Williams Manual of Pregnancy Complications, 23 ed.

CHAPTER 91. Rh Disease and Other Isoimmunization

Isoimmunization of the mother is responsible for most cases of hemolytic disease of the newborn. This occurs when mothers who lack a specific red cell antigen are exposed to that antigen through blood transfusion or to a fetus during pregnancy. The most common causes of hemolytic diseases in the fetus–neonate are ABO incompatibility and sensitization to the Rh system. Less frequent causes are other blood group incompatibilities such as those due to Kell, Kidd, and Duffy antigens (Table 91-1). The reader is referred to Chapter 29 of Williams Obstetrics, 23rd ed., for more information on these “irregular” antigens.

TABLE 91-1. Atypical Erythrocyte Antibodies and Their Relationship to Fetal Hemolytic Disease





Although incompatibility for the major blood group antigens A and B is the most common cause of hemolytic disease in the newborn, the resulting anemia is usually mild. Approximately 20 percent of all infants have an ABO maternal blood group incompatibility, but only 5 percent are clinically affected. ABO incompatibility is different from CDE incompatibility for several reasons:

• ABO disease frequently is seen in firstborn infants. This is because most group O women have anti-A and anti-B isoagglutinins antedating pregnancy. These are attributed to exposure to bacteria displaying similar antigens.

• Most species of anti-A and anti-B antibodies are immunoglobulin M (IgM), which cannot cross the placenta and therefore cannot reach fetal erythrocytes. In addition, fetal red cells have fewer A and B antigenic sites than adult cells and are thus less immunogenic. Thus, there is no need to monitor for fetal hemolysis, and there is no justification for early delivery.

• The disease is invariably milder than D-isoimmunization and rarely results in significant anemia. Affected infants typically do not have erythroblastosis fetalis, but rather have neonatal anemia and jaundice, which can be treated with phototherapy.

• ABO isoimmunization can affect future pregnancies but unlike CDE disease, rarely becomes progressively more severe. Katz and coworkers (1982) identified a recurrence in 87 percent. Of these, 62 percent required treatment, most often limited to neonatal phototherapy.

Because of these reasons, ABO isoimmunization is a disease of pediatric rather than obstetrical concern. Although there is no need for antenatal monitoring, careful neonatal observation is essential because hyperbilirubinemia may require treatment. Treatment usually consists of phototherapy or simple or exchange transfusion with O-negative blood.


This system includes five red cell proteins or antigens: C, c, D, E, and e. No “d” antigen has been identified, and Rh- or D-negativity is defined as the absence of the D-antigen. There are, however, D-antigen variants that cause hemolytic disease. Some of these include weak D, Du, and partial D.

The CDE antigens are of considerable clinical importance because many D-negative individuals become isoimmunized after a single exposure. The two responsible genes—D and CE—are located on the short arm of chromosome 1 and are inherited together, independent of other blood group genes. Like many genes, their incidence varies according to racial origin. Native Americans, Inuits, and Chinese and other Asiatic peoples have 99-percent D positivity. Approximately 93 percent of African Americans are D-positive, but only 87 percent of Caucasians are. Of all racial and ethnic groups studied thus far, the Basques show the highest incidence of D-negativity at 34 percent.

The C-, c-, E-, and e-antigens have lower immunogenicity than the D-antigen, but they too can cause erythroblastosis fetalis. All pregnant women should be tested routinely for D-antigen erythrocytes and for irregular antibodies in their serum.


Because routine administration of anti-D immunoglobulin prevents most cases of anti–D-isoimmunization, proportionately more cases of significant antenatal hemolytic disease are now caused by the less common red cell antigens. Such sensitization is suggested by a positive indirect Coombs test performed to screen for abnormal antibodies in maternal serum (Table 91-1).

Several large studies indicate that anti–red cell antibodies are found in 1 percent of pregnancies. From 40 to 60 percent of these are directed against the CDE antibodies. Anti-D is the most common, followed by anti-E, anti-c, and anti-C. A third of fetuses with either anti-C or anti-Ce alloimmunization had hemolysis but none had severe disease. In contrast, 12 of 46 anti-c isoimmunized fetuses had serious hemolysis, and 8 of these 12 required transfusions.

Anti-Kell antibodies are also frequent. A fourth of all antibodies found are from the Lewis system. These do not cause hemolysis because Lewis antigens do not develop on fetal erythrocytes and are not expressed until a few weeks after birth.

Kell Antigen

Approximately 90 percent of Caucasians are Kell negative. Kell type is not routinely determined, and 90 percent of cases of anti-Kell sensitization result from transfusion with Kell-positive blood. As with CDE antigens, Kell sensitization also can develop as the result of maternal–fetal incompatibility. Kell sensitization may be clinically more severe than D-sensitization because anti-Kell antibodies also attach to fetal bone marrow erythrocyte precursors, thus preventing a hemopoietic response to anemia. Thus, there usually is a more rapid and severe anemia than with anti–D-sensitization.

