Test Catalog

Test ID: B12    
Vitamin B12 Assay, Serum

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

Investigation of macrocytic anemia


Workup of deficiencies seen in megaloblastic anemias

Testing Algorithm Delineates situations when tests are added to the initial order. This includes reflex and additional tests.

See Vitamin B12 Deficiency Evaluation in Special Instructions.

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

Vitamin B12 (cobalamin) is necessary for hematopoiesis and normal neuronal function. In humans, it is obtained only from animal proteins and requires intrinsic factor (IF) for absorption. The body uses its vitamin B12 stores very economically, reabsorbing vitamin B12 from the ileum and returning it to the liver; very little is excreted.


Vitamin B12 deficiency may be due to lack of IF secretion by gastric mucosa (eg, gastrectomy, gastric atrophy) or intestinal malabsorption (eg, ileal resection, small intestinal diseases).


Vitamin B12 deficiency frequently causes macrocytic anemia, glossitis, peripheral neuropathy, weakness, hyperreflexia, ataxia, loss of proprioception, poor coordination, and affective behavioral changes. These manifestations may occur in any combination; many patients have the neurologic defects without macrocytic anemia.


Pernicious anemia is a macrocytic anemia caused by vitamin B12 deficiency that is due to a lack of IF secretion by gastric mucosa.


Serum methylmalonic acid and homocysteine levels are also elevated in vitamin B12 deficiency states.

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.

180-914 ng/L

Interpretation Provides information to assist in interpretation of the test results

A serum vitamin B12 level less than 180 ng/L may cause megaloblastic anemia and peripheral neuropathies.


Vitamin B12 levels less than 150 ng/L are considered evidence of vitamin B12 deficiency. Follow-up with a test for antibodies to intrinsic factor (IFBA / Intrinsic Factor Blocking Antibody, Serum) is recommended to identify this potential cause of vitamin B12 malabsorption. For specimens without antibodies and the patient is symptomatic, follow-up testing for vitamin B12 tissue deficiency may be indicated. Consider analysis of methylmalonic acid (MMAS / Methylmalonic Acid, Quantitative, Serum) and/or homocysteine (HCYSP / Homocysteine, Total, Plasma).


Patients with serum vitamin B12 levels between 150 and 400 ng/L are considered borderline deficient and should be evaluated further by functional tests for vitamin B12 deficiency. Plasma homocysteine measurement (HCYSP / Homocysteine, Total, Plasma) is a good screening test where a normal level effectively excludes vitamin B12 and folate deficiency in an asymptomatic patient. However, the test is not specific, and many situations can cause an increased level. In contrast, an increased serum methylmalonic acid (MMAS / Methylmalonic Acid, Quantitative, Serum) level is more specific for cellular-level B12 deficiency and is not increased by folate deficiency.


In patients being evaluated for vitamin B12 deficiency who have intrinsic factor blocking antibodies (IFBA), false elevations of vitamin B12 may occur due to IFBA interference, potentially obscuring a physiological deficiency of vitamin B12. If observed vitamin B12 concentrations are discordant with clinical presentation, measurement of methylmalonic acid (MMAS / Methylmalonic Acid, Quantitative, Serum) should be considered.


See Vitamin B12 Deficiency Evaluation in Special Instructions.

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

Patients who have received a vitamin B12 injection or radiolabeled vitamin B12 injection within the previous 2 weeks may have high serum vitamin B12 levels, which can interfere with this assay leading to falsely elevated results.


Many other conditions are known to cause an increase or decrease in the serum vitamin B12 concentration and should be considered in the interpretation of the assay results, including:


Increased serum vitamin B12

Decreased serum vitamin B12

Ingestion of vitamin C


Ingestion of estrogens


Ingestion of vitamin A


Hepatocellular injury


Myeloproliferative disorder

Ethanol ingestion


Contraceptive hormones






Multiple myeloma


The evaluation of macrocytic anemia requires measurement of both vitamin B12 and folate levels; ideally, they should be measured simultaneously.


Some patients exposed to animal antigens, either in the environment or as part of treatment or imaging procedure, may have circulating anti-animal antibodies present. These antibodies may interfere with the assay reagents to produce unreliable results.

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

1. Babior BM: The megaloblastic anemias. In: Williams WJ, Beutler E, Lichtman MA et al, eds. Hematology. 5th ed. McGraw-Hill; 1995:471-490

2. Roberts NB, Taylor A, Sodi R: Vitamins and trace elements. In: Rifai N, Horvath AR, Wittwer CT, eds. Tietz Textbook of Clinical Chemistry and Molecular Diagnostics. 6th ed. Elsevier; 2018:chap 37

3. Klee GG: Cobalamin and folate evaluation: measurement of methylmalonic acid and homocysteine vs vitamin B12 and folate. Clin Chem. 2000 August;46(8 Pt 2):1277-1283

4. Allen LH, Miller JW, de Groot L, et al. Biomarkers of nutrition for development (BOND): Vitamin B-12 review. J Nutr. 2018;148(suppl_4):1995S–2027S. doi: 10.1093/jn/nxy201

5. Wolffenbuttel BHR, Wouters HJCM, Heiner-Fokkema MR, van der Klauw MM: The many faces of cobalamin (vitamin B12) deficiency. Mayo Clin Proc Innov Qual Outcomes. 2019;3(2):200-214 Published 2019 May 27. doi: 10.1016/j.mayocpiqo.2019.03.002

6. Hannibal L, Lysne V, Bjorke-Monsen AL, et al: Biomarkers and algorithms for the diagnosis of vitamin B12 deficiency [published correction appears in Front Mol Biosci. 2017 Aug 08;4:53]. Front Mol Biosci. 2016;3:27. doi: 10.3389/fmolb.2016.00027

7. Green R, Kinsella LJ: Current concepts in the diagnosis of cobalamin deficiency. Neurology. 1995;45:1435-1440

8. Lahner E, Annibale B: Pernicious anemia: new insights from a gastroenterological point of view. World J Gastroenterol. 2009 Nov 7;15(41):5121-5128

9. Bizzaro N, Antico A: Diagnosis and classification of pernicious anemia. Autoimmun Rev. 2014;13(4-5):565-568

10. Toh BH: Pathophysiology and laboratory diagnosis of pernicious anemia. Immunol Res. 2017;65(1):326-330

Special Instructions Library of PDFs including pertinent information and forms related to the test