Do Not Fill in Green Boxes

Work protocol

Do not fill in green boxes

Stipulations CCD:
Stipulations IvDUtrecht:

Report changes to the IvDU ()
Report unforeseen dead animals and animals with unexpected discomfort to the IvD Utrecht () and the animal facility
The guideline to the work protocol can be found on

Seen by animal facility

/ Stamp IvD Utrecht /

Conclusion of workprotocol

Date: / Date:
Initials:

Approved IvD Utrecht

Date: / When the work protocol is concluded, send a welfare evaluation to

Does this concern an animal experiment

/ No Yes /

Type experiment (appendix CCD project):

/ Choosen.a.1234567891011121314151617181920Several, namely:

Title of the work protocol

Projectnumber (CCD):

/ AVD /

Principal investigatorof the project (name + email)

Project title:

End date CCD license:

Workprotocol number:

Optional: experiment code location

Animal facility:

/ ChooseGDLStratenumKruyt (Biology)VET-Companion AnimalsVET-Equine sciencesVET-Farm AnimalsVET-Tolakker /

Optional: GDL section(s)

/ ChooseBlueGreenInfectionLargeOrangePinkPurpleRedSPFWhiteYellowN.A. / If needed: ChooseBlueGreenInfectionLargeOrangePinkPurpleRedSPFWhiteYellowN.A.

Planned start date:

Planned enddate:

Will animals be used in experiments and registered as laboratory animals before weaning?

/ No Yes

Are biological agents (GMO yes/no) and/or biologicals and/or biological test substances and/or ionizing radiation used?

/ No Yes(If ‘yes’, please fill out the ‘hazardous substances overview’ p. 8)
Parties involved: / Name / Initials / Mobile phone / Email / Qualification / Faculty/Institute/ Department

Study Director (SD)

/ Art. 9

Alternate Study Director (ASD)

/ ChooseArt. 9Art. 13f2

Co-worker

/ ChooseArt. 9Art. 13f2

Co-worker

/ ChooseArt. 9Art. 13f2

Co-worker

/ ChooseArt. 9Art. 13f2

Co-worker

/ ChooseArt. 9Art. 13f2
Departmental Lab Animal Co-ordinator

Contact personAnimal facility

(Will be filled in by the facility)

Contact personAnimal section

(Will be filled in by the facility)

After conclusion of the work protocol, please send original study dossier and raw data to the address below(please fill in name and full postal address):

/ Approval of the principal investigator of the project
Name:
Date:

Version 6.1 - 2018Page1 of10

Work protocol

1.Description of the experiment(Guideline experimental design and statistics)

a.Abstract: Please give a short summary (5 sentences)of the experiment with the purpose of the study in simple English.
Also give the short summary in simple Dutch
b.What is the specific research question in this work protocol and how does this fit in the research strategy of the project? (refer to project and type of experiment concerned).
c.Experimental design: sketch the outline of the experimental design:

What is/are the primary and secondary read-out parameter(s)
Explain (in general terms) the experimental design
Which(and how many) experimental groups (treated/control groups) are there
What is the time schedule of the experiment
What is the number of animals per group (n)
(If desired, you can outline design and course of the experiment by means of a flow chart)

d.Statistical substantiation of the total number of animals – statistical methods.

On basis of which experimental parameter(s) is the sample size defined?
Which statistical analysis will be performed?
Describe how the group size (n) is calculated.

e.Is this a randomized experiment? If so, how? If not, why not?
f.Is this a blinded experiment with respect to animal treatment and analysis of the results? If so, how? If not, why not?