Because fewer erythrocytes are produced, there is less hemolysis and less amnionic fluid bilirubin. As a result, severe anemia may not be predicted by either the maternal anti-Kell titer or the level of amnionic fluid bilirubin. Some investigators recommend evaluation when the maternal anti-Kell titer is 1:8 or greater. In addition, some investigations suggest that the initial evaluation of the significant positive titer be accomplished by cordocentesis instead of amniocentesis, because fetal anemia from Kell sensitization is usually more severe than indicated by the amnionic fluid bilirubin level. Determination of fetal middle cerebral artery (MCA) velocity by Doppler obviates this, as discussed subsequently.

Other Antigens

Kidd (Jka), Duffy (Fya), c-, E-, and to a lesser extent C-antigens can all cause erythroblastosis as severe as that associated with sensitization to D-antigen (Table 29-5). Two Duffy antigens have been identified, Fya and Fyb, and some African Americans lack both. Fya is the most immunogenic. The Kidd system also has two antigens, Jka and Jkb, with the population distribution as follows: Jk (a+b–), 26 percent; Jk (a–b+), 24 percent; and Jk (a+b+), 50 percent. Most cases of isoimmunization to these antigens occur after blood transfusions.

If an IgG red cell antibody is detected and there is any doubt as to its significance, the clinician should err on the side of caution, and the pregnancy should be evaluated. As shown in Table 29-5, many rare or private antigenshave been associated with severe isoimmunization.


The pathological changes in the organs of the fetus and newborn infant vary with the severity of the hemolytic process due to D-isoimmunization. Excessive and prolonged hemolysis serves to stimulate marked erythroid hyperplasia of the bone marrow as well as large areas of extramedullary hematopoiesis, particularly in the spleen and liver, which may in turn cause hepatic dysfunction. There may be cardiac enlargement and pulmonary hemorrhages. When the severely affected fetus or infant shows considerable subcutaneous edema as well as effusion into the serous cavities, hydrops fetalis is diagnosed. It is defined as the presence of abnormal fluid in two or more sites such as thorax, abdomen, or skin. The diagnosis is usually made easily using sonography. The placenta is also markedly edematous, appreciably enlarged and boggy, with large, prominent cotyledons and edematous villi.

The precise pathophysiology of hydrops due to Rh disease remains obscure. Theories of its causation include heart failure from profound anemia, capillary leakage caused by hypoxia from severe anemia, portal and umbilical venous hypertension from hepatic parenchymal disruption by extramedullary hematopoiesis, and decreased colloid oncotic pressure from hypoproteinemia caused by liver dysfunction.

Fetuses with hydrops may die in utero from profound anemia and circulatory failure. A sign of severe anemia and impending death is a sinusoidal fetal heart rate (see Chapter 12). The liveborn hydropic infant appears pale, edematous, and limp at birth, often requiring resuscitation. The spleen and liver are enlarged, and there may be widespread ecchymosis or scattered petechiae. Dyspnea and circulatory collapse are common.


Less severely affected infants may appear well at birth, only to become jaundiced within a few hours. Marked hyperbilirubinemia, if untreated, may lead to kernicterus, a form of central nervous system damage that especially affects the basal ganglia. Anemia, in part resulting from impaired erythropoiesis, may persist for many weeks to months in the infant who had demonstrated hemolytic disease at birth.

Perinatal Mortality

Perinatal deaths from hemolytic disease caused by D-isoimmunization have decreased dramatically since adoption of the policy of routine preventative administration of D-immunoglobulin to all D-negative women during or immediately after pregnancy. Survival also has been increased by antenatal transfusions or preterm delivery of affected fetuses if necessary. The advent of fetal transfusion therapy has resulted in survival rates exceeding 90 percent for severe anemia alone, and 70 percent if hydrops has developed.


Blood typing and antibody screen is done at the first prenatal visit, and unbound antibodies in maternal serum are detected by the indirect Coombs test. If positive, specific antibodies are identified, and their immunoglobulin subtype is determined as either IgG or IgM. Only IgG antibodies are concerning because IgM antibodies cannot cross the placenta. If the IgG antibodies are known to cause fetal hemolytic anemia as shown in Table 91-1, the titer is quantified. The critical titer is the level for a particular antibody that requires further evaluation. This may be different for each antibody and is determined individually by each laboratory. For example, the critical titer for anti-D antibodies is usually 1:16. Thus, a titer of ≥1:16 indicates the possibility of severe hemolytic disease. The critical titers for other antibodies are often assumed to be 1:16 as well, although most laboratories have insufficient data to support this assumption. An important exception is Kell sensitization. The critical titer for anti-Kell antibody is 1:8 or even less.


