Eur J Pediatr (2011) 170:103–106 105 Fig. 1 PAS diastase positive granules in the hepatocytes. H & E, ×400 concentration 1/40,000 without antigen retrieval in DAKO immunostainer) showed clearly the accumulation of AAT protein polymers in the hepatocytes (Fig. 2). Microbiology PCR studies to identify viral agents showed no cytomegalo- virus in liver. Additional studies in microbiology showed no growth of bacterial of viral agents. Molecular analysis Genomic DNA from peripheral bloods cells was isolated according to standard procedures. Mutation analysis of Serpin 1 was performed by direct sequence analysis of the coding exons (2–5; exon 2 split into two). Primers, con- taining an M13 tail, used were: Exon 2-1 Exon 2-2 Exon 3 Exon 4 Exon 5 Forward: 5′ cacttccacgtggtgtcaat Reverse: 5′ ggttgagggtacggaggagt Forward: 5′ ttcttctccccagtgagcat Reverse: 5′ gaatccacgctgaaaagcat Forward: 5′ ggaggggactcatggtttct Reverse: 5′ tagcagtgacccagggatgt Forward: 5′ tagtgtgggtggaggacaca Reverse: 5′ cagcctgggtcttcatttgt Forward: 5′ gtccacgtgagccttgct Reverse: 5′ ggaccagctcaacccttctt PCR reactions were performed using standard conditions (25-μL reaction, containing 100-ng genomic DNA; 2′ at 94°C, followed by 29 cycles of 30″ at 94°C, 30″ at 60°C and 30″ at 72°C and a final extension of 10′ at 72°C). Sequence analysis of exon 5 of Serpin 1 showed a homozygous pattern for p.Glu342Lys in our patient. In Fig. 2 Immunohistochemical staining showing AAT protein deposition in the hepatocyte (×200) agreement with the homozygous pattern, the parents of the patient were heterozygous for this pathogenic mutation. Discussion Alpha-1-antitrypsin (AAT) deficiency has an incidence of 0.05% and can be caused by homozygosity for PI*Z [9]. The mutant Z protein can form abnormal polymers of AAT within the rough ER of hepatocytes. Chains of these polymers become interwoven to form insoluble inclusions resulting in reduced levels of this protein in the bloodstream. These inclusions form the hallmark of AAT deficiency in pathological diagnosis [8]. Clinical progression to liver disease is complex, with Z allele homozygous neonates developing hepatitis and cholestasis in about 10% of the cases [5]. Here, we reported our findings in a 4-week-old female neonate who showed harmless-looking warning bleeds at the age of 2 weeks and cascade of complications leading to her death as a result of intracranial VKDBD under standard vitamin K prophylaxis [10]. Although VKDB is described earlier as a complication of AAT deficiency in neonates, there are still two important lessons we can learn from this patient. The clinical lesson is the importance of early recognition of the warning bleeds or the cholestasis. Various reports showed a correlation between cholestasis and VKDB due to fat malabsorption. It is also reported that adjustment of the standard vitamin K prophylaxis protocols could prevent the complications due to VKDB which also could have been the case in this patient [12]. The diagnostic lesson is to use pathology as an instrument in combination with verifying genotyping to diagnose AAT deficiency. This statement contrasts with the earlier studies where the cytoplasmic inclusions were not seen in children