Iron Deficiency Anemia (IDA) – Diagnosis, Screening Tests, Investigations
Last Updated : 5/3/2010
M R Lokeshwar*, Nitin Shah**
Investigations that are required to establish the diagnosis of IDA and to determine its underlying cause are:

Screening Tests:
Red cell count, hemoglobin and Hematocrit are all decreased in IDA. MCV, MCH, and MCHC are also decreased. The peripheral blood film shows hypochromic, microcytic red cells. If anemia is severe, other morphological abnormalities such as poikilocytosis and target cells may be seen. When iron deficiency is associated with deficiency of other hematinics like vit. B12 or Folate, there may be a dimorphic picture with Hypochromic, microcytic red cells along with macrocytosis. These routine investigations may not be useful to diagnose early iron deficiency state. Reticulocyte count is normal unless the patient has had a recent acute blood loss or the patient has received hematinics, in which case it may be increased. In severe IDA, reticulocyte count may be decreased (Table 2).

Red cell osmotic fragility is decreased, and this along with low MCV may cause some confusion with beta-thalassemia trait, which causes a similar blood picture and is common in India. NESTROFT (Naked Eye Single Tube Osmotic Fragility Test), a useful screening test for thalassemia trait is positive in adult 15% of cases of I.D.A. However, if nestroft is negative, it is unlikely to be thalassemia trait.

Leucocyte count is usually normal. Hypersegmented neutrophils may be seen due to concomitant B12 or folate deficiency or due to iron deficiency - induced interference with folate utilization, or due to IDA - induced impairment of jejunal function leading to poor absorption of B12 or folate. In a majority of such cases, the abnormality is corrected by iron therapy without B12 or folate.

Thrombocytosis may occur in patients with IDA, as a result of iron deficiency per se or due to underlying condition such as malignancy or bleeding.

Red cell indices: With the availability of electronic particle counters estimation of PVC, MCV, MCH, RBC count has become accurate, reproducible and practical laboratory test for screening anemic patients. Manual determination of these red cell indices is time consuming and poorly reproducible. Low MCV and MCH with anemia favor the diagnosis of IDA. However, specific reference standard must be used for comparison. Klee G in his study showed that more than half of the 62 patients with IDA had a MCH value clearly within a normal range and nearly 70% of cases exhibited distinct microcytosis, suggesting that MCV is much more sensitive than MCH in determining changes of iron deficiency. However, 30% of cases of IDA will be misdiagnosed if physician relies only on these indices. Red cell indices in IDA are generally low i.e. MCV < 80 cu mm, MCH < 27 pg and MCHC < 33%. Microcytosis also may be seen in other conditions like abnormal hemoglobinopathies, anemia of chronic infection and inflammation, lead poisoning and in rare conditions like sideroblastic anemia, chronic renal diseases, etc. Red blood cell survival is reduce in iron deficiency due to reduced cellular deformability resulting from a reduced red cell glutathione peroxidase activity.

Red cell size Distribution Width (RDW): An estimation on electronic counter reveals elevated value of red cell distribution width (RDW) in iron deficiency anemia as compared to normal levels in anemia of chronic diseases or beta - Thalassemia trait. RDW gives an objective evidence of degree of anisocytosis.

Confirmatory Tests for IDA: As shown in Table 6, Serum iron is reduced (Normal- 50 - 180 mcg/dl), TIBC is increased (Normal- 250 - 450 mcg/dl), Transferrin saturation is low (less than 16% suggestive and less than 7% diagnostic of severe iron deficiency anemia), Serum Ferritin is less than 10 - 12 ng/ml. However, when infectious or inflammatory diseases like rheumatoid arthritis, collagen disorders, liver disorders, chronic renal disease or malignancy are also present, the serum ferritin level is usually higher, but less than 50-60ng/ml. The test still lacks sensitivity and normal value does not reliably exclude iron deficiency.

Age in yearsSerum Ferritin (ng/dl)Transferrin Saturation %RBC FEP mcg/dl
0.5 - 4< 10< 12< 80
5 - 10< 10< 14> 70
11 - 14< 10< 16> 70
> 15< 12< 16> 70

Free Erythrocyte Protoporphyrin (FEP): Protoporphyrin accumulates in the red blood cells when it does not have sufficient iron to combine with, to form Hb. The FEP can be measured rapidly by a simple fluorescence assay performed directly on the thin film of the blood. FEP value in the normal person is 15.5 + 8.3 mcg/dl of RBC. More than 80 mcg/dl of RBC below the age of 4 years, and more than 70 mcg/dl of RBC above that age , are significant values to detect IDA. FEP/Hb ratio is a useful index of iron deficiency. FEB/Hb ratio increases when the iron reserve is exhausted, even before anemia becomes apparent. The ratio is normal in thalassemia trait and renal anemia. FEB/Hb ratio remains elevated during iron therapy and returns to normal only after majority of the cells containing FEP formed during iron deficiency are replaced. FEP/Hb ratio is not subject to daily fluctuation and sudden changes as is transferrin saturation. The highest value of FEP are seen in lead intoxication - a level of FEP greater than, 160mcg/dl of RBC is taken as a cut-off value for the detection of lead intoxication. In IDA, FEP/Hb ratio is moderately elevated and never exceeds 17.5mcg/gm of Hb (equivalent to 500mcg/dl of RBC). An FEP/Hb ratio in the range of 5.5 -17.5 mcg of Hb may be attributed to either IDA or lead intoxication whereas FEP/Hb ratio greater than 17.5mcg/gm of Hb indicates lead intoxication with or without associated iron deficiency and requires immediate medical attention. Higher values may also be associated with rare genetic disorder like erythropoietic protoporphyria.

Bone marrow examination: Bone marrow aspiration is not routinely indicated in the diagnosis of IDA. The degree of cellularity and the proportion of myeloid to the erythroid cells on bone marrow examination vary depending on the severity as well as the duration of IDA. The bone marrow aspirate shows an erythroid normoblastic hyperplasia. Erythroblasts may be smaller than the normal with a narrow ragged rim of cytoplasm containing little Hb. However, morphological changes are not distinctive enough to be of diagnostic values. Prussian blue staining of iron particles revealing little or no stainable iron in reticulum cells and normoblast is diagnostic (iron granules are normally found in cytoplasm of 10% or more erythroblasts). High iron may be seen in the above marrow of the patient who have been recently transfused or received parenteral iron.


Contributor Information and Disclosures

M R Lokeshwar*, Nitin Shah**
*Visiting Pediatrician - P.D.Hinduja National Hospital, Mumbai Pediatrician and Hematologist-Oncologist,Lilavati Hospital, Bandra, Mumbai. **Division of Pediatric Hematology-Oncology, Dept of Pediatrics, LTMG Hospital & LTM Medical College, Sion, Mumbai - 400 022.

First Created : 2/8/2001
Last Updated : 5/3/2010


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