Between the biopsy and PGT lab
Several important steps will happen between the biopsy and sample delivery to the PGT laboratory.
As it is often a case for diagnostic testing, pre-analytical and analytical phases of PGT also take place in different laboratories: biopsy sample placement into the tube occurs in the IVF laboratory, while the testing is performed in the molecular laboratory.
The pre-analytical phase of PGT is logistically and technically challenging for several reasons. Logistically, each embryo has to be tracked from the biopsy Petri dish to the culture Petri dish and finally to the freezing straw. Each biopsy sample has to be tracked from the biopsy dish into the tiny PCR tube.
Embryos and biopsy tracking errors
Until recently, there were no commercially available automated systems to track individual biopsy samples and embryos. Only a short while ago, a single company started to offer such a system. It will take several years to see whether it is effective and ergonomic. We are using our in-house developed system to facilitate tracking the embryo, biopsy sample, and freezing straw. It includes the holder for the tube, the microscope shield protecting the vitrification straw from contact with the microscope stage, and the Vitrocube™, holding the vitrification straws in the numbered position.
Biopsy tubing errors
Even under a state-of-the-art binocular microscope, the sample looks like a tiny grey spec.
The biopsy sample is tiny and can sometimes get lost when transferring between the wash buffer droplets or because it is stuck to the pipette. If this happens during the transfer into the tube, the sample loss may not even be apparent to an embryologist.
Even an experienced embryologist may, on a rare occasion, confuse an embryo biopsy sample with some other spec present in the drop. If a piece of debris (usually plastic) is picked up instead of an actual sample, there will be no amplification.
Contamination with cumulus cells
A clump of cumulus cells may also be, at least in theory, occasionally confused with the actual biopsy. Even though good care is usually taken to remove all corona-cumulus cells, this is not always possible, because sometimes corona cells strongly adhere to the zona pellucida making them difficult to remove without aggressive stripping that can damage an oocyte or an embryo. Because this risk is almost impossible to eliminate completely, in the case of single-gene disorder there is an additional step during testing that ensures that the amplified sample comes from the embryo and not the cumulus. This step requires the blood from male and female partners to make sure that amplified DNA belongs to the embryo. This makes single gene testing very expensive, but the additional cost in cases of single-gene disorders is fully justified. At the same time, due to a much lower risk level with numerical chromosome testing in PGT, the step verifying the origin of the DNA in the biopsy sample is omitted making it impossible to rule out contamination with cumulus cells or any other type of DNA contamination.
Loss of the biopsy sample after transfer to the tube
The biopsy sample may occasionally get stuck to the wall of the tube and dry or its DNA may deteriorate in some way before the sample reaches the testing laboratory. This is an uncommon occurrence, but if that happens no result will be obtained.