Nowadays in-ovo injection is very popular for administering Marek's vaccine. With in-ovo injection, the hatching eggs are injected during tranfer. Due to the early adminstration that is achieved in this way, chicks are able to build-up some immunity at the time of hatch.

Crucial for in-ovo injection is the moment of injection. In an optimal situation, the vaccine should be delivered in the amniotic fluid, and then will be swallowed by the embryo. If the vaccine is delivered in the allantoic fluid, the vaccine will not be taken up by the embryo but wasted instead. If the vaccine is delivered either in the neck or in the breast of the embryo, the embryo will be vaccinated as well, but there is more chance to damage the embryo.

The site of injection depends on the development of the embryo. The needle is brought into the egg at a fixed depth, and the embryo should be developed exactly as expected so that the needle hits the amniotic fluid.
- If the embryo is not developed enough, the vaccine is delivered in the allantoic fluid, as this tissue lies on top of the amniotic fluid.
- if the embryo is developed sligthly too much, the vaccine is delivered in the breast muscle.
- if the embryo is developed even more, the vaccine is delivered in the neck.

The speed of development of the embryo is a function of the temperature inside the shell. As this temperature is not equal for each machine and each setting, nor for each place in the machine, the development of the embryo should be checked at regular intervals.
Especially a too low speed of development or a too early injection is harmful, as birds will then not be vaccinated and not develop the desired immune response.

The development can be checked by injecting a sample of eggs with a dye, and then open up the eggs to check the place of injection.
Instead of injecting with a dye, also the length of the embryo can be checked, as this will respond with the level of development and the place of injection.

Another method is to not calculate the exact moment from start of the incubation process, but calculate back from the moment of pulling. Calculating back from the moment of pulling will give a more adequate moment of development of the embryo than counting the days and hours after start of incubation.