Guidelines for Maintenance of Tumors and Hybridomas in Rodents

Cell lines should be tested for presence of murine viruses prior to introduction into the animal colony.  (Contact the VSC for information about testing services, 723-3876).  If implanting human tissues is a component of the approved protocol, issues regarding occupational health may need to be addressed, and can be done by contacting EH&S (http://www.stanford.edu/dept/EHS/prod/).

Animals inoculated with cells producing solid tumors must be observed with sufficient frequency (once a day for the first week following tumor induction, then a minimum of three times per week until tumor development becomes evident, and then daily thereafter until they are to be euthanized – including weekends and holidays) to ensure that they are euthanized before:  tumors achieve any degree of ulceration (including pinpoint lesions); reach sufficient size to interfere with normal activity; grow to be 1 cm3 in dimension or equal 10% of the animal's pre-inoculation body weight.  If tumor growth is rapid, increasing the frequency of monitoring to twice daily should be considered.  Transgenic and knockout animals that are predisposed to develop cancer may have unpredictable tumor growth.  Continuous monitoring to observe tumor growth and establish animal care parameters may be needed.

Projects that require advanced cancer (tumors greater than 1 cm3 in size or any degree of ulceration) must submit a detailed justification for an exception to these guidelines in the protocol to be reviewed by the A-PLAC.  The justification must specify the scientific reasons for this exception, why an earlier endpoint cannot be used, special husbandry needs that might occur (e.g,. single cages for animals with ulcerated tumors), provide a detailed plan for the treatment of health issues that may arise as a result of the advanced cancer (e.g., antibiotics, fluid therapy, supplemental nutritional support, topical treatments, etc.) and define endpoints/criteria for euthanasia.  If your protocol requires advanced cancer, please consult with the veterinary staff (723-3876) to design a supportive regimen to include in your protocol.  Contact the A-PLAC office (723-4550) for instructions on submitting a detailed justification for an exception in your protocol.  Please note that the exception must be approved by the A-PLAC before animals are allowed to develop advanced cancer.

Following tumor induction, animals must be observed for pain, distress, and abnormal behavior and physiology.  The overall well-being of the animal takes priority over precise tumor measurements in decisions regarding euthanasia or other interventions.  Clinical signs that should be monitored closely include (but are not limited to) rapid weight loss, loss of body condition, anorexia, the absence of normal urine and feces, abnormal respiratory rate/patterns, listlessness, piloerection (ruffled haircoat), abdominal distention, ambulatory difficulties, hemorrhage, infection, self-mutilation/cannibalism, abnormal aggression towards cage mates, vocalization, paralysis, and/or seizures.

Tumors induced in body cavities (cranium, orbit, abdomen, or thorax) may have additional limitations as to the maximum acceptable size.  These animals must be monitored very closely for any severe impairment in physiological or neurological function and be euthanized as soon as such signs become apparent.

After inoculation with an ascites-producing tumor cell line, animals must be observed at least three times per week for the first week and daily thereafter to monitor the degree of abdominal distention and signs of illness.  Ascites fluid should be removed by peritoneal tap before abdominal distention is great enough to cause discomfort or interference with normal activity.  Animals should be euthanized if they become moribund (i.e., huddling, hunched posture, increased respiratory rate and/or effort, lethargy, difficulty with normal ambulation, or ruffled coat).  Removal of peritoneal fluid with an 18-gauge or smaller needle may be done without anesthesia by skilled personnel. New personnel or students must be trained and develop proficiency using anesthetized animals prior to performing the procedure on awake animals. Training is available from VSC staff.  Ascites collection is limited to a maximum of three taps, with the animal being euthanized just prior to the third tap.

Written documentation of dates and times of all observations (including recordings of tumor size progression), status of animals and any procedures performed (i.e. euthanasia, peritoneal tap to drain ascites, supportive treatment, etc.) must be kept for review by VSC, A-PLAC, or regulatory agencies.  Contact the VSC for training in proper record keeping or to obtain sample monitoring forms.