicates that these animals
experience no deleterious effects related to Cre-mediated deletion of CCTalpha in B cells.
Animal Care and Husbandry
S4. Will animals be subjected to food or water restriction/deprivation (other than peri-operative fasting) during this project?
No
S4a. Describe the food or water restriction/deprivation regimen. Be sure to include a discussion of how the restriction
will be monitored, what actions will be taken to remedy adverse effects of such restrictions/deprivations, and
what (and where) records will be maintained.
S5. Are there any special husbandry requirements (food, bedding, caging, room temperature, lighting, or water) for the animals
described in this segment of the project?
No
S5a. Fully Describe the special Husbandry requirement(s).
S6. Is this a breeding protocol from which offspring will be transferred to a different protocol after weaning?
Yes
S6a. If ''Yes', please provide a valid protocol number to which offspring from the breeding protocol are to be
transferred. If that experimental protocol is under review concurrently, please indicate.
######
Confidential- USA Internal Use Only
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Transportation of Animals
S7. If animals assigned to this project require specific investigator attention or are found dead, should authorized personnel be
contacted outside of regular working hours? If 'Yes', every attempt will be made to contact you; however, DCM is not responsible if you cannot be reached.
No
S8. If animals assigned to this project DIE, should they be saved for your examination?
Yes
S8a. If 'Yes', please select your preferred storage method for the animal prior to your examination:
Refrigerate
Other (Contact DCM):
Segment Number: (Ex. 1, 2, 3,...) 3
Segment USDA Category C
Species: Mice
IF other 'other', please indicate:
Genetic Designation: weanlings
Allow Transfer:
Strain/Stock/Breed: weanlings (on C57BL/6 background)
Yes
If transfer, to what Protocol#ȱȱȱȱȱȱǛǛǛǛǛǛȱŠ—ȱǛǛǛǛǛǛ
Number Authorized: 2,000
Euthanasia Primary:
CO2
Euthanasia Secondary:cervical dislocation
Segment Description (field will expand to accommodate content):
This Segment is for weanling mice prior to genotype determination and assignment to an experimental protocol.
S1. Specify the Age and/or weight of the animals to be used in this segment of the project:
3 to 5 weeks of age
S2.. Specify the sex of the animals to be used in this segment of the project:
Both
Confidential- USA Internal Use Only
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S3. Will any of the following requirements or conditions apply to the animals to be procured for this project: lactating (with or
without litter), Neutered/Spayed, Proven or Retired Breeder, Untimed pregnant, Timed Pregnant, Genetically altered,
Surgically altered, or Other similar characteristics.
Please indicate Yes, the animals have at least one of these characteristics or No, none have these characteristics.
Yes
S3a. Fully describe the special conditions, alterations, or modifications for this segment of animals.
(The box will expand to fit content).
Mice hemizygous for the Cre insertion at the CD19 locus (CD19 Cre/wt) retain one functional CD19 allele and have
phenotypically normal B cells. The Cre insertion at the Cgamma1 locus (Cgamma1 Cre/wt) does not disrupt a gene
and, therefore, has no deleterious effects on the mice. There are no deleterious effects related to Cre-mediated
deletion of CCTalpha in B cells.
Animal Care and Husbandry
S4. Will animals be subjected to food or water restriction/deprivation (other than peri-operative fasting) during this project?
No
S4a. Describe the food or water restriction/deprivation regimen. Be sure to include a discussion of how the restriction
will be monitored, what actions will be taken to remedy adverse effects of such restrictions/deprivations, and
what (and where) records will be maintained.
S5. Are there any special husbandry requirements (food, bedding, caging, room temperature, lighting, or water) for the animals
described in this segment of the project?
No
S5a. Fully Describe the special Husbandry requirement(s).
S6. Is this a breeding protocol from which offspring will be transferred to a different protocol after weaning?
Yes
S6a. If ''Yes', please provide a valid protocol number to which offspring from the breeding protocol are to be
transferred. If that experimental protocol is under review concurrently, please indicate.Ǜ
#####and ######
Transportation of Animals
S7. If animals assigned to this project require specific investigator attention or are found dead, should authorized personnel be
contacted outside of regular working hours? If 'Yes', every attempt will be made to contact you; however, DCM is not responsible if you cannot be reached.
No
S8. If animals assigned to this project DIE, should they be saved for your examination?
Yes
S8a. If 'Yes', please select your preferred storage method for the animal prior to your examination:
Other (Contact DCM):
Confidential- USA Internal Use Only
Refrigerate
This form results from the IRBNet Online Document Wizard and is REQUIRED!
University of South Alabama Institutional
Animal Care and Use Committee Institutional
Biosafety Committee
IACUC/IBC Application Form
Last edited by:
IACUC
Last edited on: January 23, 2012
IBC
[xxxxxxx-x] New breeding protocol for conditional deletion in mice
I.
Principal Investigator
Name:
College:
Department:
Office:
Room:
Office Phone:
Email:
After Hours:
Cell:
II. Additional Personnel
N/A
Please include all those who will be performing the techniques described in the protocol. NOTE: If sharing
remaining tissues from a protocol with another PI, then that PI must be listed as a protocol associate.
Name
Department
III. Performance Sites
Building
Lab Room Number
Lab Phone
USA animal facility
determined by DCM
6-6239
IV. Project Funding
Funding Source:
Internal
External
Seeking Funding
No Funding
Type:
Federal
Name:
NIH
Do any personnel listed on this project, or members of their immediate families, have a significant
(personal) financial interest in the sponsoring company (awarded or seeking)?
