Transferrin is a transport protein that contains two iron binding sites by which it transports iron from storage sites to erythroid precursors.[5] TSAT (ie, the percentage of total binding sites that are occupied by iron) is a measure of iron that is available for erythropoiesis. TSAT is calculated by dividing the serum iron by the total iron binding capacity (TIBC), a measurement of circulating transferrin, and multiplying by 100.[5]

Ferritin is a storage protein that is contained primarily within the reticuloendothelial system (RES), with some amounts released in the serum. Under conditions of iron excess, ferritin production increases to offset the increase in plasma iron.[11] The level of ferritin in the serum, therefore, reflects the amount of iron in storage.

In normal patients, SF levels range from 13 to 220 ng/mL and TSAT levels range from 20% to 40%.[11] In patients without renal impairment, SF levels <12 ng/mL[12] and TSAT <16%[13] are indicative of depleted iron stores and absolute iron deficiency. In patients with chronic kidney disease, absolute iron deficiency is characterized by SF levels < 100 ng/mL and TSAT <20%.[5]

Because patients on epoetin therapy may have adequate iron stores (as reflected by serum ferritin levels ≥100 ng/mL) but still have functional iron deficiency, the use of alternative iron parameters has been suggested in these patients.

Reticulocyte hemoglobin content (CHr). Reticulocytes are immature red blood cells (RBCs) with a life span of only 1 to 2 days. When these are first released from the bone marrow, measurement of their hemoglobin content can give a picture of the amount of iron immediately available for erythropoiesis.[11] A less than normal hemoglobin content in these reticulocytes is an indication of inadequate iron supply relative to demand. The amount of hemoglobin in these reticulocytes also corresponds to the amount of hemoglobin in mature RBCs. CHr has been evaluated in several studies as a test for functional iron deficiency and has been found to be highly sensitive and specific. However, exact threshold values have not been established. Threshold values vary depending on the laboratory and assay used.[14,15,16]

Percentage hypochromic red blood cells. Epoetin is effective in stimulating production of RBCs, but without an adequate iron supply to bind to heme, the RBCs will be hypochromic, that is, have a low hemoglobin content. Thus, in states of iron deficiency, a significant percentage of RBCs leaving the bone marrow will have a low hemoglobin content. By measuring the percentage of RBCs with hemoglobin content <28 g/dL, iron deficiency can be detected. Hypochromic RBC percentages >10% have been correlated with iron deficiency.[11]

Serum transferrin receptors (sTfR). Transferrin receptors on the cell surface of RBC precursors bind iron-bound transferrin, allowing the transport of iron from the plasma into the cells. Under conditions of iron deficiency, there is an upregulation of these receptors to allow more efficient uptake of transferrin.[11] The concentration of transferrin receptors on the cell surface correlates with transferrin uptake. In hemodialysis patients who are not treated with epoetin, sTfR levels are higher among those who are iron deficient than among those who are iron replete.[17] However, in several studies, hemodialysis patients treated with epoetin had similar sTfR levels regardless of iron status.[18,19] sTfR may not be an accurate marker of iron status in hemodialysis patients.