What laboratory tests would you recommend for a patient with suspected vitamin D deficiency?

Laboratory Testing Recommendations for Suspected Vitamin D Deficiency

Yes, a vitamin D level is appropriate, but the specific test you should order is a serum 25-hydroxyvitamin D [25(OH)D] level, which should measure both 25(OH)D2 and 25(OH)D3 forms. 1, 2

Primary Laboratory Test

• Order a total serum 25-hydroxyvitamin D [25(OH)D] test as this is the most reliable biochemical marker for defining vitamin D stores and represents the standard test for vitamin D status 2, 3, 4
• The assay must measure both 25(OH)D2 (ergocalciferol) and 25(OH)D3 (cholecalciferol) forms, as both contribute to total vitamin D status 1, 2
• Serum is the sample of choice; avoid gel tubes until more data are available 2

Additional Laboratory Tests to Consider

Depending on the clinical context, you should also consider ordering:

For Bone Health and Calcium Metabolism Assessment

• Serum calcium (corrected for albumin) - to evaluate for hypocalcemia or hypercalcemia 1
• Serum phosphorus - particularly if GFR <30 mL/min per 1.73 m² 1
• Intact parathyroid hormone (iPTH) - especially if 25(OH)D is low, as vitamin D deficiency can cause secondary hyperparathyroidism 1
• Serum albumin - needed to correct calcium levels and assess nutritional status 1

For Chronic Kidney Disease Patients (GFR <30 mL/min per 1.73 m²)

• Calcium and phosphorus every 3 months 1
• iPTH at least once initially, then every 3 months if calcium/phosphorus abnormal 1
• 25(OH)D level if iPTH ≥100 pg/mL (or 1.5 times upper limit of normal) 1

For Anemia Evaluation (if GFR <30 mL/min per 1.73 m²)

• Hemoglobin - check every 3 months 1
• Complete iron studies - if hemoglobin <12 g/dL (women) or <13 g/dL (men) 1

Important Testing Considerations

Assay Selection and Quality

• Be aware that variability between assay methods and laboratories can range from 10% to 20% 2, 5
• Classification of samples as "deficient" or "nondeficient" can vary by 4% to 32% depending on which assay is used 2
• Verify that your laboratory participates in the CDC's Vitamin D Standardization Certification Program to ensure quality and comparability of results 2

Timing of Testing

• Wait at least 3 months after initiating vitamin D supplementation before retesting to allow serum levels to reach plateau and reflect true response to therapy 1, 6, 2
• Consider seasonal variation - vitamin D levels are typically lowest at the end of winter 7

Target Levels for Interpretation

• Target 25(OH)D level should be at least 30-40 ng/mL for optimal health benefits in patients with or at risk for musculoskeletal problems, cardiovascular disease, autoimmune disease, and cancer 1, 6, 2
• Choosing a threshold of 30 ng/mL ensures the patient has a true concentration >20 ng/mL when accounting for measurement uncertainty 1
• Upper safety limit is 100 ng/mL 1, 6

Common Pitfalls to Avoid

• Do not order 1,25-dihydroxyvitamin D [1,25(OH)₂D] as the initial test - this is the active hormone form but is not appropriate for assessing vitamin D stores 3, 5
• Do not use assays that only measure 25(OH)D3 - you need both D2 and D3 forms measured, especially in countries where both supplements are available 1, 2
• Do not retest too soon - waiting less than 3 months after starting supplementation will not accurately reflect steady-state levels 1, 6
• Be aware that total serum 25(OH)D may underestimate vitamin D status in African Americans due to differences in vitamin D binding protein 2