| References Bersch C. ed. The ABCs of pre-, neo-, and post-natal testing. MLO. September 2009;41.Clinical and Laboratory Standards Institute (CLSI). Procedures and Devices for the Collection of Diagnostic Capillary Blood Specimens; Approved Standard. Fifth ed. CLSI document H4-A5. NCCLS. Wayne, PA: 2004.Clinical and Laboratory Standards Institute (CLSI). Procedures for the Handling and Processing of Blood Specimens; Approved Guideline. Third ed. CLSI document H18-A3. NCCLS.Wayne, PA: 2004.Ernst DJ. Applied Phlebotomy. Baltimore, MD: Lippincott Williams & Wilkins: 2005.Garza D. Becan-McBride K. Phlebotomy Handbook. 7th ed. Upper Saddle River, NJ: Pearson Prentice Hall: 2005.Stevens B, Yamada J, Ohlsson A. Sucrose for analgesia in newborn infants undergoing painful procedures. Available at http://www.nichd.nih.gov/cochrane/Stevens/Stevens.HTM accessed January 12, 2010. | View Page |
| Inappropriate Sites/Patients for Capillary Puncture There are some instances where a dermal puncture is prohibited or not recommended.Mastectomy patientsAs a general rule, a dermal puncture, or a venipuncture, should not be performed on the side affected by a mastectomy. The body's ability to fight infection is compromised if lymph nodes were removed. A physician's permission must be obtained before performing a blood collection procedure on the same side as a mastectomy. Edematous siteDermal punctures should not be performed on previously punctured sites or swollen sites. Excess tissue fluid may contaminate the specimen.Dehydrated patientIf the patient is dehydrated or has poor circulation, it may be impossible to get a quality specimen. Fingerstick on a newborn or young infant Dermal punctures must never be performed on the fingers of a newborn or very young infant (usually defined as under 12-months-old). There is very little distance between the skin and the bone. Therefore, the bone could be easily pierced during the puncture, causing injury to the bone, infection, or gangrene. | View Page |
| Infants and Geriatric Patients: Monitor the Amount of Blood Obtained The collection of a capillary blood specimen is often used on newborns and geriatric patients. These two groups are most susceptible to blood depletion. Therefore, a dermal puncture is preferred over venipuncture where too much blood may be inadvertently collected.In some facilities, the amount of blood obtained from a patient will be charted or recorded after every procedure. This may become part of the patient's medical record and is usually entered by the nursing staff. In these cases, the nurse will interact with the laboratory staff to advise them of the safe amount of blood that can be obtained. | View Page |
| Protect Me From the Light Some specimens routinely collected for testing by using a capillary puncture are adversely affected by exposure to light. One example is a specimen collected for bilirubin testing that is obtained from a newborn. When obtaining the specimen for this testing, it is important for the phlebotomist to recognize the effect of light on the specimen. Room light or sunlight can metabolize the bilirubin in the specimen to a different compound. This will cause a falsely lower bilirubin level. A neonatal bilirubin specimen should be obtained in a dark-colored (amber) container. Alternately, a clear or white container can be immediately wrapped in aluminum foil following the blood collection, preventing the blood from exposure to light. | View Page |
| The Need for Metabolic Testing on Newborns Many state governments in the United States mandate that all newborns be tested for metabolic disorders very soon after birth. This required testing is used to determine if the infant has a metabolic disorder that could adversely affect a child's development. If discovered early, many of the effects of the metabolic disorder can be alleviated or averted. Not every state tests or screens for the same disorders, so the phlebotomist must be certain to understand the requirements for the state in which they reside. There is a movement to standardize testing throughout the United States.Typically, the method used to screen for the presence of newborn metabolic disorders is collection of capillary blood on a filter paper card. It is imperative that the phlebotomist follows the very specific directions for the collection of these samples. If a specimen is submitted to the state for testing and deemed unacceptable, the specimen would have to be re-collected. The infant would then have to be subjected to a second invasive puncture procedure, causing stress and trauma to the infant as well as the parents. More importantly, the need to obtain a second specimen can also cause a delay in treatment. | View Page |
