Test Catalog

Test Id : 2425D

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25-Hydroxyvitamin D:24,25-Dihydroxyvitamin D Ratio, Serum

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Overview

Test Catalog

Useful For
Suggests clinical disorders or settings where the test may be helpful

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
A profile is a group of laboratory tests that are ordered and performed together under a single Mayo Test ID. Profile information lists the test performed, inclusive of the test fee, when a profile is ordered and includes reporting names and individual availability.

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
A short description of the method used to perform the test

Liquid Chromatography Tandem Mass Spectrometry (LC-MS/MS)

Portions of this test are covered by patents held by Quest Diagnostics

NY State Available
Indicates the status of NY State approval and if the test is orderable for NY State clients.

Yes

Reporting Name
Lists a shorter or abbreviated version of the Published Name for a test

25HDN:24,25 Dihydroxy VitD Ratio, S

Aliases
Lists additional common names for a test, as an aid in searching

2425D

2425R

2425

24,25-Dihydroxyvitamin D

Hydroxycalcifediol

Hydroxycalcidiol

P450CC24

Cytochrome P450 Family 24

Cytochrome P450 Subfamily XXIV

Vitamin D 24-Hydroxylase

25-Hydroxyvitamin D 24-Hydroxylase

Dihydrocholesterol

2425OHD

CYP24A1

Calciferol

Ergocalciferol

Calcidiol

Cholecalciferol

Vitamin D Assay

CYP24

Specimen Type
Describes the specimen type validated for testing

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
Defines the optimal specimen required to perform the test and the preferred volume to complete testing

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.

Specimen Minimum Volume
Defines the amount of sample necessary to provide a clinically relevant result as determined by the testing laboratory. The minimum volume is sufficient for one attempt at testing.

Serum: 1.1 mL

Reject Due To
Identifies specimen types and conditions that may cause the specimen to be rejected

Gross hemolysis OK
Gross lipemia Reject
Gross icterus OK

Specimen Stability Information
Provides a description of the temperatures required to transport a specimen to the performing laboratory, alternate acceptable temperatures are also included

Specimen Type Temperature Time Special Container
Serum Refrigerated (preferred) 14 days
Ambient 7 days
Frozen 30 days

Useful For
Suggests clinical disorders or settings where the test may be helpful

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

Clinical Information
Discusses physiology, pathophysiology, and general clinical aspects, as they relate to a laboratory test

The ratio of 25-hydroxyvitamin D (25HDN) to 24,25-dihydroxyvitamin D (24,25D) provides an estimate of CYP24A1 enzymatic activity. CYP24A1 is responsible for inactivation of both 25HDN and 1,25-dihydroxyvitamin D (DHVD). Loss of function variants in the CYP24A1 gene impair vitamin D catabolism, resulting in a phenotype characterized by suppressed serum parathyroid hormone (PTH), increased serum DHVD concentrations, hypercalcemia, and hypercalciuria that may lead to nephrolithiasis. Because 24,25D production depends on substrate availability (25HDN concentration) and CYP24A1 activity, interpretation of the ratio requires simultaneous measurement of both analytes in the same specimen. The ratio is most useful in patients with suspected CYP24A1 deficiency and should be interpreted in conjunction with serum calcium, PTH, 25HDN, DHVD test results and clinical presentation.

Reference Values
Describes reference intervals and additional information for interpretation of test results. May include intervals based on age and sex when appropriate. Intervals are Mayo-derived, unless otherwise designated. If an interpretive report is provided, the reference value field will state this.

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
Provides information to assist in interpretation of the test results

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.

Cautions
Discusses conditions that may cause diagnostic confusion, including improper specimen collection and handling, inappropriate test selection, and interfering substances

For accurate interpretation, 25-hydroxyvitamin D and 24,25-dihydroxyvitamin D must be measured from the same specimen using the validated method used to establish the reference values. Calculating the ratio with analytes measured separately or by different methods may produce misleading results.

