Presentation on theme: "Second Year Medical Student Fall Pathology Lab: Transfusion Medicine and Blood Banking October 21st and 22nd 2013."— Presentation transcript:
1Second Year Medical Student Fall Pathology Lab: Transfusion Medicine and Blood Banking October 21st and 22nd 2013
2Outline, Presenters, Contact InfoBlood Donation, Component preparation, and Transfusion indicationsDr. Dan Waxman, M.D.Executive Vice President and Chief Medical Officer of Indiana Blood CenterClinical Professor of PathologyOrdering a blood transfusionDr. Connie Danielson, M.D, PhDChief of Laboratory Service at Wishard HospitalClinical Associate ProfessorBasic principles of transfusionDr. Julie Cruz, M.D.Associate Medical Director of Indiana Blood CenterVolunteer Clinical Assistant Professor of Pathology
4Transfusion Medicine Lab Blood Donation, Component Preparation, and Transfusion Indications
5Transfusion therapy is a set of processes, not just a product Recruit donorMedical reason to TXScreen donorPre-TX testingCollect unitIssue unitPrepare componentsAdminister at bedsideInfectious diseasestestingMonitor & evaluateProduct: Blood safetyEntire process: Blood transfusion safetyAfter S. Dzik, MD Blood Transfusion Service, MGH, Boston
7Whole Blood Description: 500+/- 50 ml mixed with 70 ml CPD Storage: 21 days stored at 1-6ºIndications: Recently used in military hospitals in combat areas settingsProposed clinical trials to examine feasibility and efficacy in civilian setting
9Packed Red Blood Cells Description: Storage = 42 days 200 ml of RBC with 111 ml of additive solutionPacked cell volume = 60%Storage = 42 days
10Packed Red Blood Cells Indications: Acute blood loss exceeding 15-20% of blood volume (pediatric patients ml/Kg) and failure to obtain hemodynamic stability with reasonable volume of crystalloid and/or colloid solutionsAcute blood loss of any amount if there is clinical evidence of inadequate oxygen carrying capacity
11Packed Red Blood Cells Indications: (Cont’d) Hemoglobin of ≤ 7 gm/dl (hematocrit ≤ 21%), if not due to a treatable cause (treatment of underlying case is preferable if patient is not symptomatic)Symptomatic anemia regardless of hemoglobin levelHemoglobin ≤ 8 gm/dl (hematocrit ≤ 24% ) and acute cardiac disease / or shock
12Packed Red Blood Cells Contraindications: For volume replacement In place of a hematinicTo enhance wound healingTo improve general “well-being”
22Saline Washed Red Blood Cells Indications:History of allergic or febrile reactions secondary to plasma proteins not prevented by pre-transfusion administration of antihistamines and leukocyte reductionIgA deficiency with documented IgA antibodiesHistory of anaphylactic reaction to blood components
23Irradiated Blood Products Products irradiated:Whole blood, packed red cells, platelets and granulocyte concentrates
32Platelet Concentrates Storage:Stored at 20-24º C on a rotator5-day outdate
33Platelet Concentrates Indications: prevention and cessation of bleedingSeverely thrombocytopenic (less than 10,000 or 20,000 depending on institution)Moderately thrombocytopenic and bleeding (less than 50,000)Surgery or invasive procedure (less than 50,000)Diffuse microvascular bleeding following cardiopulmonary bypass or with intra-aortic balloon pump (less than 100,000)Bleeding with qualitative platelet defectMassive Transfusion Protocols (MTP)
35Platelet Concentrates Dosage:4-6 platelet concentrates1 apheresis unitPlatelet count should increase 25,000 – 30,000/cc3Each dose has equivalent of one unit of fresh plasma** Unless using platelet additive solution (PAS)
36Frozen Plasma Description: Storage: 1 year at -18º C ml of plasma and CPDA-1, including 25 meq of citric acidJumbo plasma 400 cc or greaterFrozen within 8 hrs = FFPFrozen within 24 hrs = 24FPStorage: 1 year at -18º C
37Frozen Plasma Outdate once thawed (1-6º C) 24 hours for FFP or 24FP hours if relabeled as Thawed Plasma
40Frozen Plasma Indications: Treatment of coagulopathy due to clotting factor deficienciesPatient is bleeding actively with PT and/or PTT greater than 1.5 normal (INR > 1.8) and platelet count above 50,000Coumadin overdose with major bleeding or impending surgeryTreatment of TTPMassive Transfusion Protocol (MTP)
41Frozen Plasma Contraindications: Volume expansion Treatment of nutritional deficiencies
42CryoprecipitateDescription: each unit consists of ml residual plasma than contains 80 units of factor VIII: C and 250 mg of fibrinogenStorage: 1 year at -18º C
45New/Future Products Growth Factors to stimulate cell production Sterilized products to prevent infections*Synthetic products to replace current transfusions**Approval delayed due to regulatory reluctance.