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Yes
No
Grant Status:
Awarded
Seeking Funding
If awarded:
Start Date:
End Date:
PI on Grant:
Grant #:
V. Animal Use Information
N/A
Protocol Type:
Research
Breeding
Teaching
Tissue Harvest
Monitoring (DCM)
Experimental (Survival Surgery)
Field Studies
Observational Studies
Other:
Proposed Procedures:
Food/water restriction
Non-survival surgery
Survival surgery
Multiple survival surgeries
Multiple manipulations/procedures requiring anesthesia (other than survival surgeries)
Physical restraint
Use of paralytics
Biohazard use:
Chemical hazard use:
None of the above procedures are proposed
Will there be any acute procedures?
Acute procedures are non-survival surgeries which are performed on the animal while alive, but resulting
in the death of the animal.
Yes
No
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If yes: Describe the acute procedures:
Will an N95 mask be required?
Yes
No
N/A
Transported to off-campus locations?
Yes
No
Yes
No
If yes:
Destination:
Method of Transportation:
Animals kept outside vivarium for >12 hours?
If yes: Justify the need for extended periods outside the vivarium. Provide specific information
regarding the housing conditions in the off-site location (e.g., room temperature and control,
lighting, caging, food, water, etc.).
Domiciliary care required outside of Biologic Research Laboratory?
Yes
No
If yes: Justify the need for domiciliary care outside of the facility. Provide specific information
regarding the housing conditions in the off-site location (e.g., room temperature and control,
lighting, caging, food, water, etc.).
VI. Animal Segment Information
Species:
N/A
Source:
Mouse
Domestic
Foreign
Segment Description:
Mice on this Segment will be bred to generate pups with conditional deletion
Genetic Designation:
flox/flox; CD19 Cre/wt
Strain, Stock, Breed:
flox/flox; CD19 Cre/wt (on C57BL/6 background)
Number Requested:
66
USDA Categories:
Primary Euthanasia:
B
C
D
E
CO2
Secondary Euthanasia: cervical dislocation
Species:
Source:
Mouse
Segment Description:
Domestic
Foreign
Mice on this Segment will be bred to generate pups for conditional deletion
Genetic Designation: flox/flox; Cgamma1 Cre/wt
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Strain, Stock, Breed:
Number Requested:
flox/flox; Cgamma1 Cre/wt (on a C57BL/6 background
66
USDA Categories:
Primary Euthanasia:
B
C
D
E
CO2
Secondary Euthanasia: cervical dislocation
Species:
Source:
Mouse
Domestic
Foreign
Segment Description:
Mice on this Segment will be weanlings.
Genetic Designation:
genotypes produced by the matings of flox/flox; CD19 Cre/wt pairs
and flox/flox; Cgamma1 Cre/wt pairs
Strain, Stock, Breed:
weanlings as described above (on a C57BL/6 background)
Number Requested:
2000
USDA Categories:
Primary Euthanasia:
B
C
D
E
CO2
Secondary Euthanasia: cervical dislocation
VII. Biosafety Information
N/A
Infectious Agents:
Viruses
Bacteria
Fungi
Prions
Human cell lines
Blood/bodily fluids
Parasitic agents
Toxins
Recombinant DNA
None of the above infectious agents will be used
Select Agents or Toxins involved in this research?
Yes
No
If yes: List all Select Agents or Toxins:
Biosafety and Containment Levels:
BSL-1
BSL-2
BSL-3
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Any of the following additional considerations?
None
DNA introduced into experimental animals that fall under NIH Guidelines
Gene therapy in humans
[complete Human Gene Transfer Supplement form]
Other:
INSTRUCTIONS TO RESEARCHERS
Thank you for completing the University of South Alabama IACUC/IBC Application Form. Review the
contents of this form for accuracy and completeness before submitting this package to the appropriate
committee(s).
REGISTRATION IS COMPLETE!
You will need to finish constructing your submission package by adding some required documents if you
have not done so already. The cover sheet application you just filled out can be used for IBC and IACUC.
To complete your submission packet just print out these instructions and follow them.
For IACUC:
Download the IACUC application, which is located in the IACUC library of the Designer. Once you have
completed it and saved it to your computer, upload it back into your submission packet. This is done
by clicking the 'Add New Document' button in the Designer and uploading the file from your computer.
Also, upload a copy of the Vertebrate Animal Section (VAS) of your grant if the study is to be funded with
federal dollars.
For IBC:
Download the IBC application, which is located in the IBC library of the Designer. Once you have
completed it and saved it to your computer, upload it back into your submission packet. This is done by
clicking the 'Add New Document' button in the Designer and uploading the file from your computer.
For all submissions:
Your final submission will include the cover sheet (this completed Wizard), application, PI training
documentation (via the User Profile and linked to the package), ancillary documents, and the VAS (if
applicable).
All associate training will need to be uploaded on to their User Profile as initiated or updated. It is the
responsibility of the associates to create and maintain their own User Profile.
*Working with the IACUC, species-specific training, bio-safety/bio-security, and RCR training can be all
accessed through www.citiprogram.org.
**The NIH Bio-methodology of the Rat/Mouse CD can be obtained from the IACUC office. Verification
forms for the CD, Euthanex, Fish, etc can be found in the IACUC library.
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