| Capillary Blood Collection for Metabolic Testing The collection of these specimens requires the same attention to detail as with any phlebotomy procedure. Gather all necessary equipment Be certain to choose a device that punctures the heel to a depth appropriate to the size of the infant. Only use the filter cards provided by your state to collect the specimen. These cards are calibrated to the exact specifications needed for testing of metabolic disorders. An alternate or homemade card must not be used. Put on all necessary personal protective equipment Gloves are always required. Gowns and eye protection may also be required. Positively identify the patient Use two identifiers. The infant who is in the nursery should have an identification band attached to the ankle or wrist. In special care nurseries an alternate form of identification may be used. However, a crib card should never be used as a form of identification. Follow the practice for your facility. Position the infant Be certain that the heel can be easily accessed. Follow all nursery requirements that apply to safe handling of newborns. Warm the heel using an approved warming device Clean the site with alcohol or the approved disinfectant. Allow the site to air dry before proceeding with collection of the specimen. Grasp the heel firmly but not tightly, activate the puncture device, wipe away the first drop of blood, and begin collection of the specimen.Allow the blood to wick onto the card. Completely saturate the circle with one continuous drop of blood. Avoid touching the card to the skin. Apply the blood only to one side of the card. Do not layer the blood by applying a second drop on top of the first. Repeat the procedure to completely fill each circle on the card. Each circle should be completely and uniformly saturated as shown in the bottom image on the right. Follow the policy of your institution or state to determine how many circles must be completely filled. Apply pressure to the puncture site using a sterile gauze Gently raising the infant's leg above the level of the heart will also aid in clotting the puncture site. Bandage according to site-specific policy. | View Page |
| The directions for this testing facility requires the filling of all 5 circles on the filter card.Which of the cards that were collected for metabolic testing on newborns is filled correctly? | View Page |
| Foreword This course is a refresher on current concepts and practices in hemolytic disease of the fetus and newborn (HDFN). As such it is a survey course that provides a broad overview of the field and presents an opportunity to review significant aspects of HDFN and its laboratory investigation and prevention. Because it is a survey course with many topics, not all will be covered in depth. However, Rh immune globulin (RhIg) will be reviewed extensively since it prevents the most severe form of HDFN and is one of the biggest success stories of modern medicine. The course assumes that participants have a basic background knowledge of immunohematology theory and practice. Reading the resources in Further Reading for more information on any topic is encouraged. In brief, the course will: Recap relevant background information on HDFN and its treatment Review the characteristics and uses of Rh immune globulin (RhIg) Discuss typical laboratory findings and their interpretations Examine current best practices in perinatal testing programsThe course is a companion to "Rh negative female with anti-D at delivery: A case study on dealing with the issues" and complements its content. | View Page |
