B-12 Tests
Vitamin B-12 Tests
Testing for Pernicious Anemia
Patients are not diagnosed with pernicious anaemia as fast as they should be getting diagnosed after experiencing symptoms of the disease.
The main reason may be that vitamin B 12 deficiency testing methods are not flawless.
British Committee for Standards in Haematology (BCSH) has recently published new, updated guidelines for the Diagnosis and Treatment of Cobalamin (B12) and Folate Disorders, which state that:
The clinical picture is the most important factor in assessing the significance of test results assessing cobalamin status since there is no ‘gold standard’ test to define deficiency.
Here is a link to the BCSH guidelines: https://onlinelibrary.wiley.com/doi/full/10.1111/bjh.12959
Tests to determine patients’ B-12 status
1. Full Blood Count (FBC)
An FBC is one of the most commonly requested blood tests because as its name implies, it gives a full picture or snapshot as to information about the kinds and numbers and sizes of cells in the blood.
Checking the size of the patient’s red blood cells (RBCs) can point to a deficiency in vitamin B12 if the size of the RBCs is increased past a certain range. This enlargement of RBCs is called macrocytosis, and the enlarged RBCs themselves are called megaloblasts. To detect macrocytosis, the test is called Mean Corpuscular Volume (MCV).
However, having large red blood cells does not automatically mean you have a vitamin B12 deficiency. Only 60% of those with vitamin B12 deficiency will have macrocytosis. There is also another problem with relying on MCV to test for B12 deficiency. Some other deficiencies, such as an iron deficiency, can lead to a reduction on the MCV, i.e. iron deficiency can make your RBCs smaller. If you have an iron deficiency as well as vitamin B12 deficiency, each will cancel out the other’s effect on the size of the cell.
As more doctors are becoming aware that B12 deficiency is more common than initially thought, more B12 tests are ordered along with the FBC to interpret both results at the same time, giving a clear picture.
2. Serum B12 Test
In its guidelines, the BCSH states that
Definitive cut-off points to define clinical and subclinical deficiency states are not possible, given the variety of methodologies used and technical issues, and local reference ranges should be established.
The serum B12 test is not very accurate due to a couple of things. First, because threshold figures vary across labs, the threshold used to diagnose a deficiency may be too low. You can have patients who have symptoms of B12 deficiency, but whose levels are within ‘normal’ range. These patients should be treated anyway. There are also problems with some of the machines used to test for B12, and queries around the validity of their results.
3. Red Blood Cell Folate (RBC Folate) and Serum Folate
These are tests to measure folate level in the RBCs, on a smaller scale, as well as the blood, on a larger scale. Both are as effective in determining a patient’s folate level.
If a patient has been supplementing using folic acid tablets, then this increase in folic acid will prevent red blood cells from turning into megaloblasts. This means that high folate levels can hide one of the test results, which points to vitamin B12 deficiency.
Further, the lower a patient’s folate level, the more B12 will be excreted via urine. This is a rather unexpected effect of low folate level.
Another problem posed by low levels of folic acid is that patients are not able to convert the injectable vitamin B12 cyanocobalamin into the ‘active’ forms of B12, namely Methylcobalamin and Adenosylcobalamin.
Tell your doctor if you have been using a folic acid supplement before testing your vitamin B12 levels; the test may show normal or even high levels of B12 even if the ‘active’ form is not present—current testing does not distinguish between active and inactive forms of B12.
The BCSH has mentioned this in their guidelines as well:
Serum folate offers the equivalent diagnostic capability to red cell folate and is the first-line test of choice to assess folate status.
4. Ferritin (iron stores)
Ferritin is a blood protein that contains iron, and the test helps doctors understand how much iron is stored in the body. If ferritin is low, this indicates that you have an iron deficiency and that your body’s iron stores are depleted. This low iron can be a result of blood loss, lack of iron in the diet, or an inability to absorb iron from food. A doctor will search behind the reasons for a low ferritin level through history-taking and examination.
5. Serum Holotranscobalamin (HoloTC) – The ‘Active’ B12 Test
This is a test that distinguishes between the active form of B12 holotranscobalamin) and the inactive form (holohaptocorrin). It is not readily available in a lot of hospitals in the UK. The results it provides remain inconclusive, but in some cases, there are differences between the patient’s total serum B12 results and the HoloTC result. Therefore, while this test has the potential to remove some of the inaccuracies associated with current testing methods for vitamin B12, there does need to be more investigation into its effectiveness.
Here’s what the new guidelines say:
Serum holotranscobalamin has the potential as a first-line test, but an indeterminate ‘grey area’ may still exist.
6. Plasma Methylmalonic Acid (MMA)
This test is a good early indicator of B12 deficiency; the lower B12 levels, the higher MMA levels. However, it is not routinely carried out. High levels of MMA are sometimes also seen in patients who have kidney disease.
7. Plasma Total Homocysteine (tHcy)
Another test which can reliably point to low B12 levels early on, tHcy comes with a couple of problems. First, this test must be carried out within two hours of the blood being drawn from the patient provided that the sample is kept cool during this period. Second, tHcy is also increased in diseases such as kidney failure, hypothyroidism, vitamin B6 deficiency, or folate deficiency.
Tests for Pernicious Anaemia
After a patient is investigated and a vitamin B12 deficiency is discovered, there should be a further inquiry into why they have that deficiency. The following tests are those which can point to a diagnosis of pernicious anaemia (which causes vitamin B12 deficiency, among other things).
1. Anti-intrinsic Factor Antibody Test
Intrinsic factor is a protein produced by a type of specialized cells that line the stomach. During digestion, stomach acids release vitamin B12 from food and bind to intrinsic factor to form a complex. This complex is necessary for the absorption of vitamin B12 in the small intestine.
If positive, the patient is diagnosed with auto-immune pernicious anaemia. However, this test is often criticized by doctors for not being sensitive, and therefore not reliable.
From the BCSH guidelines:
…and the finding of a negative intrinsic factor antibody assay does not, therefore, rule out pernicious anaemia.” The guidelines also state that recent B12 injections compromise the test, but do not specify a timeframe in which injections are considered ‘recent.
2. Parietal Cell Antibodies
These antibodies are present in about 90% of pernicious anaemia patients, but may also be present in a variety of other conditions and in up to 10% of the general healthy population.
Other Tests
1. Mean Corpuscular Hemoglobin (MCH)
The MCH shows the average quantity of haemoglobin present in a single red blood cell. A high concentration is indicative of B12 deficiency.
2. Mean Corpuscular Hemoglobin Concentration (MCHC)
This measures the average concentration of haemoglobin in red blood cells.
3. Red Blood Cell Distribution Width (RDW)
A measure of the range of variation of red blood cell volumes. If RDW is high, it could indicate a B12 deficiency, an iron deficiency, or a folate deficiency.