| 26 |
There are too many embryos to easily process in 50-60 min |
More mice superovulated than is necessary |
Only superovulate 10-15 mice per session |
| 27 |
Embryos are being lost when manipulated using a mouth pipette |
Pipette tip is flicked against cell culture dish |
Use a cell culture dish with low edges or an unpturned dish with drops of medium |
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Embryos are sucked up beyond the tip of the pipette |
Aspirate a small amount of M2 into the pipette tip and include a small air bubble to sit behind the embryos when manipulating |
| 39 |
Embryos are not lysed but cell division looks abnormal |
Embryo patterning differs in the absence of sperm entry |
Allow the embryos to culture for longer |
| 42 |
Blastocyst is close to breaching it's zona and hatching |
The blastocysts were at an optimal stage for acid treatment over night |
Perform the zona removal, being prepared to take them out of the acid sooner |
| 46 |
The zona does not dissolve rapidly |
The acid Tyrodes is not sufficiently warm |
Warm the acid in an incubator at 37 °C and use a heat plate on the microscope |
| 49 |
It is hard to visualise the dissolving of the zona |
Magnification of the microscope is insufficient |
Use x100 magnification capable objective |
| 50 |
Zona disappears but then reappears when in M2 |
Variation in osmotic gradient between mediums may give the impression the zona is shrinking away |
Ensure the acid is suitably warmed |
| 52 |
Embryos are lost when retrieving them from the vial |
Too much medium in vial |
Use no more than 50 μl in the vial. |
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Flush the inner sides and lid of the vial with M2 to dislodge and collect embryos that may have got attached to these surfaces |
