Diagnostic criteria flowchart for PBD-ZSD. Given the current availability of next generation sequencing panels, clinicians have moved from evaluation of biochemical markers to genetic analysis future reproductive options, carrier testing in relatives, eligibility purposes in clinical trials and for patients that are difficult to diagnose. In difficult cases, it may still be necessary to evaluate cultured fibroblasts, and this may be important also to ascertain responses of specific mutations for future interventional trials. Abbreviations: PBD-ZSD, peroxisome biogenesis disorder-Zellweger spectrum disorder; X-ALD, X-linked adrenoleukodystrophy; VLCFA, very long chain fatty acids.