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HelpTest Code 2425D 25-Hydroxyvitamin D:24,25-Dihydroxyvitamin D Ratio, Serum
Ordering Guidance
The preferred initial test for assessing vitamin D status and most accurately reflects the body's vitamin D stores is the 25-hydroxyvitamin D test; order 25HDN / 25-Hydroxyvitamin D2 and D3, Serum.
In the presence of kidney disease or hypercalcemia, testing of 1,25-dihydroxy vitamin D may be needed to adequately assess vitamin D status; order DHVD / 1,25-Dihydroxyvitamin D, Serum.
Specimen Required
Supplies: Sarstedt Aliquot Tube, 5 mL (T914)
Collection Container/Tube: Red top
Submission Container/Tube: Plastic vial
Specimen Volume: 1.7 mL Serum
Collection Instructions: Within 2 hours of collection, centrifuge and aliquot serum into a plastic vial.
Useful For
Screening for CYP24A1 deficiency
Evaluation of parathyroid hormone-independent hypercalcemia
Evaluation of recurrent nephrolithiasis or nephrocalcinosis associated with suspected vitamin D-mediated disease
Selection of patients for confirmatory molecular testing
Profile Information
| Test ID | Reporting Name | Available Separately | Always Performed |
|---|---|---|---|
| 2425R | 24,25 Dihydroxy Vitamin D | No | Yes |
| 25HDN | 25-Hydroxyvitamin D2 and D3, S | Yes | Yes |
Method Name
Liquid Chromatography Tandem Mass Spectrometry (LC-MS/MS)
Portions of this test are covered by patents held by Quest Diagnostics
Reporting Name
25HDN:24,25 Dihydroxy VitD Ratio, SSpecimen Type
SerumSpecimen Minimum Volume
Serum: 1.1 mL
Specimen Stability Information
| Specimen Type | Temperature | Time |
|---|---|---|
| Serum | Refrigerated (preferred) | 14 days |
| Frozen | 30 days | |
| Ambient | 7 days |
Reference Values
Interpretative commentary provided based on 25-hydroxyvitamin D (25HDN) to 24,25-dihydroxyvitamin D (24,25D) ratio result.
25HDN to 24,25D Ratio
<25: Consistent with normal CYP24A1 activity, although overlap with heterozygous carriers has been reported.
25-80: May be observed in vitamin D deficiency or heterozygous CYP24A1 variants. Correlation with clinical findings is recommended. Molecular testing may be appropriate when clinical suspicion is high.
>80: Strongly suggests markedly reduced CYP24A1 activity. Confirmatory molecular testing is recommended.
Interpretation
Results should be interpreted in conjunction with serum calcium, parathyroid hormone, 25-hydroxyvitamin D (25HDN), 1,25-dihydroxyvitamin D test results and clinical presentation.
If the 25HDN concentration is less than 20 ng/mL, the ratio may be falsely elevated because of reduced substrate availability and should be interpreted cautiously.
Interpretation guidance for 25HDN to 24,25D ratios:
Ratio <25: Consistent with normal CYP24A1 activity, although overlap with heterozygous carriers has been reported.
Ratio 25-80: May be observed in vitamin D deficiency or heterozygous CYP24A1 variants. Correlation with clinical findings is recommended. Molecular testing may be appropriate when clinical suspicion is high.
Ratio >80: Strongly suggests markedly reduced CYP24A1 activity. Confirmatory molecular testing is recommended.
Clinical Reference
1. Kaufmann M, Gallagher JC, Peacock M, et al. Clinical utility of simultaneous quantitation of 25-hydroxyvitamin D and 24,25-dihydroxyvitamin D by LC-MS/MS involving derivatization with DMEQ-TAD. J Clin Endocrinol Metab. 2014;99(7):2567-2574. doi:10.1210/jc.2013-4388
2. Ketha H, Kumar R, Singh RJ. LC-MS/MS for identifying patients with CYP24A1 mutations. Clin Chem. 2016;62(1):236-242
3. Demay MB, Pittas AG, Bikle DD, et al. Vitamin D for the Prevention of Disease: An Endocrine Society Clinical Practice Guideline. J Clin Endocrinol Metab. 2024;109(8):1907-1947. doi:10.1210/clinem/dgae290
4. Institute of Medicine (US) Committee to Review Dietary Reference Intakes for Vitamin D and Calcium, Food and Nutrition Board. In: Ross AC, Taylor CL, Yaktine AL, Del Valle HB, eds. Dietary Reference Intakes for Calcium and Vitamin D. National Academies Press: 2011
5. Vitamin D-Fact Sheet for Health Professionals. US Department of Health and Human Services, National Institutes of Health. Updated June 27, 2025. Accessed August 12, 2026. Available at https://ods.od.nih.gov/factsheets/VitaminD-HealthProfessional/
6. Fraser W. Bone and mineral metabolism. In: Rifai N, Horvath AR, Wittwer CT, eds. Tietz Fundamentals of Clinical Chemistry and Molecular Diagnostics. 8th ed. Elsevier; 2020:466-487
7. Herrmann M. Assessing vitamin D metabolism - four decades of experience. Clin Chem Lab Med. 2023;61(5):880-894. doi:10.1515/cclm-2022-1267
8. Shah AD, Hsiao EC, O'Donnell B, et al. Maternal hypercalcemia due to failure of 1,25-dihydroxyvitamin-D3 catabolism in a patient with CYP24A1 mutations. J Clin Endocrinol Metab. 2015;100(8):2832-6. doi:10.1210/jc.2015-1973
Day(s) Performed
Wednesday
Report Available
2 to 15 daysPerforming Laboratory
Mayo Clinic Laboratories in Rochester
Test Classification
This test was developed and its performance characteristics determined by Mayo Clinic in a manner consistent with CLIA requirements. It has not been cleared or approved by the US Food and Drug Administration.CPT Code Information
82306
82542
LOINC Code Information
| Test ID | Test Order Name | Order LOINC Value |
|---|---|---|
| 2425D | 25HDN:24,25 Dihydroxy VitD Ratio, S | 94674-9 |
| Result ID | Test Result Name | Result LOINC Value |
|---|---|---|
| 2897 | 25-Hydroxy D2 | 49054-0 |
| 63416 | 25HDN:24,25 Dihydroxy VitD Ratio, S | 94673-1 |
| 2898 | 25-Hydroxy D3 | 1989-3 |
| 83670 | 25-Hydroxy D Total | 62292-8 |
| 90601 | 24,25-Dihydroxy VitD Total | 94672-3 |