Version 6.1 - 2018Page1 of10

Work protocol

2.Animals

a.Species to be used1ChooseCatCattleDogDonkeyDoveFerretGoatGuinea pigHorseMouseParrotPigPoultryRabbitRatSheepToadZebra finchZebra FishOther (please clarify)

Geno- type / Breed/strain name (Please indicate the official strain name and/or supplier reference number) / Supplier / GDL strain name / Number of animals / Sex / GA2 / Immune competent / Origin code / Age / Weight
A / ChooseMFBothNo preference / No
Yes / Chooseyesnounknown / Choose12345
B / ChooseMFBothNo preference / No
Yes / Chooseyesnounknown / Choose12345
C / ChooseMFBothNo preference / No
Yes / Chooseyesnounknown / Choose12345
1Please specify animal species ‘other’:
2If genetically modified, please give corresponding IG-number: .State the risk assessment article according to Appendix 5 (GMO legislation): .

Version 6.1 - 2018Page1 of10

Work protocol

b.Justify the choice of species, strain, sex and (if applicable), the reason for the use of transgenic/k.o. animals
c.Re-use:

If re-use is possible, origin of the animals: Were the animals used in previous procedures? No Yes

If so, what is theWP number? Level of discomfort in previous procedures: ChooseMildModerateSevere

d.Surplusanimals:

Surplus animalschoosecancannotbe used because:

Version 6.1 - 2018Page1 of10

Work protocol

3. Housing

Experimental group / Group housing1 / Bedding3 / Enrichment3 / Food2,3,4 / Water2,3,4 / Housing types3 / Restriction level
ChooseYesNo / ChooseStandard animal facilitySandSawdust/wood shavingsStrawNo beddingOther (please clarify) / ChooseBird, toyBird, water bathCat, toyCattle, brushDog, toyFerret, play tubeGoat, brushGuinea pig, shelterHorse, motionMouse, paper shelter + tissuesMouse, plastic shelter + tissueMouse, tissuePig, biting stickPig, chainPoultry, grainPoultry, perchPoultry, perch + grainPoultry, scratch padRabbit, shelterRat, gnawing stickRat, paper towelRat, plastic shelterRat, plastic shelter + paper towelOther (please clarify) / ChooseStandard animal facility ad libitumGDL treated ad libitumOther (please clarify) / ChooseStandard animal facilityOther (please clarify) / ChooseAviaryBox stableFerret cageFilter top cageGrazingInsulatorIsocageIVCKennelLoose housing for diary cattleMultifunctional stablesOpen cageOpen cagePaddockStanchionOther (please clarify) / ChooseNo restrictionD-IDM-IDM-IIDM-III
ChooseYesNo / ChooseStandard animal facilitySandSawdust/wood shavingsStrawNo beddingOther (please clarify) / ChooseBird, toyBird, water bathCat, toyCattle, brushDog, toyFerret, play tubeGoat, brushGuinea pig, shelterHorse, motionMouse, paper shelter + tissuesMouse, plastic shelter + tissueMouse, tissuePig, biting stickPig, chainPoultry, grainPoultry, perchPoultry, perch + grainPoultry, scratch padRabbit, shelterRat, gnawing stickRat, paper towelRat, plastic shelterRat, plastic shelter + paper towelOther (please clarify) / ChooseStandard animal facility ad libitumGDL treated ad libitumOther (please clarify) / ChooseStandard animal facilityOther (please clarify) / ChooseAviaryBox stableFerret cageFilter top cageGrazingInsulatorIsocageIVCKennelLoose housing for diary cattleMultifunctional stablesOpen cageOpen cagePaddockStanchionOther (please clarify) / ChooseNo restrictionD-IDM-IDM-IIDM-III
ChooseYesNo / ChooseStandard animal facilitySandSawdust/wood shavingsStrawNo beddingOther (please clarify) / ChooseBird, toyBird, water bathCat, toyCattle, brushDog, toyFerret, play tubeGoat, brushGuinea pig, shelterHorse, motionMouse, paper shelter + tissuesMouse, plastic shelter + tissueMouse, tissuePig, biting stickPig, chainPoultry, grainPoultry, perchPoultry, perch + grainPoultry, scratch padRabbit, shelterRat, gnawing stickRat, paper towelRat, plastic shelterRat, plastic shelter + paper towelOther (please clarify) / ChooseStandard animal facility ad libitumGDL treated ad libitumOther (please clarify) / ChooseStandard animal facilityOther (please clarify) / ChooseAviaryBox stableFerret cageFilter top cageGrazingInsulatorIsocageIVCKennelLoose housing for diary cattleMultifunctional stablesOpen cageOpen cagePaddockStanchionOther (please clarify) / ChooseNo restrictionD-IDM-IDM-IIDM-III
ChooseYesNo / ChooseStandard animal facilitySandSawdust/wood shavingsStrawNo beddingOther (please clarify) / ChooseBird, toyBird, water bathCat, toyCattle, brushDog, toyFerret, play tubeGoat, brushGuinea pig, shelterHorse, motionMouse, paper shelter + tissuesMouse, plastic shelter + tissueMouse, tissuePig, biting stickPig, chainPoultry, grainPoultry, perchPoultry, perch + grainPoultry, scratch padRabbit, shelterRat, gnawing stickRat, paper towelRat, plastic shelterRat, plastic shelter + paper towelOther (please clarify) / ChooseStandard animal facility ad libitumGDL treated ad libitumOther (please clarify) / ChooseStandard animal facilityOther (please clarify) / ChooseAviaryBox stableFerret cageFilter top cageGrazingInsulatorIsocageIVCKennelLoose housing for diary cattleMultifunctional stablesOpen cageOpen cagePaddockStanchionOther (please clarify) / ChooseNo restrictionD-IDM-IDM-IIDM-III

