To begin, place three to six dechorionated embryos in a Petri dish containing Danios medium supplemented with 0.02%tricaine. Once the embryos are anesthetized, transfer them to a 35 by 15 millimeter glass bottom dish with minimal medium. Quickly add 300 to 400 microliters of 1%low melting point agarose containing tricaine to create a dome.
Using forceps, position the embryos so they lie straight in contact with the glass with the ventral side facing up. Next, deposit approximately 0.5 microliters of magnetic beads on top of the agarose. Then add a drop of Danios medium containing 0.02%tricaine on top of the agarose and the beads.
Based on the sample and heart size, select the appropriate bead and use tweezers to place it on top of the embryo yolk. Create a yolk lesion or hole in the center of the yolk using one arm of the tweezers. Place the bead into the lesion and gently push it anteriorly until it reaches the venous pole.
If suction alone is insufficient to move the bead, gently push it toward the atrium, applying enough pressure to guide it into the pericardial cavity. After grafting the magnetic beads into the embryos, add one to two milliliters of Danios medium containing PTU to the glass bottom dish. Then using tweezers, gently break the low melting point agarose and carefully remove the embryos.
Finally, using a Pasteur pipette, transfer the embryos to a new Petri dish containing Danios medium with PTU and allow them to recover and develop at 28.5 degrees Celsius. The magnetic bead was correctly positioned within the atrium of the zebrafish heart, allowing for unobstructed blood flow and no observed hemorrhage with the heart walls maintained.