 

Vitamin D deficiency may increase the ratio independent of CYP24A1 deficiency. Results should always be interpreted in conjunction with serum calcium, parathyroid hormone, and the overall clinical presentation.

 

Serum gel and SST tubes significantly increase chances of interference with the 24,25 dihydroxyvitamin result and may increase laboratory time. Therefore, red-top tubes are preferred for specimen collection.

Clinical Reference
Recommendations for in-depth reading of a clinical nature

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

Method Description
Describes how the test is performed and provides a method-specific reference

Analytes of interest and deuterated internal standard are extracted, derivatized, and analyzed by liquid chromatography tandem mass spectrometry.(Ketha H, Kumar R, Singh RJ. LC-MS/MS for identifying patients with CYP24A1 mutations. Clin Chem. 2016;62(1):236-242)

PDF Report
Indicates whether the report includes an additional document with charts, images or other enriched information

No

Day(s) Performed
Outlines the days the test is performed. This field reflects the day that the sample must be in the testing laboratory to begin the testing process and includes any specimen preparation and processing time before the test is performed. Some tests are listed as continuously performed, which means that assays are performed multiple times during the day.

Wednesday

Report Available
The interval of time (receipt of sample at Mayo Clinic Laboratories to results available) taking into account standard setup days and weekends. The first day is the time that it typically takes for a result to be available. The last day is the time it might take, accounting for any necessary repeated testing.

2 to 15 days

Specimen Retention Time
Outlines the length of time after testing that a specimen is kept in the laboratory before it is discarded

2 weeks

Performing Laboratory Location
Indicates the location of the laboratory that performs the test

Mayo Clinic Laboratories - Rochester Superior Drive
CLIA Number: 24D1040592

Fees
Several factors determine the fee charged to perform a test. Contact your U.S. or International Regional Manager for information about establishing a fee schedule or to learn more about resources to optimize test selection.

  • Authorized users can sign in to Test Prices for detailed fee information.
  • Clients without access to Test Prices can contact Customer Service 24 hours a day, seven days a week.
  • Prospective clients should contact their account representative. For assistance, contact Customer Service.

Test Classification
Provides information regarding the medical device classification for laboratory test kits and reagents. Tests may be classified as cleared or approved by the US Food and Drug Administration (FDA) and used per manufacturer instructions, or as products that do not undergo full FDA review and approval, and are then labeled as an Analyte Specific Reagent (ASR) product.

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
Provides guidance in determining the appropriate Current Procedural Terminology (CPT) code(s) information for each test or profile. The listed CPT codes reflect Mayo Clinic Laboratories interpretation of CPT coding requirements. It is the responsibility of each laboratory to determine correct CPT codes to use for billing.

CPT codes are provided by the performing laboratory.

82306

82542

LOINC® Information
Provides guidance in determining the Logical Observation Identifiers Names and Codes (LOINC) values for the order and results codes of this test. LOINC values are provided by the performing laboratory.

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
Applies only to results expressed in units of measure originally reported by the performing laboratory. These values do not apply to results that are converted to other units of measure.
2897 25-Hydroxy D2 49054-0
2898 25-Hydroxy D3 1989-3
83670 25-Hydroxy D Total 62292-8
90601 24,25-Dihydroxy VitD Total 94672-3
63416 25HDN:24,25 Dihydroxy VitD Ratio, S 94673-1

Test Setup Resources

Setup Files
Test setup information contains test file definition details to support order and result interfacing between Mayo Clinic Laboratories and your Laboratory Information System.

Excel | Pdf

Sample Reports
Normal and Abnormal sample reports are provided as references for report appearance.

Normal Reports | Abnormal Reports

SI Sample Reports
International System (SI) of Unit reports are provided for a limited number of tests. These reports are intended for international account use and are only available through MayoLINK accounts that have been defined to receive them.

SI Normal Reports | SI Abnormal Reports