1If ‘no’ please explain

2Clarify food/water restriction if applicable

3If ‘other’ please clarify and/ordescribe additional requirements of housing conditions

4In case of nonstandard food and/or water: Please fill out the form ‘hazardous substances overview’

4.Antibiotics, anaesthesiaand analgesia

a.Are antibiotics and/or heterologous anti-microbial prophylactics and/or therapeutics administered?No YesIf so, please fill out the table below.

Experimental group / Substance (including brand name) / Dose / Route of administration / Frequency / Duration treatment

b.Will anaesthesia be administered? No YesIf so, please fill out the table below.

Anaesthesia / Substance (including brand name) / Route of administration / Dose
Premedication agent
Introductory agent
Maintenance agent
Insufflation with intubation: No Yeso
Optional: explanation anaesthesia

c.Will analgesia be administered? No Yes1If so, please fill out the table below.

Analgesia / Substance (including brand name) / Route of administration / Dose / Interval / Duration
Pre-surgical agent
Agent during surgery
Post-surgical agent
Analgesia NOT related to any performed surgery
1If applicable, please explain why no analgesia is administered.

5.Describe consecutive procedures

a. Acclimatization: 1 week other (please define):

Table5.b. Schedule of experimentalprocedures

Line / Relative day in experiment / Experimental group / Description of the procedure / Duration proce-dure / Description of discomfort during and as a result of the procedure / Duration discomfort / Estimated level of discomfort / Who (see initials on page 1)
ChooseNon-recoveryMildModerateSevereN.A.
ChooseNon-recoveryMildModerateSevereN.A.
ChooseNon-recoveryMildModerateSevereN.A.
ChooseNon-recoveryMildModerateSevereN.A.
ChooseNon-recoveryMildModerateSevereN.A.
ChooseNon-recoveryMildModerateSevereN.A.
ChooseNon-recoveryMildModerateSevereN.A.
ChooseNon-recoveryMildModerateSevereN.A.
ChooseNon-recoveryMildModerateSevereN.A.
ChooseNon-recoveryMildModerateSevereN.A.
ChooseNon-recoveryMildModerateSevereN.A.
ChooseNon-recoveryMildModerateSevereN.A.
ChooseNon-recoveryMildModerateSevereN.A.
ChooseNon-recoveryMildModerateSevereN.A.

c. Duration entire experiment (excluding acclimatization):

6.Welfare check and humane end point

a.Regularwelfare check

Starting point is that the animal and the accommodation are checked daily at all times (from the moment the animals enter the facility). This is a routine check for calamities.