47TRANSFUSION MEDICINE LAB ORDERING A BLOOD TRANSFUSION:What does your patient need?Why? How much? When? Alternatives?Obtain ConsentWrite OrderObtain Specimen and send to Blood Bank
48Case PresentationThe patient is a 64 year old man 2 days after a total hip arthroplasty. He has a history of coronary artery disease and had a MI 5 years ago. He complains that during physical therapy he becomes short of breath and develops chest pain which resolves with rest and sublingual nitroglycerine. Today his Hgb/HCT= 7.5 gms/dL/23%. You decide to order 1 unit of packed red blood cells to be transfused. Why?
49packed Red Blood Cells *1 unit RBC/Adult increases Hgb 1 gm/dL* Indication – patient is anemia AND symptomaticHgb < 8gm/dLPatient complaining of shortness of breath*1 unit RBC/Adult increases Hgb 1 gm/dL*Since the 1990’s it has become clear that “over-transfusion” can increase morbidity and mortality.
50Transfusion Guidelines: In the USA currently accepted guidelines for transfusion of red cells is generally:Hgb ≤7 g/dL many need transfusionHgb 7-9 g/dL variable need for transfusionHgb≥ 10 g/dL very few need transfusionBlood is a drug with a variable safety profile (discussed later)
51Normal levels of Hgb in adults Men: 13.8 to 18.0 g/dLWomen: 12.1 to 15.1 g/dLIf the concentration is below normal, this is called anemia.Hematocrit, the proportion of blood volume occupied by red blood cells, is typically about three times the hemoglobin concentration measured in g/dL.
70Basic Principles of Transfusion ABO, Rh, Other Blood GroupsPretransfusion Testing
71N-acetylgalactosamine B D-galactose ABO Blood Groups4 Types: A, B, AB and Othe name of the blood group corresponds to the presence of the antigen associated with the sugar residue (ie, a person whose red blood cells carry the A antigen has blood group type A. Type O red blood cells lack these terminal sugars, and the corresponding antigen is called the “H” antigen.Antigen SpecificityImmunodominant sugarHL-fucoseAN-acetylgalactosamineBD-galactose
72ABO Antigens and Antibodies Blood GroupAntigen on Red Blood CellAntibodies in serumGroup OH (neither A nor B)Anti-A, Anti-BGroup AAAnti-BGroup BBAnti-AGroup ABA and BNeither anti-A nor anti-B
73ABO compatibility is the primary concern when selecting a unit of blood for transfusion Other antigens are present on red blood cells, and antibodies to these antigens are also important. However, these antibodies occur only after the individual has been exposed (or sensitized) to the antigen through transfusion or pregnancy. This is also called alloimmunization.
