What is the workup for suspected vitamin B12 (cobalamin) deficiency?

Workup for Vitamin B12 Deficiency

Start with either total serum B12 or active B12 (holotranscobalamin) as your initial test, then add methylmalonic acid (MMA) testing when results fall in the indeterminate range to confirm functional deficiency. 1

Initial Testing Strategy

First-Line Test Selection

• Total serum B12 is the standard initial test, costing approximately £2 with rapid turnaround in local laboratories 1
• Active B12 (holotranscobalamin) measures the biologically active form available for cellular use and is more accurate, but costs £18 per test with longer turnaround times due to external laboratory processing 1
• Either test is acceptable as first-line; the choice depends on local availability and cost considerations 1

Interpreting Initial B12 Results

For Total Serum B12:

• <180 ng/L (133 pmol/L): Confirmed vitamin B12 deficiency—proceed directly to treatment 1
• 180-350 ng/L (133-258 pmol/L): Indeterminate result requiring MMA testing 1
• >350 ng/L (258 pmol/L): Deficiency unlikely, but consider functional markers if high clinical suspicion persists 1, 2

For Active B12 (Holotranscobalamin):

• <25 pmol/L: Confirmed deficiency 1
• 25-70 pmol/L: Indeterminate—measure MMA 1
• >70 pmol/L: Deficiency unlikely 1

Confirmatory Testing with Methylmalonic Acid

When to Order MMA

• Indeterminate B12 results (180-350 ng/L total B12 or 25-70 pmol/L active B12) 1
• High clinical suspicion despite normal B12, particularly in elderly patients (>60 years) where up to 50% with "normal" serum B12 have metabolic deficiency 2
• Patients with neurological symptoms and borderline B12 levels 2

MMA Interpretation

• MMA >271 nmol/L: Confirms functional B12 deficiency with 98.4% sensitivity 2
• MMA detects an additional 5-10% of patients with B12 deficiency who have low-normal B12 levels 2
• In polyneuropathy patients, 44% had B12 deficiency based solely on abnormal MMA when serum B12 was normal 2

Cost-Effectiveness Considerations

• MMA testing costs £11-80 per test and requires specialized equipment 1
• Cost-effectiveness analysis shows MMA testing before treatment is cost-effective at £3,946 per quality-adjusted life year when B12 results are indeterminate 1
• Using MMA as first-line screening is not cost-effective due to expense and processing delays 2

Additional Diagnostic Tests

Complete Blood Count

• Check for macrocytic anemia (MCV >100 fL), though megaloblastic anemia is absent in one-third of B12-deficient patients 2
• Evaluate for pancytopenia in severe cases 3

Homocysteine (Optional)

• Homocysteine >15 μmol/L supports B12 deficiency diagnosis 2
• Less specific than MMA—elevated in 95.9% of B12 deficiency but also in 91% of folate deficiency 2
• Elevated homocysteine + elevated MMA = B12 deficiency 2
• Elevated homocysteine + normal MMA = folate deficiency 2

Etiologic Workup (After Confirming Deficiency)

When No Clear Cause Exists

Test for pernicious anemia:

• Intrinsic factor antibodies (highly specific for pernicious anemia) 2
• Gastrin levels: Markedly elevated levels (>1,000 pg/mL) indicate pernicious anemia 2
• Anti-parietal cell antibodies (less specific but more sensitive) 2

Evaluate for other causes:

• Helicobacter pylori testing to assess for atrophic gastritis 4
• Tissue transglutaminase (tTG) antibodies and total IgA for celiac disease, found in 13.3% of B12-deficient patients 2

High-Risk Populations Requiring Screening

• Age >75 years: 18.1% have metabolic deficiency; 25% of those ≥85 years have B12 <170 pmol/L 2
• Metformin use >4 months 2, 5
• PPI or H2 blocker use >12 months 2, 5
• Gastric or ileal resection (especially >20 cm of distal ileum) 2, 5
• Autoimmune conditions: thyroid disease, type 1 diabetes, Sjögren syndrome 2, 5
• Inflammatory bowel disease, particularly Crohn's disease with ileal involvement 2, 5
• Vegans or strict vegetarians 3, 4

Critical Clinical Pitfalls to Avoid

Never Give Folic Acid Before Treating B12 Deficiency

• Folic acid may mask anemia while allowing irreversible neurological damage to progress 6, 7
• This is the most dangerous error in B12 deficiency management 6

Don't Rely Solely on Serum B12 in Elderly Patients

• Serum B12 misses functional deficiency in up to 50% of cases in patients >60 years 2
• Always consider MMA testing in elderly patients with suggestive symptoms despite "normal" B12 2

Recognize MMA Limitations

• MMA can be falsely elevated in hypothyroidism, renal insufficiency, and hypovolemia 2
• Interpret MMA cautiously in these conditions 2

Don't Stop B12 Supplements Before Testing

• If patient is already taking B12 supplements, measure MMA as the primary test rather than stopping supplements and waiting 2
• MMA reflects actual cellular B12 status regardless of supplementation 2

Monitoring During Initial Treatment

First 48 Hours

• Monitor serum potassium closely and replace if necessary, as treatment can precipitate hypokalemia 6

Days 5-7 of Therapy

• Repeat hematocrit and reticulocyte count daily from the fifth to seventh days 6
• Continue frequent monitoring until hematocrit normalizes 6

If Inadequate Response

• If reticulocytes have not increased after treatment or don't continue at least twice normal while hematocrit <35%, reevaluate diagnosis or treatment 6
• Repeat iron and folate determinations to identify complicating illness inhibiting marrow response 6

Special Considerations

Pregnancy and Lactation

• Testing recommendations differ for pregnant women—refer to full guidelines 1
• Vitamin B12 requirements increase during pregnancy and lactation 6

Nitrous Oxide Use

• Recreational nitrous oxide use requires different testing approach—refer to full guidelines 1

Post-Bariatric Surgery

• Deficiencies can occur even when serum concentrations are 300 pmol/L (approximately 406 pg/mL) 2
• These patients require lifelong monitoring and supplementation 2