Reticulocytes occupy an intermediate position between nucleated RBCs in the bone marrow and mature (nonnucleated, fully hemoglobinated) RBCs. After the normoblast (metarubricyte) nucleus is extruded from the cell, some cytoplasmic microsomes and ribosomes remain for 1-2 days that are not ordinarily visible on peripheral blood smears using Wright’s or Giemsa stain but that can be seen by using vital staining techniques and dyes such as methylene blue or cresyl blue. The material then is seen microscopically in the form of dark blue dots or thin short irregular linear structures arranged in loose aggregates or reticulum. The reticulocyte count is an index of the production of mature RBCs by the bone marrow. Increased reticulocyte counts mean an increased number of RBCs being put into the peripheral blood in response to some stimulus. In exceptionally great reticulocyte responses, there may even be nucleated RBCs pushed out into the peripheral blood due to massive RBC production activity of the bone marrow. Except in a very few diseases, such as erythroblastosis, peripheral blood nucleated RBCs are usually few in number and of a later maturity stage when they do appear. Reticulocytes are not completely mature RBCs; therefore, when reticulocytes appear in the peripheral blood, they may be slightly larger than normal RBCs and may be sufficiently large to be recognizable as macrocytes. When present in sufficient numbers, these macrocytes may increase the MCV index. Early reticulocytes sometimes appear blue-gray or gray with Wright’s stain in contrast to the red-orange appearance of the normal RBC; this phenomenon is called polychromatophilia and is produced by immature bluish cytoplasmic material to which reddish staining hemoglobin is added. In some conditions a reticulocyte may display small, evenly distributed dotlike aggregates of cytoplasmic ribosomes visible with Wright’s stain, a phenomenon known as basophilic stippling.

Reference limits for the reticulocyte count are usually considered to be 0.5%-1.5%. Some investigators have reported somewhat higher values, especially in women. There is a substantial problem concerning reproducibility of reticulocyte counts, with statistical variation on the order of ±1 reticulocyte percent unit at normal reticulocyte levels of 0.5%-1.5% standard manual (i.e., statistical variation of more than 50%) and somewhat greater variation at levels of 5% or more. More recently, it has become possible to count reticulocytes using a fluorescent nucleic acid stain in a flow cytometer. This method generally has statistical error less than 15%. Howell-Jolly bodies and medical parasites may interfere if large numbers are present. Some automated cell counters can be adapted to count reticulocytes in the red cell counting channel with preliminary reports suggesting accuracy comparable to flow cytometry. There may be differences in the reference range produced by different equipment and reports.

Reticulocytes are traditionally reported as a percentage of total RBCs (total RBC includes both mature RBCs and reticulocytes). Automated meter methods are frequently reported as the absolute (quantitative) number of retics. In some clinical situations the number of mature RBCs may decrease while the absolute (total) number of reticulocytes remains the same; this increases the reticulocyte percentage and gives a false impression of increased reticulocyte production. Therefore, some authorities recommend that reticulocyte counts be corrected for effects of anemia. This may be done by multiplying the reticulocyte count (percent) by the patient hematocrit and dividing the result by an average normal hematocrit (47 for men and 42 for women). An alternate method is to obtain the absolute number of circulating reticulocytes by multiplying the patient RBC count by the reticulocyte count (after converting reticulocyte % to a decimal fraction). If polychromatophilic RBCs are present, some experts recommend that the (already) corrected reticulocyte count be divided by 2 to correct for the longer stay of younger reticulocytes in the peripheral blood.

As noted previously, reticulocyte counts are used as an index of bone marrow activity. Any substantial change in bone marrow RBC production theoretically should be reflected in reticulocyte count change. A normal reticulocyte count has traditionally been considered evidence against a substantial degree of hemolytic anemia and can be used as an index of success in therapy for factor-deficiency anemia. One difficulty is that it usually takes 48-96 hours, sometimes even longer, to establish a reticulocyte count elevation following acute episodes of blood loss, onset of hemolysis, or beginning of factor therapy. Also, reticulocyte counts are not above reference range in some patients with hemolytic anemia (as many as 20%-25% in some studies, depending on the etiology of the disorder), with the degree of reticulocyte response having some correlation with the severity of the hemolytic process. In some cases, failure to obtain expected degrees of reticulocyte response by be due to superimposed factor deficiency (e.g., iron or folate). Another problem may be failure to suspect a hemolytic process or blood loss because the hemoglobin level may remain within population reference range if increased RBC production balances RBC loss or destruction.

In certain hemolytic anemias such as sickle cell anemia and congenital spherocytosis, temporary aplastic “crisis” may develop in which the anemia worsens because of a halt in RBC production rather than an increase in rate of hemolysis. These crises are sometimes due to parvovirus B-19 infection. The reticulocyte count will become normal or decrease after time is allowed for reticulocytes already in the peripheral blood to disappear.

In certain anemias caused by ineffective erythropoiesis, the reticulocyte count is normal or decreased unless therapy is given. Some examples include deficiencies of iron, folic acid, vitamin B12, or pyridoxine, or in many patients with anemia associated with chronic disease. In the case of factor deficiency, blood transfusion or hospital diet may contain a sufficient quantity of the deficient factor to increase RBC production.