77ABO Typingforward type involves testing the patient’s red blood cells with commercial anti-sera containing anti-A and anti-sera containing anti-B. Reactivity with the specific anti-sera indicates that antigen is present on the patient’s red blood cellsreverse type combines the patient’s serum with reagent red blood cells that are either type A or type B. Reactivity indicates the patient has circulating antibody to the corresponding antigen on the reagent red blood cell.Reactivity occurs in the form of agglutination – which is the clumping together of the red blood cells resulting from the interaction of the antibody and its corresponding antigen. This clumping can be observed visually or detected photometrically in automated equipment. Strength of agglutination is graded from 0 (no agglutination) to 4+ (very strong).
80ABO Forward and Reverse Groupings Forward group: Patient’s cells with reagentsReverse Group: Patient’s serum with reagentsBlood GroupAnti-AAnti-BAntigen(s) on RBCsA1 cellsB cellsAntibody(ies) in serumONo A or B+A and BABAB
81The Rh Blood Group System RhDThe RhD gene is either present or absent – it’s absence is denoted by the placeholder “d”. There is no “d” gene, and no “d” antigen.An individual who possesses no copies of the RhD gene (did not inherit RhD from either parent) is indicated by d, and said to be “Rh negative”. Eighty-five percent of Caucasians express the D antigen.Some RBC’s exhibit a weaker than normal form of the D antigen, giving weaker or negative reactivity with standard anti-D reagent. This antigen expression is known as weak D.
82RhCEThe RhCE gene codes for either RhCe, RhcE, Rhce, or RhCE proteins. Again, these genes are inherited as a haplotype from each parent. The following table shows the possiblehaplotypes and their frequencies utilizing Fisher-Race and Weiner terminology.Prevalence (%)HaplotypeFisher-RaceWeinerWhiteBlackAsianDCeR1421770dcer37263DcER2141121DceR0444dCer’2dcEr”1<0.01DCERzdCEry
83Calculating prevalence Since one haplotype is inherited from each parent, the phenotype frequencies may be obtained by multiplying the haplotype frequencies for the population. For example, the frequency of the dce/dce (rr) phenotype in the Caucasian population is .37 X .37 = .14 or 14%. So, if an alloimmunized individual required red blood cells of this type for transfusion, approximately 14% of randomly collected donor units would be appropriate.What frequency would you expect to find the dce/dce (rr) phenotype in the Black population?
84Other Blood Group Systems LewisAntibodies to the antigens in the Lewis system are rarely clinically significant. The antigens are not synthesized on the red cells, but are absorbed from the plasma. The importance of the Lewis system is more its association with the Se gene and the determination of an individual’s secretor status.KellAfter ABO and Rh, the K antigen is the most immunogenic blood group antigen causing both HDFN and delayed hemolytic transfusion reactions. About 10% of whites and 2% of blacks are positive for the K antigen. The antithetical antigen, k, (pronounced Cellano) is a high incidence antigen, meaning <1% of the population lacks this antigen.
85Other Blood Group Systems KiddTwo antigens, Jka and Jkb are responsible for the common phenotypes. The Kidd antigens are located on a membrane glycoprotein involved in urea transport in the red cell. They are often weakly reacting antibodies showing dosage. Dosage refers to stronger expression of the antigen when the individual inherits the gene homozygously, vs. heterozygously. It is important that antibodies to these antigens are ruled out on homozygous reagent cells, since a heterozygous cell may be nonreactive due to the low expression of the antigen. Kidd antibodies are notorious for weakening over time and even becoming undetectable. Despite this, they cause severe delayed hemolytic transfusion reactions. They are also implicated in HDN, but it is usually a milder form.
86Other Blood Group Systems MNSThe most important antigens that affect transfusion are M, N, S, s and U. The M and N antigens are carried on glycophorin A, and S,s, and U are carried on glycophorin B. Anti-M and anti-N can occur as naturally occurring agglutinins (IgM) and as such usually react at room temperature or below. They do not cause HDFN. These antibodies can usually be ignored for transfusion, unless they are also reactive at 37 C due to the presence of an IgG component. In contrast, S and s are IgG reactive at 37 C, are clinically significant and cause HDFN. Anti-U is a rare antibody which is associated with HDFN and hemolytic transfusion reactions, usually seen in the African American population. It occurs in individuals who are S-s-U-, and individuals who have anti-U are incompatible with greater than 99% of the donor population.