| Symptoms and Laboratory Findings in Severe HDFN Due to Anti-D Anti-D causes the most severe HDFN. Symptoms and laboratory findings in HDFN due to anti-D typically include:1. Anemia: Cord Hb can be less than 10 g/dL (100 g/L) and as low as 3–5 g/dL (30–50 g/L).2. Jaundice (icterus gravis): Jaundice occurs after delivery, as fetal bilirubin is cleared by the mother during pregnancy. Extravascular fetal red cell destruction by maternal antibody produces high bilirubin levels. The newborn, who is unable to produce adequate amounts of the liver enzyme glucuronyl transferase, is unable to conjugate the bilirubin into its water-soluble, excretable form.3. Kernicterus: If indirect bilirubin levels reach approximately 20 mg/dL (340 mmol/L) the fat soluble unconjugated bilirubin deposits in the fat-rich brain cells causing brain cell damage. Cerebral palsy, deafness, mental retardation, and other serious disorders can result.4. Hydrops fetalis: Gross edema occurs in severely affected infants, and often results in stillbirth or death soon after birth. Liver failure and hypoproteinemia also play a role in this syndrome.5. Enlarged organs, e.g., liver, spleen and heart6. Laboratory findings include a positive direct antiglobulin test (DAT), low hemoglobin (as above), increased reticulocyte count, and increased nucleated red cells. | View Page |
| Postnatal Treatment: Exchange Transfusion Whenever possible, a hallmark of HDFN treatment is to induce labor as early as possible once lung maturity has been attained so that the newborn will be able to survive. Once the infant is born, the main treatment for severe HDFN due to anti-D (and other antibodies causing severe disease) is exchange transfusion. In exchange transfusions, up to 85–90% of the infant's blood can be exchanged with donor blood by a process of removing 5–20 mL of blood at a time, and injecting an equivalent amount until the exchange is complete. An exchange transfusion accomplishes the following: Removes bilirubin and thus helps prevent kernicterus; Removes sensitized red cells that have not been broken down yet; Removes circulating maternal antibody; Provides antigen-negative red cells that will not be destroyed by the maternal antibody, thus will survive and provide oxygen to the tissues. | View Page |
| Which symptom of HDFN is associated with low levels of glucuronyl transferase? | View Page |
| Other Postnatal Treatment Besides exchange transfusion, postnatal treatment of HDFN may include the following:RBC TransfusionMany infants who have received IUTs also require simple RBC transfusions in the first few weeks of life to treat ongoing hemolysis caused by persistent maternal antibody in the newborn's circulation.Phototherapy Phototherapy is used to treat jaundice in preterm infants without HDFN and in infants with mild HDFN. Intensive phototherapy has also been used to treat moderate and severe HDFN and decrease the need for exchange transfusion. The newborn is placed under a "blue light" which chemically alters the bilirubin in the surface capillaries to a harmless substance. Human Serum AlbuminHuman serum albumin can also be transfused, either separately or as part of an exchange transfusion in place of FFP. Albumin binds unconjugated bilirubin, thus preventing its deposition in the fat-rich brain cells. Albumin must be used judiciously, because it can aggravate congestive heart failure. | View Page |
| ABO HDFN - Etiology and Symptoms ABO HDFN is the most common type of HDFN, in that anti-A is the antibody most often found bound to the red cells of a newborn. While the disease is usually so mild as to not require treatment, severe HDFN is possible. EtiologyABO HDFN is caused by maternal IgG anti-A or anti-B, which can be produced as a result of prior pregnancy or prior inoculation (some common inoculations contain A or B substances). In Caucasians, most often the mother is group O and the child is group A, although other combinations are possible. Group O people tend to produce IgG ABO antibodies more commonly than other blood groups.Just as in other types of HDFN, maternal IgG antibody crosses the placenta and destroys fetal red cells.SymptomsTypical symptoms of ABO HDFN include mild anemia and especially jaundice appearing in the first 24 hours. In rare severe cases the infant can have the more severe symptoms of Rh HDFN, except that prenatal death is unlikely. Rationales to explain the mild nature of ABO HDFN include Fewer A and B antigens on fetal cells Poorly developed fetal A and B antigens Presence of A and B antigens on cells and tissues other than red cells | View Page |
