The collection of blood samples is a common practice in murine research, used for diagnostic testing, drug toxicity screening, and genetic analysis. Because mice are small, their physiological systems are highly sensitive to changes in blood volume. Strict adherence to ethical guidelines for blood collection is necessary to ensure humane animal care and prevent data from being skewed by stress or blood loss. Establishing the maximum volume of blood that can be safely drawn maintains both animal welfare and research integrity.
Defining Safe Blood Collection Limits
Regulatory guidelines, often set by Institutional Animal Care and Use Committees (IACUCs), establish maximum limits for blood collection to protect the mouse’s health. The universally accepted standard for a single, non-terminal blood draw is 10% of the animal’s total circulating blood volume (TBV). This 10% limit is considered safe for collection every two to four weeks without requiring fluid replacement.
If fluid replacement, such as a warmed isotonic solution, is administered immediately after the draw, the safe single-collection limit can be extended up to 15% of the TBV. This 15% is the absolute maximum for a single, survival procedure and is typically reserved for healthy animals.
For repeated or chronic blood sampling, the volume must be significantly reduced to allow the mouse time to regenerate blood components. The maximum volume that can be safely collected over a week is generally restricted to 7.5% of the total blood volume. If collections are performed daily, the volume must be less than 1.0% of the TBV every 24 hours. These lower limits account for the slow rate at which mice replace lost red blood cells and plasma components.
Calculating Total Blood Volume and Safe Draw Amount
Determining the safe blood collection volume requires first estimating the mouse’s total blood volume (TBV). The circulating blood volume for a healthy mouse is generally estimated to be between 70 and 80 milliliters of blood per kilogram of body weight (ml/kg).
To calculate the TBV, the mouse’s weight is used with this ratio. For example, a common laboratory mouse weighing 25 grams has a TBV of approximately 1.8 to 2.0 milliliters. Once the TBV is established, the safe collection amount is determined by applying the regulatory percentage limit.
Using the standard 10% limit for a single, survival collection, the maximum safe volume for a 25-gram mouse is about 0.2 milliliters. This small volume necessitates the use of microsampling techniques to minimize the impact on the animal. Researchers must calculate the specific volume for each animal based on its current body weight and the frequency of collection.
Physiological Consequences of Excessive Blood Loss
Exceeding defined blood collection limits can lead to two main physiological crises: hypovolemic shock and severe anemia. Hypovolemic shock is the immediate concern, resulting from a rapid loss of circulating fluid volume that causes a sudden drop in blood pressure. If this fluid loss compromises blood flow to the heart and brain, it can quickly become fatal.
Removing less than 15% of the total blood volume is usually managed by the mouse’s body to maintain blood pressure. However, removing 15% or more can lead to hypovolemic shock, with symptoms including a fast, weak pulse, pale mucous membranes, and lethargy. The loss of more than 30% to 40% of the TBV is expected to cause death in at least half of the animals.
Anemia, the loss of oxygen-carrying capacity, is the longer-term consequence of excessive or too-frequent collection. While plasma volume restores quickly, the body requires about two weeks to regenerate lost red blood cells. If blood is collected before the red blood cell count recovers, the animal’s health will decline, potentially causing pale extremities and decreased activity.
Acute vs. Survival Blood Collection Schedules
The research timeline determines whether an acute or survival blood collection schedule is used, which changes the maximum allowable blood volume.
Survival (Chronic) Collection
A survival, or chronic, schedule is used when the mouse must remain healthy for an extended study period. This requires adhering to the strict limits of 7.5% to 15% of the TBV, depending on the frequency of the draws (daily, weekly, or bi-weekly).
Acute (Terminal) Collection
An acute collection is typically performed as a terminal procedure, meaning the animal is euthanized immediately after the blood is drawn. During this procedure, the maximum volume is substantially higher, often involving exsanguination. Exsanguination involves collecting the maximum possible volume, which can be approximately 50% to 75% of the total blood volume, or roughly 4% to 5% of the mouse’s body weight. Terminal collection must be performed under deep anesthesia to minimize distress.