heart lab2[v6.0]

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Procedure

1.Prepare a clean working area. Wipe off tools with 70% ethanol.

Prewarm Howard's Ringers solution in petri dish and culture media (2% FCS in DMEM) in organ culture dish to 37ºC on slide warmer.

2.Prepare and sterilize the filter paper discs in the autoclave.

3.Wipe off egg with 70% ethanol. Puncture egg at wide end (above air space) using point of sharp forceps. Carefully remove shell above air space. Using forceps, peel back the shell membrane to expose the embryo.

4. Drop a filter paper disc around 2-day embryos. Hold on to the filter paper with fine forceps and cut around the ring with sharp scissors. Gradually lift the paper disc with the forceps, the embryo should remain within the center of the disc. Transfer to warm Howard's Ringers solution. Dissect away the extra-embryonic membranes.

5. The 3-day embryo can be removed using a plastic spoon. Keeping hold of the embryo with the forceps, cut around it with fine scissors. Pick up a plastic spoon with your cutting hand and slide it under the embryo. Continue to hold onto the embryo, lift with the spoon and transfer to the petri dish filled with Howard's Ringers solution.

6. Using fine forceps, sever the trunk above and below the heart region. Then, carefully remove the dorsal region of the embryo, leaving the heart intact.

7. Identify the sinus venosus, atrium and ventricle. Measure the heart beat, and determine the direction of blood flow.

8. Transfer the heart to warm culture media. Check for a change in heart beat rate or pattern.

9. Incubate in CO₂incubator overnight. Check for beat and evidence of development the next day, and thereafter.

@Cebra-Thomas, 2000

Last Modified: 2 August 2001

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		Procedure
		1.Prepare a clean working area. Wipe off tools with 70% ethanol. Prewarm Howard's Ringers solution in petri dish and culture media (2% FCS in DMEM) in organ culture dish to 37ºC on slide warmer.
		2.Prepare and sterilize the filter paper discs in the autoclave.
		3.Wipe off egg with 70% ethanol. Puncture egg at wide end (above air space) using point of sharp forceps. Carefully remove shell above air space. Using forceps, peel back the shell membrane to expose the embryo.
		4. Drop a filter paper disc around 2-day embryos. Hold on to the filter paper with fine forceps and cut around the ring with sharp scissors. Gradually lift the paper disc with the forceps, the embryo should remain within the center of the disc. Transfer to warm Howard's Ringers solution. Dissect away the extra-embryonic membranes. 5. The 3-day embryo can be removed using a plastic spoon. Keeping hold of the embryo with the forceps, cut around it with fine scissors. Pick up a plastic spoon with your cutting hand and slide it under the embryo. Continue to hold onto the embryo, lift with the spoon and transfer to the petri dish filled with Howard's Ringers solution. 6. Using fine forceps, sever the trunk above and below the heart region. Then, carefully remove the dorsal region of the embryo, leaving the heart intact. 7. Identify the sinus venosus, atrium and ventricle. Measure the heart beat, and determine the direction of blood flow. 8. Transfer the heart to warm culture media. Check for a change in heart beat rate or pattern. 9. Incubate in CO₂incubator overnight. Check for beat and evidence of development the next day, and thereafter.