| HDFN Due to Other Antibodies After anti-D, the antibodies that are most often associated with HDFN include: anti-K anti-c anti-E anti-Fya (rarely) anti-Jka (rarely) anti-M,-N,-S,-s,-U (all rarely)Of these antibodies, anti-K, anti-c, and anti-E are more common causes. Anti-K typically causes more severe HDFN (hydrops and neonatal death) than the others. Anti-c has also been known to cause severe HDFN.Antibodies to low frequency antigens have also been known to cause HDFN, albeit rarely. Examples include anti-Mia, -Dia, -Wra and anti-Rd. In such cases the maternal antibody screen is usually negative and the only unexpected test is a positive DAT on the newborn. In theory any IgG antibodies directed against antigens that are well developed on fetal red cells can cause HDFN. The complete list of antibodies documented to cause HDFN is long and will not be covered in this survey course. | View Page |
| ABO HDFN - Diagnostic Tests Before ABO HDFN is considered as a possible cause of jaundice and anemia in the newborn, other causes should be considered, for example, erythrocyte membrane defects or red cell enzyme deficiencies. The diagnosis of ABO HDFN in the laboratory differs from diagnosing Rh and other types of HDFN in which clinically significant antibodies must be identified. Diagnosis may be difficult, because the DAT on the newborn's red cells is unreliable. Indeed, many labs do not routinely do a DAT on infants born to Rh positive females, since many will be positive in the absence of clinically significant hemolysis. Cord blood is often retained (e.g., for 7 days) should the infant develop signs of HDFN and required testing.If ABO HDFN is possible, based on incompatible ABO blood groups and a positive DAT, and the mother's antibody screen is negative, many laboratories do not investigate the positive DAT as would be done for unexpected antibodies like anti-D or anti-K (the laboratory does not perform an elution on the newborn's red cells). Instead, the infant's plasma is tested against group A1 (or B cells) and group O screen cells using the indirect antiglobulin test (IAT). A positive reaction with A1 or B cells, but not group O cells, would suffice to report a case of possible ABO HDFN. | View Page |
| ABO HDFN - Treatment Prenatal treatment Prenatal management and treatment of ABO HDFN is not routinely done because: Titers of anti-A and anti-B do not correlate well with severity of disease; The risks of fetal monitoring (e.g., amniocentesis, cordocentesis) and fetal transfusion are greater than the risk of ABO HDFN since it is usually mild and subclinical. However, if a woman has a history of infants with moderate to severe ABO HDFN requiring treatment, she may be monitored so that the infant can be treated for possible HDFN as soon as possible. Postnatal TreatmentTreatment of ABO HDFN usually consists of phototherapy in which the newborn is placed under a "blue light" that chemically alters bilirubin in the surface capillaries to a harmless substance.For more severe cases, exchange transfusion may be performed. Donor RBC for exchange transfusion in cases of ABO HDFN must meet these criteria: Group O; Rh compatible with infant; Less than or equal to 7 days old (or fresher); Reconstituted with AB FFP to obtain a prescribed hematocrit; CMV negative (or equivalent, e.g., leukoreduced by filtration); Negative for hemoglobin S to prevent blood from sickling under conditions of reduced oxygen concentration in the newborn; Irradiated to prevent graft-versus-host disease. Exchange transfusion is also discussed later in the course in the section related to HDFN due to anti-D and other antibodies. Red Blood Cells are crossmatched with maternal plasma, although the infant's plasma can be used if a maternal blood specimen is unavailable. | View Page |
| For which of the following antibodies is the DAT most likely to be negative when testing a newborn for possible HDFN? | View Page |
| ABO HDFN - Expected Findings Diagnosis of ABO HDFN is supported by these findings: ABO incompatibility between mother and child, with mother typically group O; Maternal antibody screen negative; Cord DAT weakly positive or negative; Newborn hyperbilirubinemia with jaundice occurring in first 24 hours; Increased spherocytes and reticulocytosis in the newborn; Presence of IgG anti-A or anti-B in cord plasma / serum. | View Page |
| Follow-up Investigative Tests (Newborn) If the mother has a clinically significant antibody, routine serologic tests done on the newborn include ABO and Rh; Direct antiglobulin test (DAT). If the newborn's DAT is positive: Elution of newborn's red cells to prepare an eluate containing the sensitizing antibody (unless assessed to be passive anti-D from RhIg*); Antibody identification using eluate. Antibody in the eluate should correspond to at least one antibody found in the maternal serum.* The exception is a positive DAT in a newborn whose mother received RhIg antenatally and who has anti-D at delivery. If other maternal antibodies have been excluded, the positive DAT is assumed to be from RhIg and no elution is performed. | View Page |