At least once per 7 days a thorough check of the animal’s welfare is performed on the basis of behaviour, posture, gait/mobility, food intake/weight loss, grooming/auto mutilation and other striking clinical symptoms.Findings of this welfare check are noted in the Welfare Logbook.

b.Additional welfare checks within the scope of possible experiment-specific clinical symptoms

Depending on the experiment, so-called experiment-specific clinical symptoms can occur.

Is this the case? No Yes

If so, please indicate in the table below:

At what time-points the animals need to be assessed. Describe the experiment-specific clinical symptoms and when in the experiment these are expected. If possible, please quantify the clinical symptom/parameter, e.g. volume of a tumour, circumference of a swollen knee joint, etc.
Please also describe the estimated level of discomfort (mild, moderate, severe).

Table 6.b.: Points (relative day in experiment) of observation and the experiment-specific clinical symptoms that need to be assessed.

Line / Day/period in experiment / Observation frequency / Experimental group / Assessment of experiment-specific clinical symptoms / Estimated level of discomfort / Who (initials see p1)
1 / ChooseNon-recoveryMildModerateSevere
2 / ChooseNon-recoveryMildModerateSevere
3 / ChooseNon-recoveryMildModerateSevere
4 / ChooseNon-recoveryMildModerateSevere
5 / ChooseNon-recoveryMildModerateSevere
6 / ChooseNon-recoveryMildModerateSevere
7 / ChooseNon-recoveryMildModerateSevere
8 / ChooseNon-recoveryMildModerateSevere
9 / ChooseNon-recoveryMildModerateSevere
10 / ChooseNon-recoveryMildModerateSevere

c.Humane endpoint / Premature withdrawal of animal(s)

When a humane end point (HEP) is applied, the experiment is prematurely terminated for an individual animal or a group(i.e. before the planned end of the experiment). The goal of the HEP is to prevent avoidable discomfort. There can be three reasons to apply a HEP. You are requested to indicate for I and II which criteria you use.

I.The discomfort of an individual animal that is inherent to the experiment exceeds the maximum discomfort described in the project.Describe for which clinical symptoms (specify degree and seriousness) and on basis of what criteria (opional) a HEP is applied.

Describe the HEP also in Dutch.

II.The scientific goal is reached.Describe –onanimal level–when the scientific end point has been reached and what the criteria are to withdraw an animal from the experiment.

III.The scientific goal can no longer be reached.For example: due todropoutssignificant results can no longer be reached; the readout is affected by external factors as a result of which the animal is no longer suitable to produce reliable results.If this is the case, make a note in the Welfare Logbook and report to the IvD Utrecht!

7.Estimated maximum discomfort per experimental group

Experimental group / Estimated maximum discomfort / Description of discomfort (on the basis of type and number of procedures – see table 5.b- and clinical symptoms- see table 6.b))
ChooseNoneNon-recoveryMildModerateSevere
ChooseNoneNon-recoveryMildModerateSevere
ChooseNoneNon-recoveryMildModerateSevere
ChooseNoneNon-recoveryMildModerateSevere

8.Ill and dead animals during the experiment

What to do with unexpected ill animals?

What to do with unexpected dead animals?

