My thesis involves experimental study with Wistar puppies. I am dividing the puppies equally in relation to the groups. However I only have 4 mothers and I will use all of their puppies. There is 4 groups, male and females (8 rats per gp). I need, in total, 64 puppies.

Here comes the question, will this be a problem in the statistical analysis? I know that there is a calculation that can be done to assess "the litter effect" in studies of this type. Is 6 mothers good to avoid this effect?

Do you think I should ask for more female mothers? So that the rats are not from the same litter. there is some calculation for this?

Thanks in advance,

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