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List the classifications of Hemolytic Disease of the Newborn and the most antibody specificities involved.
State the testing to perform on the mother to monitor the severity of HDN.
List the laboratory tests and values which indicate that an infant is affected by HDN both in the fetus and newborn.
State the treatment options for intrauterine treatment of HDN.
State the treatment options for HDN in the moderately and severely affected newborn.
State the requirements of blood to be used for transfusion of the fetus and newborn.

 Maternal IgG antibodies directed against an antigen of paternal origin present on the fetal red blood cells.
 IgG antibodies cross the placenta to coat fetal antigens, cause decreased red blood cell survival which can result in anemia.
 Produced in response to previous pregnancy with antigen positive fetus OR exposure to red blood cells, ie transfusion.

 ABO
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“Other” – unexpected immune antibodies other than anti-D – Jk, K, Fy, S, etc.
 Rh – anti-D alone or may be accompanied by other Rh antibodies – anti-C, -c, -E or – e.

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Mother group O, baby A or B
Group O individuals have anti-A, -B and –A,B in their plasma, fetal RBCs attacked by 2 antibodies
 Occurs in only 3%, is severe in only 1%, and
<1:1,000 require exchange transfusion.
 The disease is more common and more severe in African-American infants.

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Uncommon, occurs in ~0.8% of pregnant women.
Immune alloantibodies usually due to anti-E, -c,
-Kell, -Kidd or -Duffy.
Anti-K
 disease ranges from mild to severe
 over half of the cases are caused by multiple blood transfusions
 is the second most common form of severe HDN
Anti-M very rare

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 Anti-D is the commonest form of severe HDN.
The disease varies from mild to severe.
Anti-E is a mild disease
Anti-c can range from a mild to severe disease is the third most common form of severe HDN
Anti-e - rare
Anti-C - rare antibody combinations (ie anti-c and anti-E antibodies occurring together) - can be severe

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Maternal antibodies destroy fetal red blood cells
 Results in anemia.
 Anemia limits the ability of the blood to carry oxygen to the baby's organs and tissues.
Baby's responds to the hemolysis by trying to make more red blood cells very quickly in the bone marrow and the liver and spleen.
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Organs enlarge - hepatosplenomegaly.
New red blood cells released prematurely from bone marrow and are unable to do the work of mature red blood cells.
As the red blood cells break down, bilirubin is formed.
 Babies unable to get rid of the bilirubin.
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Builds up in the blood (hyperbilirubinemia ) and other tissues and fluids of the baby's body resulting in jaundice.
The placenta helps get rid of some of the bilirubin, but not all.

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Severe anemia with enlargement of the liver and spleen
When these organs and the bone marrow cannot compensate for the fast destruction of red blood cells, severe anemia results and other organs are affected.
Hydrops Fetalis
This occurs as the baby's organs are unable to handle the anemia. The heart begins to fail and large amounts of fluid build up in the baby's tissues and organs. A fetus with hydrops is at great risk of being stillborn.

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Varies from mild jaundice and anemia to hydrops fetalis (with ascites, pleural and pericardial effusions)
Chief risk to the fetus is anemia .
Extramedullary hematopoiesis due to anemia results in hepatosplenomegaly .
Risks during labor and delivery include:
 asphyxia and splenic rupture .
Postnatal problems include:
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Asphyxia
Pulmonary hypertension
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Pallor (due to anemia)
Edema (hydrops, due to low serum albumin)
Respiratory distress
Coagulopathies (↓ platelets & clotting factors)
Jaundice
Kernicterus (from hyperbilirubinemia)
Hypoglycemia (due to hyperinsulinemnia from islet cell hyperplasia)

 Kernicterus (bilirubin encephalopathy) results from high levels of indirect bilirubin
( >20 mg/dL in a term infant with HDN ).
 Kernicterus occurs at lower levels of bilirubin in the presence of acidosis, hypoalbuminemia, prematurity and certain drugs ( e.g.
, sulfonamides) .

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Affected structures have a bright yellow color.
Unbound unconjugated bilirubin crosses the blood-brain barrier and, because it is lipid soluble, it penetrates neuronal and glial membranes.
Bilirubin is thought to be toxic to nerve cells
The mechanism of neurotoxicity and the reason for the topography of the lesions are not known.
Patients surviving kernicterus have severe permanent neurologic symptoms
(choreoathetosis, spasticity, muscular rigidity, ataxia, deafness, mental retardation) .