| For infants born to Rh negative females, a test for weak D is optional when initial D typing shows the newborn to be Rh negative. | View Page |
| Newborn Serologic Testing Protocols Protocols for testing newborns vary internationally and within countries. The table below summarizes some of the more common protocols. Scenario Typical Newborn Testing Protocol Comments Mother is D-negative with no unexpected antibodies Newborn is tested at delivery for: ABO and Rh Test for weak D (mandatory) if initial Rh typing appears to be D-negative Direct antiglobulin test (DAT)* A positive DAT does not always mean that the newborn has clinically significant hemolysis. A positive DAT commonly occurs due to ABO incompatibility, yet infants seldom require treatment. Infants born to mothers who received antenatal RhIg sometimes have a positive DAT that does not cause clinically relevant hemolysis. Mother is Rh positive and a blood group other than group O Routine testing not performed Cord blood retained for a specified period of time (e.g., seven days) in the event that the mother has an unexpected antibody at delivery or the newborn develops signs of red cell hemolysis. Routine testing would result in many positive DATs due to ABO incompatibility- not clinically significant. Mother is group O Rh positive Newborn is tested- especially important if women and their infants are discharged within 24 hours since hyperbilirubinemia due to ABO HDFN may develop later. Optional only if there is appropriate surveillance and risk assessment before discharge and provided there is follow-up (American Academy of Pediatrics). *Policies for DAT testing of newborns whose mothers have received antenatal RhIg vary internationally. For example, the British Committee for Standards in Haematology guidelines state that a DAT should not be performed on cord blood routinely since in some cases it may be positive due to antenatal RhIg prophylaxis. A DAT is recommended only if HDFN is suspected because of a low cord blood hemoglobin or the presence of unexpected maternal antibodies. However in North America, DATs are always performed on infants born to Rh negative mothers who are RhIg candidates. | View Page |
| A rosette test to screen for FMH is contraindicated if the newborn is weak D. | View Page |
| Literature and Online Resources The following published literature and online resources, while useful, should not be used as a substitute for technical and clinical judgment. Medical and technical information becomes obsolete quickly and current sources relevant to the user's location should always be consulted.References indicated by * provide a broad overview of HDFN and are highly recommended.LITERATUREAvent ND, Reid ME. The Rh blood group system: a review. Blood 2000 Jan 15;95 (2):375-87.Bowman J. Thirty-five years of Rh prophylaxis. Transfusion 2003 Dec;43(12):1661-6.* Eder AF. Update on HDFN: new information on long-standing controversies. Immunohematology 2006;22(4):188–195. (scroll to article)Eder, AF, Manno, C.S. Alloimmune hemolytic disease of the fetus and newborn. In Wintrobe's Clinical Hematology, 11th ed. (Greer JP, Foerster J, Lukens JN, Rodgers GM, Paraskevas F, Glader BE, (eds). Philadelphia, PA: Lippincott, Williams & Wilkins, 2004.Flegel WA. Molecular genetics of RH and its clinical application. Transfus Clin Biol. 2006 Mar-Apr;13(1-2):4-12. Kennedy MS, McNanie J, Waheed A. Detection of anti-D following antepartum injections of Rh immune globulin. Immunohematology 1998;14(4):138-40.Koelewijn JM, de Haas M, Vrijkotte TG, van der Schoot CE, Bonsel GJ. Risk factors for RhD immunisation despite antenatal and postnatal anti-D prophylaxis. BJOG. 2009 Sep;116 (10): 1307-14. Epub 2009 Jun 17.