What blood tests are used to diagnose vitamin B12 deficiency?

Blood Tests for Diagnosing Vitamin B12 Deficiency

Use either total B12 (serum cobalamin) or active B12 (serum 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 Options

You have two acceptable choices for initial screening 1:

• Total B12 (serum cobalamin): Costs approximately £2 per test with rapid turnaround in local laboratories 1
• Active B12 (serum 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

Despite active B12 being more accurate, evidence is insufficient to mandate it as the standard over total B12, so either test is acceptable when clinical suspicion exists 1.

Interpreting Initial Results

Total B12 Thresholds 1:

• <180 ng/L (<133 pmol/L): Confirmed vitamin B12 deficiency—start treatment immediately
• 180-350 ng/L (133-258 pmol/L): Indeterminate result—proceed to MMA testing
• >350 ng/L (>258 pmol/L): Deficiency unlikely

Active B12 Thresholds 1:

• <25 pmol/L: Confirmed vitamin B12 deficiency—start treatment immediately
• 25-70 pmol/L: Indeterminate result—proceed to MMA testing
• >70 pmol/L: Deficiency unlikely

Second-Line Confirmatory Testing

When to Measure Methylmalonic Acid (MMA)

Order MMA testing when initial B12 results fall in the indeterminate range to identify functional B12 deficiency that reflects actual cellular B12 status 1, 2. This is critical because:

• 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 metabolites when serum B12 was normal 2
• MMA has 98.4% sensitivity for B12 deficiency 2
• Standard serum B12 testing misses functional deficiency in up to 50% of cases 2

MMA Testing Considerations

• 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
• Do not use MMA as first-line screening—it is not cost-effective due to expense and processing delays 2

Important MMA Interpretation Caveats

MMA can be falsely elevated in 2:

• Hypothyroidism
• Renal insufficiency (elevated creatinine)
• Hypovolemia

Interpret MMA cautiously in these conditions and consider lowering MMA after vitamin B12 treatment to confirm true deficiency in patients with elevated creatinine 2.

Additional Testing Considerations

Complete Blood Count

Order a CBC to assess for 3, 4:

• Macrocytic anemia
• Megaloblastic changes

Critical pitfall: Megaloblastic anemia may be absent in one-third of cases, and concurrent folic acid supplementation can mask the anemia while allowing irreversible neurologic damage to progress 2, 5.

When NOT to Test Further

Do not order B12 testing in patients with levels >350 ng/L (>258 pmol/L) unless there are specific clinical indications, as this makes deficiency unlikely 1, 2.

Diagnostic Algorithm Summary

1. Start with total B12 or active B12 based on local availability and cost 1
2. If <180 ng/L (total) or <25 pmol/L (active): Diagnose deficiency and treat immediately 1
3. If 180-350 ng/L (total) or 25-70 pmol/L (active): Order MMA testing 1
4. If MMA elevated: Confirm functional B12 deficiency and treat 1, 2
5. If >350 ng/L (total) or >70 pmol/L (active): Deficiency unlikely, consider alternative diagnoses 1

Common Clinical Pitfalls

• Never rely solely on serum B12 to rule out deficiency, especially in patients >60 years where metabolic deficiency is common despite normal serum levels 2
• Do not give folic acid without B12 in suspected deficiency—folic acid doses >0.1 mg daily may produce hematologic remission while allowing irreversible neurologic damage to progress 5
• Recognize that vitamin B12 levels alone are not always a good predictor of deficiency due to methodological problems affecting sensitivity and specificity 6
• Check medications: Metformin (>4 months use), proton pump inhibitors (>12 months), H2 blockers, colchicine, and anticonvulsants can all cause B12 deficiency 2, 6, 3