Management is individualized and consists of maternal antibody titer surveillance, sonographic monitoring of the fetal middle cerebral artery peak systolic velocity, amnionic fluid bilirubin studies, or fetal blood sampling. Accurate pregnancy dating is critical. The gestational age at which fetal anemia developed in the last pregnancy is important because anemia tends to occur earlier and be sequentially more severe. In the first sensitized pregnancy, a positive antibody screen with a titer below the critical level should be followed with repeated titers at timely intervals, usually monthly. Once the critical titer has been met or exceeded, subsequent titers are not helpful, and further evaluation is required. If this is not the first sensitized pregnancy, then the pregnancy is considered to be at risk, and maternal antibody titers are unreliable.

Amnionic Fluid Spectral Analysis

Almost 50 years ago, Liley demonstrated the utility of amnionic fluid spectral analysis to measure the bilirubin concentration to estimate the severity of hemolysis, and thus to indirectly assess anemia. MCA Doppler flow is increasingly being used in conjunction with amniotic fluid analysis.

Because amnionic fluid bilirubin levels are low, the concentration is measured by a spectrophotometer and is demonstrable as a change in absorbance at 450 nm—this is referred to as ΔOD450. The likelihood of fetal anemia is determined by plotting the ΔOD450 value on a graph that is divided into several zones. The original Liley graph (not shown) is valid from 27 to 42 weeks and contains three zones. Zone 1 generally indicates a D-negative fetus or one with only mild disease. Zone 2 indicates that anemia is present. In lower zone 2, the anticipated hemoglobin level is 11.0 to 13.9 g/dL, whereas in upper zone 2, hemoglobin values range from 8.0 to 10.9 g/dL. Zone 3 indicates severe anemia with hemoglobin values below 8.0 g/dL (Liley, 1961).

Use of the “Liley curve” allowed decisions regarding management. At that time, the options were fetal intraperitoneal transfusions or preterm delivery. The Liley graph was subsequently modified by Queenan and associates (Figure 91-1). These investigators studied 845 amnionic fluid samples from 75 D-immunized and 520 unaffected pregnancies and constructed a Liley-type curve that begins at 14 weeks. As can be seen, the naturally high amnionic fluid bilirubin level at midpregnancy results in a large indeterminate zone. Importantly, bilirubin concentrations in this zone do not accurately predict fetal hemoglobin concentration. For this reason, when evaluation indicates that severe fetal anemia or hydrops before 25 weeks is likely, many forego amniocentesis in favor of fetal blood sampling.


FIGURE 91-1 Proposed amnionic fluid ΔOD450 management zones in pregnancies from 14 to 40 weeks. (Queenan JT, Thomas PT, Tomai TP, et al: Deviation in amniotic fluid optical density at a wavelength of 450 nm in Rh isoimmunized pregnancies from 14 to 40 weeks’ gestation. A proposal for clinical management. Am J Obstet Gynecol 168(5):1370–1376, 1993.)

Fetal Blood Sampling

Cordocentesis is used for obtaining fetal blood. There is a risk of fetal loss with this technique, but it allows both determination of fetal hemoglobin and transfusion if indicated.


The goal of management is delivery of a reasonably mature healthy fetus. When management includes serial amnionic fluid ΔOD450 measurement or fetal transfusions, fetal well-being should be closely monitored with techniques discussed in Chapters 9 and 12.

Exchange Transfusion in the Newborn

Some advocate that the last transfusion for the severely affected fetus be given at 30 to 32 weeks with antenatal corticosteroid administration and delivery at 32 to 34 weeks. Others continue transfusion until 36 weeks. In either case, cord blood is obtained at delivery for hemoglobin concentration and direct Coombs testing. If the infant is overtly anemic, it is often best to complete the initial exchange transfusion promptly with recently collected type-O, D-negative red cells. For infants who are not overtly anemic, the need for exchange transfusion is determined by the rate of increase in bilirubin concentration, the maturity of the infant, and the presence of other complications. Most fetal transfusion survivors develop normally.


Anti-D immunoglobulin is a 7S immune globulin G extracted by cold alcohol fractionation from plasma containing high-titer D antibody. Each dose provides not less than 300 μg of D antibody, which can neutralize 15 mL of fetal red blood cells. It is given to the D-negative nonsensitized mother to prevent sensitization to the D antigen.

Such globulin given to the previously unsensitized D-negative woman within 72 hours of delivery is highly protective. Any pregnancy-related events that could result in fetal–maternal hemorrhage require D-immunoglobulin and include miscarriage, abortion, or evacuation of a molar or ectopic pregnancy. In addition to these situations, anti-D-immunoglobulin is also given prophylactically to all D-negative women at about 28 weeks’ gestation.

In the case of a large fetal–maternal hemorrhage, one dose of D-immunoglobulin may not be sufficient to neutralize the transfused cells. The Kleihauer–Betke test can be used to estimate the amount of fetal blood in circulation, and D-immunoglobulin given accordingly.

For further reading in Williams Obstetrics, 23rd ed.,

see Chapter 29, “Diseases and Injuries of the Fetus and Newborn.”