* Kumar S, Regan F. Management of pregnancies with RhD alloimmunisation. BMJ. 2005 May 28;330(7502):1255-8. (UK perspective but much valuable information relevant to all)* Murray NA, Roberts IAG. Haemolytic disease of the newborn. Arch Dis Child Fetal Neonatal Ed 2007 Mar; 92(2): F83–F88. Oepkes D, Seaward PG, Vandenbussche FP, Windrim R, Kingdom J, Beyene J, Kanhai HH, Ohlsson A, Ryan G; DIAMOND Study Group. Doppler ultrasonography versus amniocentesis to predict fetal anemia. N Engl J Med. 2006 Jul 13;355(2):156-64.Ramsey G. Inaccurate doses of Rh immune globulin after Rh-incompatible fetomaternal hemorrhage: survey of laboratory practice. Arch Pathol Lab Med 2009 Mar; 133(3):465-9. Reid ME. The Rh antigen D: a review for clinicians. Blood Bulletin 2008 Apr; 10(1).Sandler SG. Effectiveness of the RhIg dose calculator. Arch Pathol Lab Med 2010 Jul;134(7): 967-8.Shulman IA, Calderon C, Nelson JM, Nakayama R. The routine use of Rh-negative reagent red cells for the identification of anti-D and the detection of non-D red cell antibodies. Transfusion 1994 Aug;34(8):666-70.Tamul KR. Determining fetal-maternal hemorrhage with flow cytometry. Advance 2000. Posted online June 5, 2000.Westhoff CM, Sloan SR. Molecular genotyping in transfusion medicine. Clin Chem 2008;54(12): 1948-50.ONLINE RESOURCESPaxton A. Bringing new rigor to RhIg calculations. CAP TODAY. May 2008. Accessed January 18, 2011.*Wagle S, Deshpande PG. Hemolytic disease of the newborn. eMedicine / WebMD. Updated Apr. 9, 2010. Accessed January 18, 2011. | View Page |
| Introduction This case concerns a common scenario in the transfusion service (TS) laboratory, the detection of anti-D at delivery in a female who has received Rh immune globulin (RhIg) during pregnancy.Distinguishing between passive and immune anti-D is important clinically: If passive anti-D is misinterpreted as immune, RhIg prophylaxis may be omitted leading to D sensitization. If immune anti-D is misinterpreted as passive, appropriate follow-up of the antibody may be curtailed putting the fetus at risk.Unfortunately, differentiating between immune and passive anti-D is often impossible. This case study presents an opportunity to review perinatal testing programs and the crucial role of RhIg in preventing hemolytic disease of the fetus and newborn (HDFN) due to anti-D. The case also examines practical aspects of routine serologic testing involving neonates and women who have received RhIg during pregnancy. The case is a companion to "Hemolytic Disease of the Fetus and Newborn" and complements its content.In brief, the case will: Guide participants through laboratory findings that need to be interpreted and resolved; Examine current best practices in perinatal testing programs; Review the characteristics of RhIg and its use in pregnancy; Review and investigate key issues associated with detection of anti-D in women who have received antenatal RhIg; Discuss crossmatch and LIS policies related to RhIg-derived passive anti-D. | View Page |
| When given during pregnancy, RhIg may cross the placenta and cause a positive DAT in the newborn. | View Page |
| Introduction As noted earlier, in this case study the laboratory's protocol is to set up a mini-panel, providing these criteria are met: Mother is Rh-negative and has been tested on two separate occasions; Laboratory has confirmed administration of RhIg prophylaxis; Result of current antibody screen is positive and typical of anti-D due to RhIg; There is no record or history of an unexpected antibody. All criteria were met and a selected mini-panel was set up to confirm the presence of anti-D and exclude possible co-existing maternal antibodies. Other clinically significant antibodies have implications for possible HDFN and for transfusion to both the mother and newborn, thus must be excluded. | View Page |
| Serologic Tests on Newborn Based on the results of the mini-panel, the laboratory concluded that only anti-D was present and that it was consistent with administration of RhIg at 28 weeks.Patient A.D. delivered a 5 lb 13 oz female by C. section with serologic test results on cord blood as follows. Well washed cord red cells were used for ABO and Rh(D) typing to remove possible Wharton's jelly.Before proceeding to the next page, evaluate if the infant's ABO and Rh(D) types are valid. You will be asked questions that assess basic knowledge of blood grouping practices and test results for newborns. ABO Forward Group ABO Reverse Group Rh anti-A anti-B A1 cells B cells anti-D* 0 0 NT NT 3+ NT = not tested / * monoclonal IgM anti-D DAT Reagent DAT CC Polyspecific AHG w+ 2+ W+ = microscopic positiveAHG = antihuman globulin serum CC = IgG sensitized cells Note: It is the lab's policy to add IgG sensitized cells to weak antiglobulin test results. | View Page |
| The positive DAT on the newborn means that the infant probably has clinically significant hemolysis. | View Page |