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Vary with severity of HDN and include:
Anemia
Hyperbilirubinemia
Reticulocytosis (6 to 40%)
↑ nucleated RBC count (>10/100 WBCs)
Thrombocytopenia
Leukopenia
Positive Direct Antiglobulin Test
Hypoalbuminemia
Rh negative blood type or ABO incompatibility
Smear: polychromasia, anisocytosis, no spherocytes

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Polychromasia
Anisocytosis
Increase NRBCs no spherocytes

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Total Serum Bilirubin (TSB) monitored to determine risk of kernicterus.
Measure bilirubin in cord blood and at least every 4 hours for the first 12 to 24 hours. Plot bilirubin concentrations over time.

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 Transcutaneous bilirubinometry can be adopted as the first-line screening tool for jaundice in well, full-term babies.
This leads to about 50% decrease in blood testing.
http://tinyurl.com/36jazx

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Given to the fetus to prevent hydrops fetalis and fetal death.
Can be done as early as 17 weeks, although preferable to wait until
20 weeks
Severely affected fetus, transfusions done every 1 to 4 weeks until the fetus is mature enough to be delivered safely. Amniocentesis may be done to determine the maturity of the fetus's lungs before delivery is scheduled.
After multiple IUTs, most of the baby’s blood will be D negative donor blood, therefore, the Direct Antiglobulin test will be negative, but the Indirect Antiglobulin Test will be positive.
After IUTs, the cord bilirubin is not an accurate indicator of rate of hemolysis or of the likelihood of the need for post-natal exchange transfusion.

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An intrauterine fetal blood transfusion is done in the hospital. The mother may have to stay overnight after the procedure.
The mother is sedated, and an ultrasound image is obtained to determine the position of the fetus and placenta.
After the mother's abdomen is cleaned with an antiseptic solution, she is given a local anesthetic injection to numb the abdominal area where the transfusion needle will be inserted.
Medication may be given to the fetus to temporarily stop fetal movement.
Ultrasound is used to guide the needle through the mother's abdomen into the fetus's abdomen or an umbilical cord vein.
A compatible blood type (usually type O, Rh-negative) is delivered into the fetus's abdominal cavity or into an umbilical cord blood vessel.
The mother is usually given antibiotics to prevent infection. She may also be given tocolytic medication to prevent labor from beginning, though this is unusual.

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Increasingly common and relatively safe procedure since the development of high resolution ultrasound particularly with colour Doppler capability.
MCA Doppler velocity as a reliable non-invasive screening tool to detect fetal anemia.
 The vessel can be easily visualized with color flow Doppler as early as 18 weeks’ gestation.
 In cases of fetal anemia, an increase in the fetal cardiac output and a decrease in blood viscosity contribute to an increased blood flow velocity

 The risk of these procedures is now largely dependent on the prior condition of the fetus and the gestational age at which transfusion is commenced.

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Titer greater than 32 for anti-D and 8 for anti-K OR four fold increase in titer indicates need for analysis of amniotic fluid.
Amniocentesis
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Perform at 28 wks if HDN in previous child
Perform at 22 wks if previous child severely affected
Perform if maternal antibody increases before 34 th wk.
High values of bilirubin in amniotic fluid analyses by the Liley method or a hemoglobin concentration of cord blood below 10.0 g/mL.
Type fetus -recent development in fetal RhD typing involves the isolation of free fetal DNA in maternal serum. In the United Kingdom, this technique has virtually replaced amniocentesis for fetal RhD determination in the case of a heterozygous paternal phenotype
Maternal plasma exchange may be instituted if the fetus is too young for intrauterine transfusion.

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CPD, as fresh as possible, preferably <5 days old.
A hematocrit of 80% or greater is desirable to minimize the chance of volume overload in the fetus.
The volume transfused ranges from 75-175 mL depending on the fetal size and age.
CMV negative
Hemoglobin S negative
IRRADIATED
O negative, lack all antigens to which mom has antibodies and Coomb’s compatible.

 Phototherapy is the treatment of choice.
 Phototherapy process slowly decomposes/converts bilirubin into a nontoxic isomer, photobilirubin, which is transported in the plasma to the liver.
 HDN is judged to be clinically significant
(phototherapy treatment) if the peak bilirubin level reaches 12 mg/dL or more.

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The therapy uses a blue light (420-470 nm) that converts bilirubin so that it can be excreted in the urine and feces.
Soft eye shields are placed on the baby to protect their eyes from damage that may lead to retinopathy due to the bili lights.

