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BIOMETHODOLOGY OF THE MOUSE Office of Laboratory Animal Care University of Tennessee, Knoxville.

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Presentation on theme: "BIOMETHODOLOGY OF THE MOUSE Office of Laboratory Animal Care University of Tennessee, Knoxville."— Presentation transcript:

1 BIOMETHODOLOGY OF THE MOUSE Office of Laboratory Animal Care University of Tennessee, Knoxville

2 General Behavior  Nocturnal  Non-aggressive towards humans  Social  Barbering is common  Males are more likely to fight if housed with non- littermates

3 Weight Gain Chart C57BL/6

4 Reproduction  Estrus Cycle 4–5 d  Gestation 19-21 d  Litter Size 10-12 pups  Eyes open 14 d  Weaning 21 d  Postpartum estrus  Reproductive life ~8 mo

5 Sexing  Newborns have a subtle difference in anogenital distance  At 9 days of age, nipples are evident in the female and absent in the male  Adults have a marked difference in anogenital distance

6 Housing  Adequate housing should provide the following:  Clean, dry and safe area with adequate ventilation, food and water  Visualization by personnel  Sufficient space to turn around and make normal postural movements

7 Enclosure Recommendations  Primary enclosure space recommendations per the Guide for the Care and Use of Laboratory Animals  UTK Mouse Cage Density Policy http://www.vet.utk.edu/olac/pdf/Mouse_Cage_Density_Policy.pdf

8 Primary Enclosure  Cage Bottom  Easy visualization  Solid bottom flooring  Bedding and enrichment  Microisolator top  Reduces spread of pathogens  Wire-bar lid  Water bottle  Feed

9 Environmental Conditions  Temperature 68 – 79 ⁰ F  Humidity 30 -70%  Ventilation 10 -15 air changes/hr.  Noise ≥85 db can cause  Stress  Metabolic changes  Reduced fertility Room Recommendations

10 Identification Ear PunchesEar Tags

11 Identification  Cage Card

12 Procedures  Institutional Animal Care and Use Committee (IACUC) protocol describes all procedures that can be performed  A procedure is defined as any activity carried out on a mouse such as:  Behavioral observation  Venipuncture  Surgery

13 Manual Restraint  Lift mouse by the base of the tail and place on wire bar lid  Push the mouse against the wire bar to prevent escape and advance the hand toward the head.  Grasp the scruff of the neck  Tuck tail between finger and palm

14 Manual Restraint Click to Watch Video

15 Mechanical Restraint Plastic Adjustable RestrainerDecapicone

16 Blood Collection  Survival Procedures  Orbital sinus  Tail vein prick  Facial vein  Non-Survival Procedures  Cardiac puncture  Cranial vena cava puncture  Axillary cut down

17 Blood Collection  Blood Collection Guidelines  Single blood draw  ≤1.0 ml per 100 grams of body weight  Multiple blood draws  Maximum of 1.5 ml per 100 grams of body weight within a 2 week period  Note: Average mouse body weight is ~20 grams but can be highly variable between stock/strain

18 Blood Collection  Orbital sinus  For collecting up to 0.2 ml of blood  Anesthetize mouse  Hold the head steady  Insert pipette in the medial canthus of the eye  Rotate the tube between thumb and finger  Keeping eyelids closed, apply direct pressure using gauze for hemostasis

19 Blood Collection  Tail Nick  For collecting up to 0.2 ml of blood  Warm tail to dilate blood vessel  Use proper restraint  Prep with 70% alcohol  Use #11 scalpel blade or needle to nick the lateral tail vein  Apply direct pressure for hemostasis

20 Blood Collection  Facial Vein  Hold the mouse securely  Locate the puncture site slightly caudal to the freckle  Apply petroleum based lubricant to the site  Use a lancet or 18 gauge needle to puncture the skin  Collect blood  Apply direct pressure for hemostasis

21 Blood Collection  Cardiac  For collecting up to 1.0 ml of blood  Anesthetize mouse  Insert ≤25 gauge needle under sternum at a 20 ⁰ angle  Aspirate slowly  Euthanize mouse

22 Blood Collection Click to Watch Video

23 Tissue Collection  Tail Biopsy  Limited to a maximum of 2 times  Maximum of 5 mm  Analgesia/Anesthesia is required for mice 21 days of age and older

24 Compound Administration  25 to 27 gauge needle Maximum Administration Volumes (in ml/kg)

25 Compound Administration  Subcutaneous (SC or SQ)  The sides of the mouse serve as good site for SQ injections  Insert needle underneath skin  Aspirate negative pressure  Inject compound and watch for SQ bleb  To reduce leakage from the injection site, pause before retracting the needle

26 Compound Administration Click to Watch Video

27 Compound Administration  Intraperitoneal (IP)  Locate lower right quadrant for injection site  Aspirate  If an unintended subcutaneous bleb occurs, reposition the needle