| Newborn's Clinical Status The newborn showed no clinical evidence of HDFN or early newborn hyperbilirubinemia, with related laboratory tests as follows (laboratory's reference ranges for newborns in brackets): Test USA SI Hemoglobin 16 g/dL (13.5 - 21.0 g/dL) 160 g/L(130.5 - 226.0 g/L)* Hematocrit 52% (43-62%) 0.52 (0.43-0.62) Total bilirubin (cord blood) 2.1 mg/dL (<2.5 mg/dL) 35.9 µmol/L (<43 µmol/L) *Most countries that adopted SI do not use the official SI unit for Hb (mmol/L), but rather use g/L. | View Page |
| A rosette test to screen for FMH is contraindicated if the newborn is weak-D. | View Page |
| Routine Serologic Tests - Mother Tests done routinely as part of perinatal testing programs vary from country to country and within countries. Below is one example of serologic tests typically done when pregnant females lack clinically significant antibodies. Other test protocols exist.Mother ABO, Rh, and antibody screen at first prenatal visit; Optional (not mandated by blood safety standards): Test for weak D, if initial Rh typing appears to be D-negative; D-negative females: Tested again (ABO, Rh, and antibody screen) at ~ 28 weeks weeks gestation prior to administration of RhIg (depending on the country) and again at delivery. Note: The application of DNA analysis to typing blood group antigens started in the early 1990s but is not yet widely available. When available, the mother can be typed for D using molecular methods, but this is usually not done unless she is weak D. The purpose is to determine using molecular methods which D variant the mother has, weak D or partial D, since the latter can produce anti-D. (see Further Reading) Molecular typing is reviewed more fully in Refresher on Hemolytic Disease of the Fetus and Newborn and Its Prevention, a companion course that complements this one. | View Page |
| For infants born to Rh negative females, a test for weak D is optional when initial D typing shows the newborn to be Rh negative. | View Page |
| Routine Serologic Tests - Newborn Protocols Protocols for testing newborns vary internationally and within countries.if the mother is D-negative and has no unexpected antibodies, newborns are always tested at delivery.Many labs do not test all newborns if the mother is Rh positive and especially do not test if the mother is a blood group other than group O. If all infants born to Rh positive women were tested, many positive DATs due to ABO incompatibility would be detected that are of no clinical significance. Instead cord blood is retained for a period (e.g., 7 days) should it be needed, for example, if the mother has an unexpected antibody at delivery or if the newborn develops signs of red cell hemolysis.However, some clinical practice guidelines, such as those of the American Academy of Pediatrics specify that testing infants born to group O Rh positive mothers is optional only if there is appropriate surveillance and risk assessment before discharge and provided there is follow-up. (See Further Reading) Not testing becomes an issue if group O women and their infants are discharged within 24 hours as occurs in some locations, since hyperbilirubinemia due to ABO HDFN may develop later. Therefore, some facilities where early discharge occurs require that all infants born to group O Rh positive mothers be tested.Typical protocols: Infants born to Rh negative mothers are tested; Infants born to Rh positive mothers who are group O are often tested, especially if early discharge is common (limiting surveillance); Infants born to Rh positive mothers who are not group O are often not tested and this is acceptable good practice. Cord blood is typically retained for a period should it be needed for testing later. | View Page |
| Routine Serologic Tests - Newborn Tests on Newborn ( mandatory if mother is Rh negative) ABO and Rh*; Mandatory: Test for weak D if initial Rh typing appears to be D-negative; DAT**. * ABO typing of the infant does not require a reverse serum group with A1 and B cells since the newborn is not expected to have ant-A or anti-B (unless of maternal origin).* If cord blood is used for ABO and Rh(D) typing, the red cells should be well washed to remove possible Wharton's jelly.