Laboratory Diagnosis Of Megaloblastic Anemias

The megaloblastic anemias show striking similarities in their clinical and hematological presentations. Common features of the megaloblastic anemias include

• Pancytopenia

• Increased MCV and MCHC

• Hypersegmented neutrophils (five lobes or more in segmented neutrophils)

• Increased bilirubin

• Increased LDH

• Hyperplasia in the bone marrow

• Reticulocytopenia

The differential diagnosis of these disorders depends on a more sophisticated battery of laboratory tests that can help determine if the patient is lacking vitamin B12, folic acid, or intrinsic factor. Several tests are used to distinguish between these possibilities; they include serum B12, folic acid, or red cell folate determination by radioimmunoassay, gastric analysis to determine achlorhydria or lack of hydrochloric acid in the stomach, or tests to denote intrinsic factor or parietal cell antibodies performed by enzyme-linked immunosorbent assay (ELISA). Parietal cell antibodies are seen in 90% of individuals at the time of initial diagnosis,8 although the presence of these antibodies is not specific for a diagnosis of pernicious anemia, because parietal cell antibodies are seen in some individuals with endocrine disorders. Intrinsic factor antibody evaluations are cost effective, reliable, and highly specific for a diagnosis of pernicious anemia.9 There are two classifications of intrinsic antibody: blocking antibodies and binding antibodies. Blocking antibodies inhibit the binding of vitamin B12 to intrinsic factor, while binding antibodies prevent the attachment of the intrinsic fac-tor-B12 complex to receptors in the small intestine. Radioimmunoassay testing can delineate the nature of the intrinsic factor antibody. The reference procedure for the determination of pernicious anemia, however, continues to be the Schilling test, which measures the intestinal absorption of vitamin B12. This test is performed in two parts, and although it is costly and labor intensive, it provides valuable information on the cause of pernicious anemia. The procedure in part 1 is to give the patient an oral dose of vitamin B12 and then, within 2 hours, a flushing dose of vitamin B12 via intramuscular injection. The flushing dose saturates all of the liver B12 binding sites. The urine is collected in a 24-hour period and the amount of vitamin B12 is measured. If intrinsic factor was present and vitamin B12 was absorbed, then 5% to 30% of the initial radiolabeled B12 will be excreted. If less than this amount is excreted, some type of malabsorption has taken place. In part 2 of the test, intrinsic factor is added to the oral vitamin B12 dose and the test proceeds as in part 1, including the flushing dose of B12. If the excretion of B12 is in the proper amount, then intrinsic factor is determined as the deficiency. If the excretion of B12 is less than expected, then the patient is diagnosed with a malabsorption syndrome. Normal kidney function and conscientious urine collection are essential for the correct interpretation of this test. Once the results are analyzed, the physician will have a clear picture as to whether the lack of vitamin B12 absorption is due to intrinsic factor deficiency or malabsorption syndrome (Fig. 6.5).

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