High-frequency Ultrasonography Based Early Pregnancy Characterization: A Technique to Study Embryo Implantation and Pregnancy Progression in Pregnant Murine Model

Begin with preparing the ultrasound machine. Turn on the transducer and then attach the 3D motor stage to the transducer. Next, attach the probe to the transducer clamp. Now, position the mouse supine on the monitoring platform. Provide it anesthetic via nose cone and gently tape all of its paws to the heart rate monitoring pads on the platform.

Next, rub about 1 or 2 milliliters of ultrasound transmission gel onto the abdomen. Then, position the ultrasound probe on the lower abdomen using the manual motor stage. Using the ultrasound, locate the bladder, which should appear as a fluid-filled dark circle just cephalad to the vaginal opening.

Once the bladder is located, move the probe very slowly cephalad to visualize the pregnant uterus, which should appear as a cylindrical shape with round areas at pregnancy sites. This can be described as looking like pearls on a chain. Now, proceed with 2D imaging.

Once the pregnant uterus has been identified, begin at the pregnancy site closest to the bladder and slowly and sequentially move cephalad to determine location of all the pregnancy sites. Save the image. If the kidney, spleen, or liver can be seen, then the scan is too far cephalad. Reposition the probe closer to the bladder and save the image.

Next, image the contralateral uterine horn in the same manner. Freeze the image when the ultrasound frame is at the center of the pregnancy or implantation site and save the image for analysis. Now or later, use the saved images to measure implantation positions using the hyperechoic decidualization reaction as a marker.

Manually click on the "measure" tool and click on the location of an implantation site. Then, drag the cursor to the next implantation site and click again to draw a line. The software then reports the length of the line. After all the implantation sites are measured, use the same technique and proceed to measure the gestational sac size and the fetal pole size.