Diagnostic Workup for Suspected Hypophosphatasia
When hypophosphatasia is suspected based on bone pain, fractures, premature tooth loss, or unexplained musculoskeletal symptoms, immediately measure serum alkaline phosphatase (ALP), serum pyridoxal 5'-phosphate (vitamin B6), urinary phosphoethanolamine, and proceed with ALPL gene testing to confirm the diagnosis. 1
Initial Laboratory Testing
The diagnostic workup should prioritize the following tests in order:
Serum alkaline phosphatase (ALP): Persistently low ALP (on majority of measurements) is the key biochemical hallmark that warrants full hypophosphatasia workup 1, 2. Markedly reduced serum ALP activity is the defining feature of this condition 3, 4.
Serum pyridoxal 5'-phosphate (PLP/vitamin B6): High serum concentrations represent a sensitive and specific biochemical marker for hypophosphatasia 2. This substrate accumulates due to TNSALP enzyme deficiency 5.
Urinary phosphoethanolamine (PEA): Elevated levels support the diagnosis, though this marker is less reliable than PLP 6, 2. Plasma ALP and urinary phosphoethanolamine are specifically recommended for childhood-type hypophosphatasia 6.
Serum PTH and 25(OH) vitamin D: These should be checked to exclude other metabolic bone disorders in the differential diagnosis 1.
Age-Specific Clinical Features to Assess
Newborns and Infants (Perinatal/Infantile Forms)
- Severe undermineralization and bone deformity on radiographs 6
- Respiratory complications and premature craniosynostosis 3
- Widespread demineralization and rachitic changes in metaphyses 3
- Hypercalcemia and hyperphosphataemia due to blocked mineral entry into skeleton 2
- Pyridoxine-dependent seizures from vitamin B6 deficiency 5, 2
Children (Childhood Type)
- Premature loss of primary teeth (fully rooted teeth exfoliating early) 6, 3
- Rachitic bone changes with fractures 6
- Skeletal deformities, short stature, and waddling gait 3
- Undermineralization of bones on imaging 6
Adults
- Stress fractures and recurrent fractures 1, 3
- Skeletal pain and thigh pain 3, 7
- Chondrocalcinosis and calcific periarthritis 7, 2
- Dental problems including severe caries 3
- Marked osteoarthropathy 3
Genetic Confirmation
ALPL gene sequencing: Identifies TNSALP mutations and confirms molecular diagnosis 4, 2. Approximately 95% of mutations are detected in severe (perinatal and infantile) forms 3. Around 300 predominantly missense ALPL mutations have been identified 2.
Gene analysis showing a pathogenic variant in ALPL confirms the diagnosis, though a substantial proportion of patients show normal results after sequencing ALPL exons 7.
Radiographic Evaluation
Obtain skeletal radiographs to assess for characteristic findings including undermineralization, fractures, and rachitic changes 6, 3.
In severe pediatric cases, pathognomonic radiographic changes aid diagnosis, showing marked bone hypomineralization 7, 2.
Critical Differential Diagnosis Considerations
The differential must exclude:
X-linked hypophosphatemia: Presents with hypophosphatemia but elevated (not low) ALP and elevated FGF23 1. This is the opposite ALP pattern from hypophosphatasia.
Osteogenesis imperfecta: May present with fractures and blue sclera, but typically has normal or elevated ALP 6, 8. Requires COL1A1/COL1A2 mutation testing if suspected 6.
Non-accidental injury: Must be considered in infants with unexplained fractures, though hypophosphatasia can be missed and incorrectly attributed to abuse 6.
Immediate Management Considerations
Discontinue antiresorptive therapy immediately if the patient is on bisphosphonates or denosumab, as these drugs are absolutely contraindicated in hypophosphatasia and may worsen symptoms 1.
Refer to an endocrinologist or metabolic bone disease specialist for confirmed or suspected hypophosphatasia, particularly if enzyme replacement therapy (asfotase alfa) is being considered 1, 2.
Common Pitfalls to Avoid
Do not dismiss low ALP as clinically insignificant—persistently low values demand investigation 1, 7.
Do not treat presumed "osteoporosis" with bisphosphonates without first excluding hypophosphatasia in patients with unexplained fractures and low ALP 1.
Blue sclera can occur normally in infants before 12 months of age, but beyond infancy requires pathologic investigation 6, 8.
In infants with fractures, do not rely solely on clinical examination—biochemical or DNA testing may be necessary to distinguish hypophosphatasia from other conditions 6.