1. Ill/dead animals must always be reported to a staff member of the animal facility andIvD Utrecht.

2. Investigation into a possible cause of illness/death is highly recommended.

9.Destiny of animals at the end of the experiment

Experimental group / Destiny / If euthanasia: method
ChooseEuthanasiaRe-useAdoption / ChooseMouse/Rat/Hamster CO2/O2 asphyxiationMouse/Rat/Hamster decapitationMouse/Rat/Bird cervical dislocationPoultry, electrocutionAll animal species overdose euthesate
ChooseEuthanasiaRe-useAdoption / ChooseMouse/Rat/Hamster CO2/O2 asphyxiationMouse/Rat/Hamster decapitationMouse/Rat/Bird cervical dislocationPoultry, electrocutionAll animal species overdose euthesate
ChooseEuthanasiaRe-useAdoption / ChooseMouse/Rat/Hamster CO2/O2 asphyxiationMouse/Rat/Hamster decapitationMouse/Rat/Bird cervical dislocationPoultry, electrocutionAll animal species overdose euthesate
ChooseEuthanasiaRe-useAdoption / ChooseMouse/Rat/Hamster CO2/O2 asphyxiationMouse/Rat/Hamster decapitationMouse/Rat/Bird cervical dislocationPoultry, electrocutionAll animal species overdose euthesate

Clarificationother method of euthanasia

Clarification when re-use or adoption is possible

Date of availability for re-use or adoption

Version 6.1 - 2018Page1 of10

Work protocol –hazardous substancesoverview

Hazardous substances

Are biological agents (GMO yes/no) and/or biological substances and/or test substances and/or non- feed and/or ionizing radiation being used?

Biological agents, both GMO and non-GMO (micro-organisms, transduced cells/cell lines) No Yes
If ‘yes’, please fill in appendix RIE Micro-organismen. Describe side effects causing discomfort to the animals at E.

Experimental group

Name agent

Concentration, volume, administering route and location

PRIS number agent

(if known)

IG-number

Classification for IG license after 01-03-2015

Other biological products (for instance organs, tissue, cells, cytokines, antibodies)No Yes
If ‘yes’, please fill in appendixRIE Biologische Producten.Describe side effects causing discomfort to the animals at E.

Experimental group

Name product

Concentration, volume, administering route and location

PRIS number agent

(if known)

Frequency

Duration of treatment

Test-/auxiliary substances (toxic substances, cytostatics, pharmaceutical substances,etc.) No Yes
If ‘yes’, please fill in appendixRIE Teststoffen.Describe side effects causing discomfort to the animals at E.

Experimental group

Name substance

Concentration, volume, administering route and location

PRIS number agent

(if known)

Frequency

Duration of treatment

Non-standard feed (includingadditives/supplements) No Yes
If ‘yes’, please fill in appendix RIE Voer.Describe side effects causing discomfort to the animals at E.

Experimental group

Name of feed

Feed additive including concentration (mg/kg)

PRIS number agent

(if known)

Durationof administration of non-standard feed

E.Describe possible side effectsof substances mentioned in A. to D., including level of discomfort.

F.Ionizing Radiation

Substances labelled radioactive No Yes
If ‘yes’, please fill in appendixRIE Ioniserende Straling

Experimental group

Name substance

/ Concentration and volume, / Administration route and location /

SIT number[1]

Frequency

Duration of treatment

Body irradiation No Yes

Experimental group

Origin

Dose/animal

SIT number1

C-arm imaging system (diagnostic X-ray) No Yes

Nameof authorizedemployee

Education level(3, 4A, 5A)

SIT number1

Micro-CT scanner No Yes

Name ofemployee[2]

SIT number1

Version 6.1 - 2018Page1 of10

Work protocol – Registration NVWA

Table Code numbers according to NVWA-registration. Please provisionally fill in the table below! Attention: use new codes (since 01-01-2016), see next page.

The Study Director is required to send the completed Welfare Evaluation form (see to the IvD Utrecht () and to the lab animal co-ordinator of their own department within two weeks of conclusion of the work protocol and give the actual encountered discomfort of the animals in column 12. The data for legally required annual registration are retrieved from this table.