** A positive DAT does not indicate that the newborn has clinically significant hemolysis. For example, a positive DAT commonly occurs due to ABO incompatibility, yet infants seldom require treatment. Also, infants born to mothers who received antenatal RhIg sometimes have a positive DAT that does not cause clinically relevant hemolysis.Also note that policies for DAT testing of newborns whose mothers have received antenatal RhIg vary internationally. For example, the British Committee for Standards in Haematology guidelines state that a DAT should not be performed on cord blood routinely since in some cases it may be positive due to antenatal RhIg prophylaxis. A DAT is recommended only if HDFN is suspected because of a low cord blood hemoglobin or the presence of unexpected maternal antibodies.However in North America, DATs are always performed on infants born to Rh negative mothers who are RhIg candidates. | View Page |
| A group A Rh positive mother is about to deliver her infant. Is it acceptable good practice not to test the newborn (ABO, Rh, DAT)?Answer Y (for yes) and N (for no) | View Page |
| Case Summary A group O Rh negative female who had received RhIg at 28 weeks gestation had a weak anti-D when a type and screen was done prior to performance of a Cesarean section (C-section). A mini-panel of selected red cells confirmed the presence of anti-D and excluded other antibodies. The laboratory decided that the anti-D was likely passive and consistent with RhIG administration. A group O Rh positive child was delivered by C-section. The newborn had a weakly positive DAT but was healthy and required no treatment. A rosette test to screen for FMH was negative and A.D. was injected with 1500 IU (300 µg) of RhIG within 72 hours of delivery. | View Page |
| Literature and Online Resources The following published literature and online resources, while useful, should not be used as a substitute for technical and clinical judgment. Medical and technical information becomes obsolete quickly and current sources relevant to the user's location should always be consulted.References indicated by * provide a broad overview of HDFN and are highly recommended.LITERATUREAvent ND, Reid ME. The Rh blood group system: a review. Blood. 2000 Jan 15;95 (2):375-87.Bowman J. Thirty-five years of Rh prophylaxis. Transfusion 2003 Dec;43(12):1661-6.* Eder AF. Update on HDFN: new information on long-standing controversies. Immunohematology. 2006;22(4):188–195. (scroll to article).Eder, AF, Manno, C.S. Alloimmune hemolytic disease of the fetus and newborn. In Wintrobe's Clinical Hematology, 11th ed. (Greer JP, Foerster J, Lukens JN, Rodgers GM, Paraskevas F, Glader BE, (eds). Philadelphia, PA: Lippincott, Williams & Wilkins, 2004.Flegel WA. Molecular genetics of RH and its clinical application. Transfus Clin Biol. 2006 Mar-Apr;13(1-2):4-12. Kennedy MS, McNanie J, Waheed A. Detection of anti-D following antepartum injections of Rh immune globulin. Immunohematology 1998;14(4):138-40.Koelewijn JM, de Haas M, Vrijkotte TG, van der Schoot CE, Bonsel GJ. Risk factors for RhD immunisation despite antenatal and postnatal anti-D prophylaxis.BJOG. 2009 Sep;116 (10): 1307-14. Epub 2009 Jun 17.* Kumar S, Regan F. Management of pregnancies with RhD alloimmunisation. BMJ. 2005 May 28;330(7502):1255-8. (UK perspective but much valuable information relevant to all)* Murray NA, Roberts IAG. Haemolytic disease of the newborn. Arch Dis Child Fetal Neonatal Ed 2007 Mar; 92(2): F83–F88. Oepkes D, Seaward PG, Vandenbussche FP, Windrim R, Kingdom J, Beyene J, Kanhai HH, Ohlsson A, Ryan G; DIAMOND Study Group. Doppler ultrasonography versus amniocentesis to predict fetal anemia. N Engl J Med. 2006 Jul 13;355(2):156-64.Ramsey G. Inaccurate doses of Rh immune globulin after Rh-incompatible fetomaternal hemorrhage: survey of laboratory practice.Arch Pathol Lab Med 2009 Mar; 133(3):465-9. Reid ME. The Rh antigen D: a review for clinicians. Blood Bulletin 2008 Apr; 10(1).Sandler SG. Effectiveness of the RhIg dose calculator. Arch Pathol Lab Med 2010 Jul;134(7): 967-8.Shulman IA, Calderon C, Nelson JM, Nakayama R. The routine use of Rh-negative reagent red cells for the identification of anti-D and the detection of non-D red cell antibodies. Transfusion 1994 Aug;34(8):666-70.Tamul KR. Determining fetal-maternal hemorrhage with flow cytometry. Advance 2000. Posted online June 5, 2000.Westhoff CM, Sloan SR. Molecular genotyping in transfusion medicine. Clin Chem 2008;54(12): 1948-50.ONLINE RESOURCESPaxton A. Bringing new rigor to RhIg calculations. CAP Today May 2008. *Wagle S, Deshpande PG. Hemolytic disease of the newborn. eMedicine / WebMD. Updated Apr. 9, 2010. | View Page |