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Lightweight, fiberoptic pad delivers up to 45 microwatts of therapeutic light for the treatment of jaundice while allowing the infant to be swaddled, held and cared for by parents and hospital staff.
Compact unit is ideal for hospital and homecare.

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Full-term infants rarely require an exchange transfusion if intense phototherapy is initiated in a timely manner.
It should be considered if the total serum bilirubin level is approaching 20 mg/dL and continues to rise despite intense inhospital phototherapy.
The procedure carries a mortality rate of approximately 1% and there may be substantial morbidity

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Remove sensitized cells.
Reduce level of maternal antibody.
Removes about 60 percent of bilirubin from the plasma, resulting in a clearance of about 30 percent to 40 percent of the total bilirubin.
Correct anemia by providing blood that will have normal survival.
Replacement with donor plasma restores albumin and any needed coagulation factors.
Rebound – usually a 2 volume exchange is needed as bilirubin in tissues will return to blood stream.

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Antibody elution testing from cord red blood cells.
ABO/D typing
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If baby received intrauterine transfusions will type as O negative
If baby’s Direct Antiglobulin Test is strongly positive due to anti-D may get FALSE NEGATIVE immediate spin reaction with reagent anti-D (blocking phenomenon), weak D (Du) test will be
STRONGLY positive
Antibody screen
Coomb’s crossmatch antigen negative donor.

 Type and screen on mom.
 Identification of unexpected antibodies.
 More than 40 antigens have been identified as causing HDN.
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Select blood that lacks antigens to which mom has antibodies.
Perform coomb’s crossmatch with Mom and baby’s blood.

 CPD, as fresh as possible, preferably <5 days old.
 CMV negative
 Hemoglobin S negative
 Irradiated if possible

 Physician will specify a hematocrit.
 Reconstitute donor unit with plasma.
 Most facilities prefer to use group O red cells and AB plasma.
 Reference for procedure at end of this presentation.

 Three types of HDN vary in severity.
 Laboratory testing key to diagnosing and monitoring- great care to be taken when interpreting ABO/D typing on affected infants.
 Therapy dependent on severity: phototherapy alone or with transfusion.

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References
Urgent Clinical Need for Accurate and Precise Bilirubin Measurements in the United States to Prevent
Kernicterus, 2004 http://www.clinchem.org/cgi/content/full/50/3/477
Identifying newborns at risk of significant hyperbilirubinaemia: a comparison of two recommended approaches, http://adc.bmj.com/cgi/content/full/90/4/415
Predictive Ability of a Predischarge Hour-specific Serum Bilirubin for Subsequent Significant
Hyperbilirubinemia in Healthy Term and Near-term Newborns, 1999, http://pediatrics.aappublications.org/cgi/content/full/103/1/6
Neonatal Jaundice, 2005, http://int-pediatrics.org/PDF/volume_20/20_1/47_54_ip20_1.pdf
HDN, Answers.com, http://www.answers.com/topic/hemolytic-disease-of-the-newborn
Hemolytic Disease of the Newborn, UCSF Children’s Hospital, http://www.ucsfhealth.org/childrens/health_professionals/manuals/42_Hemol.pdf
Lucile Packard Children’s Hospital at Stanford, http://www.lpch.org/DiseaseHealthInfo/HealthLibrary/hrnewborn/hdn.html
Intrauterine Transfusion, http://www.sufw.com.au/default.asp?cid=1%7C6%7C1
The Procedure for Preparation of Whole Blood for Neonatal Exchange Transfusion, http://www.cbbsweb.org/enf/attachments/sop_reconstitute_blood_mass.pdf
California Blood Bank Society discussion on neonatal transfusions http://www.cbbsweb.org/enf/2002/txneonatalpract.html
Irradiation of Blood Products, recommendations, http://www.scbcinfo.org/publications/bulletin_v2_n1.htm
Intrauterine Tranfusion: IOI Consecutive Cases treated at Queen Charlotte's Maternity Hospital, http://www.pubmedcentral.nih.gov/picrender.fcgi?artid=1587950&blobtype=pdf
Management of Rhesus Alloimmunization in Pregnancy, 2005, http://www.touchbriefings.com/pdf/1421/ACFA2C2.pdf
Molecular Diagnosis in Prenatal Medicine, http://pages.unibas.ch/diss/2004/DabsB_7129.pdf
Rh phenotype prediction by DNA typing and its application to practice, http://www.uniulm.de/~wflegel/RH/TME/trans173.pdf
Many references to molecular blood group testing http://www.ihop-net.org/UniPub/iHOP/bng/91718.html
Created April 2007