87Other Blood Group Systems DuffyThe Duffy system is made up of six antigens that reside on an acidic glycoprotein . The glycoprotein carrier molecule is also known as the Duffy antigen receptor for chemokines (DARC) and functions as a cytokine receptor. It is also a receptor for the malarial parasite Plasmodium vivax. The null phenotype Fy(a-b-) is rare in Caucasians but common in blacks. It is thought that this developed as a survival advantage in malarial regions of Africa, since such individuals are naturally resistant to infection by P. vivax. Anti-Fya is a relatively common antibody, while anti-Fyb is less common. Both are clinically significant and associated with hemolytic transfusion reactions and HDFN.
88Antibody ScreenThe goal of antibody screening is to detect unexpected clinically significant red cell antibodies. In general, clinically significant antibodies are antibodies known to have caused hemolytic transfusion reactions, hemolytic disease of the fetus and newborn (HDFN), or shortened survival of transfused red blood cells. The method used must be capable of detecting clinically significant antibodies, which requires that the antibody screen method include a 37 C incubation with reagent red cells followed by an indirect antiglobulin test (IAT).
89The Indirect Antiglobulin Test (IAT): The IAT is used to detect in-vitro sensitization and detects anti-red cell antibodies in patient’s serum or plasma. Procedural steps are as follows:Patient’s plasma or serum is incubated at 37 C with red cells (screen or panel cells of known antigenic composition, ie. reagent red cells, or donor cells as in the crossmatch).A potentiator (LISS, PEG) may or may not be addedDuring incubation, if an antibody is present in the plasma or serum and the corresponding antigen is present on the red cells, the cells become sensitized by the antibody adsorbing to antigens on the red cell surfaceAfter incubation, the red cells are washed with saline to remove unbound antibodyAntihuman globulin serum (anti-IgG or polyspecifc AHG, usually the former) is added and forms RBC agglutinates if the antibody has attached to the antigen sites during incubation
91Crossmatch Methods: Immediate spin crossmatch: The immediate spin (IS) crossmatch is performed only after an antibody screen is done and found to be negative on a current specimen. It is performed at room temperature without enhancement. The patient should have no history of clinically significant antibodies. The immediate spin crossmatch is meant to detect ABO incompatibility. It can also detect cold reactive (generally clinically insignificant) antibodies that react at room temperature.Antihuman Globulin (AHG) crossmatchThe AHG is performed as described for the IAT above, and tests the patient’s serum or plasma against the donor RBCs intended for transfusion to the patient. It will detect ABO incompatibility as well as other clinically significant antibodies, such as antibodies in the Rh, Kell, Kidd, and Duffy systems.
92Computer-Assisted (Electronic) Crossmatch Principle:Using a validated computer system, patients with no history of clinically significant antibodies and a negative antibody screen on the current specimen are issued ABO specific or compatible donor units. Specific conditions must be met:Two independent determinations of the ABO group of the patientNo history of clinically significant antibodiesA negative antibody screen on a current specimenDonor units are ABO confirmedThe computer system is validated on site, results are entered directly into the computer, and there is logic in the system to recognize incompatibility
93In the last section:You ordered type and cross for 1 unit of red blood cells for your patient.Now, you are the blood bank technician:You receive an appropriately labeled and collected specimen with an order to type and cross for one unit of red blood cells. First, you perform the ABO and Rh Typing:
94Forward type: tube on left contains anti-B, tube on right contains anti-A.
95Reverse type: Tube on left contains A1 Cells, Tube on right contains B cells ForwardReverseAnti-AAnti-BType?A CellB Cell
101Antibody is present to what antigen? What does this imply for transfusion?