28 Compound Administration Click to Watch Video

29 Compound Administration  Intravenous (IV)  Lateral tail vein  Proper restraint  Heat source to dilate vessel  Apply direct pressure after injection

30 Compound Administration  Gastric Gavage (PO)  Use a bulb-tipped gastric gavage needle  Measure length of needle from mouth to last rib  DO NOT FORCE the needle down the esophagus  Inject solution  Observe mouse for signs of distress

31 Compound Administration Click to Watch Video

32 Anesthesia  When planning any procedure involving anesthesia and/or surgery, please consult one of the laboratory animal veterinarians in the Office of Laboratory Animal Care (OLAC) at 974-5634.  The veterinarian can provide guidance and detailed information in selecting the most appropriate anesthetic and analgesic protocol for your mice and procedure.

33 Aseptic Technique  Surgical Prep  After induction of anesthesia, clip hair from the surgical site  Prep skin with povidone iodine, chlorhexidine or other appropriate skin antiseptic  Scrub in a circular pattern, beginning in the center and spiraling outward  Follow povidone iodine scrub with a 70% alcohol prep  Repeat Twice  End procedure with a light coat of povidone iodine solution (not scrub) to the surgical site

34 Aseptic Technique  Place a sterile drape over the mouse  Anything that touches the surgical site must be sterile  Non-absorbable sutures/clips should be removed in 7-14 days

35 Surgical Monitoring  Prevent pain, hypoxia, and hypothermia  Monitor withdrawal reflex  Provide a source of external heat  Provide appropriate analgesics for post- operative pain management

36 Record of Medical Care

37

38 Euthanasia  Inhalant Anesthetic Overdose  Isoflurane  End the procedure with a thoracotomy or cervical dislocation  Carbon Dioxide  Place mouse in the chamber  Turn on CO 2 flow into the chamber  Once the mouse has stopped breathing, wait at least 1 minute before removing the mouse from the chamber  End the procedure with a thoracotomy or cervical dislocation  Substantially prolonged in neonates  Cervical Dislocation  Performed on anesthetized mice  Must be performed by skilled personnel

39 Occupational Health and Safety  PPE (Personal Protective Equipment)  Protects handler and mouse  May include: gloves, gowns, lab coats, shoe covers, hair bonnets, face masks

40 Prevention of Infectious Disease  Colony Health Surveillance  Sentinel mice detect for presence of infectious pathogens and parasites in the research colony  Mouse Antibody Production Test (MAP Test) and PCR  A test for cell lines and tumors for murine viruses  Included in every animal use protocol

41 Procurement of Mice  APPROVED vendors include Charles River, Jackson Labs, Harlan, NCI-Frederick, and Taconic  Mice that are shipped from a non-approved vendor source must be approved by OLAC before the mice are procured.  An animal requisition form must be submitted to the facility manager: http://www.vet.utk.edu/olac/pdf/animal_acquisition_form.pdf

42 Quarantine  Quarantine is required if receiving mice from an unapproved vendor  The minimum quarantine period is six (6) weeks  No experimental manipulations or breeding can be initiated during the quarantine period unless approval has been granted by an OLAC veterinarian

43 Health Concerns  General appearance  Lethargy  Aggressiveness  Hunched posture  Coat  Piloerection  Hair loss  Unkempt appearance

44 Health Concerns Abnormal massesSkin lesions

45 Health Concerns  Teeth NormalMalocclusion

46 Body Condition Scoring Ullman-Cullere, et al. 1999

47 Tumor Production  Four criteria for euthanasia:  Tumor size of 1.5 cm in diameter or tumor ulceration  Body condition score of 1  Tumor interferes with function of vital organs  Tumor significantly interferes with locomotion

48 Reporting Signs of Pain or Distress  For Mice That Require Veterinary Care  Complete the red “Sick Animal” cage card  Attach card to cage  Notify facility manager or  Print Clinical Case Request Form http://www.vet.utk.edu/olac/pdf/CLINICAL_CASE_REQUEST.pdf http://www.vet.utk.edu/olac/pdf/CLINICAL_CASE_REQUEST.pdf  Fax form to the OLAC office 974-5649  Assessment of the mouse’s condition and treatments will be recorded on the “Sick Animal” card

49

50 10 Question Quiz

51 References  GUIDE FOR CARE AND USE OF LABORATORY ANIMALS. National Academy Press, 2011.  Lawson, PT. 2005. ASSISTANT LABORATORY ANIMAL TECHNICIAN (ALAT) TRAINING MANUAL. The American Association for Laboratory Animal Science, Memphis, TN.  Suckow M, Danneman P, Brayton C. 2001. THE LABORATORY MOUSE. CRC Press, Boca Raton, FL.  Ullman-Cullere, M.H. and Foltz, C.J. 1999. Body Condition Scoring: A Rapid and Accurate Method for Assessing Health Status in Mice. Lab Anim Sci 49(3) 319-323.  Presentation: Chris Carter, OLAC UTK


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