OWE number (department; see right column on the next page):

Experimental group / Animal particulars / Species / Origin of animals (place of birth) or reuse / Number of animals / Purpose of experiment / Specification of ‘other purpose’ (column 5 ) / Legal stipulations / Toxicity tests and other statutory safety research / Special techniques / Anaesthesia / Analgesia, post-operative of or unrelated to surgery / Degree of discomfort / Condition of the animal at the end of the experiment / Number CCD project application / Specification other species (column 2) / Specification other birthplace NHP (column 3) / Specification other tox research (column 8) / Description
1 / 2 / 3 / 4 / 5 / 6 / 7 / 8 / 9 / 10 / 11 / 12 / 13 / 14: / 15: / 16 / 17
AVD
AVD
AVD
AVD
AVD
AVD
AVD
AVD

Version 6.1 — 2018Page1

COLUMN 1 / COLUMN 16
Code / Type of animal / Code / Purpose (continued) / Specification of ‘other birthplace NHP’
1 / Animal other than B, C or D (not genetically altered, no NHP) / Regulated production
2 / Genetically altered without harmful phenotype / 50 / Blood based products / COLUMN 17
3 / Genetically altered with harmful phenotype / 51 / Monoclonal antibodies / Specification of ‘other toxicity tests’
5 / Animal taken from the wild other than NPH / 59 / Other*
6 / NHP FO
7 / NHP F1 / Quality control
8 / NHP F2 or greater / 60 / Batch safety testing
9 / NHP self-sustaining colony / 61 / Pyrogenicity testing
62 / Batch potency testing
COLUMN 2 / 69 / Other quality controls
Code / Species
1 / Mice (Mus Musculus) / Legislative required tox research
2 / Rats (Rattus Norvegicus) / 70 / Acute/subacute: LD50, LC50
4 / Guinea-pigs (Cavia porcellus) / 71 / Acute/subacute: other lethal methods
5 / Mongolian Gerbil (Meriones uneuiculatus) / 72 / Acute/subacute: non-lethal methods
6 / Hamster (Syrian) (Mesocricetus auratus) / 73 / Skin irritation / corrosion / OWE / NAME
7 / Hamster (Chinese) (Cricetulus griseus) / 74 / Skin sensitization / 10863 / BIO/Afd. Celbiologie
9 / Other Rodents (other Rodentia)* / 75 / Eye irritation / corrosion / 10857 / BIO/Leerstoelgroep Ontwikkelingsbiologie, afd. Aquaria en
11 / Rabbits (Oryctolagus cuniculus) / 76 / Repeated dose ≤ 28 days / Volières
21 / Dogs (Canis familiaris ) / 77 / Repeated dose 29-90 days / 10856 / BIO/Leerstoelgroepen Animale Ecologie, Ecologie &
22 / Cats (Felis catus) / 78 / Repeated dose > 90 days / Biodiversiteit, en Onderwijsinstituut Biologie
23 / Ferrets (Mustela putorius furo) / 79 / Carcinogenicity / 10805 / DGK/B&C
29 / Other carnivores (other Carnivora)* / 80 / Genotoxicity / 10813 / DGK/DGG
31 / Prosimians (Prosimia) / 81 / Reproductive toxicity / 10803 / DGK/DGL
32 / Marmoset and tamarins (e.g. Callithrix jacchus) / 82 / Developmental toxicity / 10809 / DGK/DGL/Afd. Voeding
33 / Rhesus monkey (Macaca mulatta) / 83 / Neurotoxicity / 10801 / DGK/DGP
34 / Cynomolgus monkey (Macaca fascicularis) / 84 / Kinetics (pharmacokinetics, toxicokinetics, residue depletion) / 10818 / DGK/DWM
35 / Vervets Chlorocebus spp. (usually either pygerythrus or sabaeus) / 85 / Pharmaco-dynamics (including safety pharmacology) / 10810 / DGK/I&I/Afd. Klinische Infectiologie
36 / Baboons (Papio spp.) / 86 / Phototoxicity / 10811 / DGK/I&I/Afd. Moleculaire Infectiologie/Sectie Immunologie
37 / Squirrel monkey (e.g.Saimiri sciureus) / 87 / Ecotoxicity/Acute toxicity / 10823 / DGK/I&I/Afd. Moleculaire Infectiologie/Sectie Virologie