Protocols


Screen (pdf version)
ScreenBone Screen
Common NameDEXA
Posted On01/04/2021 6:16 PM
AuthorJonathan Rios
Background

Numerous genes regulate various aspects of skeletal development, including skeletal patterning and integrity. Many genes regulating these processes in humans have been modeled in mice and shown to largely phenocopy major skeletal defects observed in human patients. The goal of this screen is to identify novel genes regulating skeletal patterning and integrity. This is achieved using in vivo live animal imaging performed in a high-throughput manner.

Reagents and Solutions

Anesthesia: Isoflurane delivered through precision vaporizer

 

Imaging: Faxitron UltraFocusDXA

Method

Anesthesia. Up to two mice from the same cage are anesthetized in a chamber with isoflurane (1.5-2%) delivered through a precision vaporizer. Once mice are sufficiently anesthetized, both are transferred to the UltraFocus instrument and anesthesia continued via nose cone.

 

Imaging. Once mice are successfully transferred to the UltraFocus, mice are positioned in opposite orientation to allow sufficient space for imaging both mice simultaneously. Mice are positioned in the prone position with the head straight and body balanced as able. Proper positioning is required to avoid non-physiologic bending of the spine. Limbs are splayed outward to allow imaging of the femur and tibia. Once positioned, DXA scans are acquired (approx. 3 minutes). Following DXA scans, a single X-ray is acquired (approx. 10 seconds). Mice are transitioned from the prone position to their side, and lateral X-rays are acquired (approx. 10 seconds). Once completed, mice are transferred out of the instrument, anesthesia discontinued, and mice are monitored until fully recovered.

 

DEXA Analysis. Analysis of DEXA images is performed using the Faxitron Vision software. Using the “Bone Map”, arbitrary region-of-interest analyses are performed to measure bone mineral density (mg/cm^2) and bone mineral content (grams) of both the left and right tibia and femur. For statistical analysis, bone mineral densities and bone mineral content measures of the tibia and femur are averaged. Following, quantitative measures of fat mass (grams) and lean mass (grams) are measured using the arbitrary region-of-interest analysis feature for the entire body with the head excluded.

 

X-ray Analysis. Analysis of X-ray images is performed using the Faxitron Vision software. Subjective measures of skeletal patterning include observations for limb deformities, kinked or short tails, scoliosis, kyphosis, and vertebral or limb abnormalities. Quantitative measure (in millimeters) of the left and right limbs include tibia length, femur length, and length from the distal femur to the fibula. Each quantitative phenotype is averaged for statistical analysis.