This study utilises retrospectively retrieved NBS (at birth) and dried blood spots (DBS; created at the time of assessment) collected from a large Australian cohort of children affected with FXS to determine: (i) sensitivity and specificity of FREE2m for FM alleles in NBS; (ii) whether the previously reported epigenotype–phenotype associations for FREE2m in older children [20,21], are conserved from birth; (iii) if and how FREE2m levels in NBS versus DBS are related to FMR1 transcriptional changes, intellectual functioning and autism features throughout neurodevelopment in FXS in both sexes. This evidence concerns the gene FMR1 and fragile X syndrome.