102A unit of RBCs that is Type A, Rh positive and confirmed negative for the Jkb antigen is selected for the patient. The AHG crossmatch is performed and the unit is compatible. Is this unit appropriate for the patient?
105Outline Definition Types of transfusion reactions Acute febrile transfusion reactionsAcute non-febrile transfusion reactionsDelayed febrile transfusion reactionsDelayed non-febrile transfusion reactionsRisk of Transfusion reactionsApproach and follow-up to suspected transfusion reactions
106Definition Unfavorable events that occur during or after transfusion Can happen with any blood componentPRBC, Plasma, Platelets, CryoprecipitateRho (D) immune globulin, IVIG, Factor concentrateStem cells, Human albumin, Granulocytes
108Acute hemolytic transfusion reactions (AHTR) Most severe reaction (may be fatal!)All reactions are hemolytic until proven otherwiseABO incompatibility most commonly from clerical errorIgM antibodies bind to the RBCs → complement activation → formation of MAC → RBC lysisCoagulation cascade activation from Ag-Ab complexes →Intravascular thrombi → schistocytesCoagulation cascade and complement → cytokine production → additional symptomsTreatment is supportive
109Signs and Symptoms Fever/Chills 1°C or 2° F (most common) TachycardiaHypotensionBleedingDyspneaImpending doomNausea/VomitingPain: flank, back, chest, head, infusion siteDark urine (hemoglobinuria)Death
110Laboratory Findings Hemoglobinemia (pink or red serum/plasma) Hemoglobinuria (NOT hematuria)Usually positive direct antiglobulin test (DAT) but can be negative (all Ab coated cells already lysed)Elevated indirect bilirubin and LDHDecreased haptoglobinHyperkalemiaPeripheral smear: Schistocytes
112Transfusion Related Acute Lung Injury (TRALI) #1 cause of fatal transfusion reactions in the USUsually donor anti-HLA or anti-neutrophil antibodies attach to neutrophils in lung → neutrophil activation → vascular damage → edema and impaired gas exchangeBegins during or within 6 hours of transfusionNo pre-existing acute lung injuryPrevented by use of male plasmaWhy???
113Symptoms and Other Findings DyspneaFeverTachycardiaHypotensionFrothy endotracheal aspirateDeathCXR shows white-out/edema
114Suspected TRALIDonor testing for anti-HLA and anti-neutrophil antibodiesPermanent deferral for donor if positive
115Bacterial Contamination Contaminated blood product → sepsisMost common in platelet productsWhy???Symptoms:Rapid high fever, hypotension, tachycardia, and shockMay also have violent rigors, nausea, vomiting, diarrheaShock and death
116Febrile non-hemolytic transfusion reactions (FNHTR) Most frequently reportedTemperature increase of >1°C or >2°F with no other explanationCytokines from WBC in product during storage or antibody activation of WBCUsually mild symptomsFeverChills (may be only symptom if pt is premedicated)Treatment:Antipyretics (acetaminophen)Leukoreduced products (pre-storage)
118Transfusion associated circulatory overload (TACO) Fluid overload precipitated by blood productRisk factors: Renal impairment and heart failureSigns and symptoms: Dyspnea, tachycardia, hypertension, jugular venous distensionTreatment: Support respiratory function and diuretics
120Allergic Reaction Second most common type of transfusion reaction Only reaction that allows the restart of procedure if symptoms resolveCaused by type I hypersensitivity to donor plasma proteinsSymptoms: Pruritis, urticaria, erythemaTreatment/PreventionBenadrylCould give plasma deficient products (washing)
121Anaphylactic Reaction Most severe allergic reactionSymptoms: Rapid onset of laryngeal edema leading to dyspnea, tachypnea, hypotension, tachycardia, afebrileOften associated with recipients that lack IgA AND have anti-IgACan give washed or IgA deficient products
123Delayed Hemolytic Transfusion Reactions (DHTR) Anamnestic response = resurgence of antibody faster and more severe after second exposureIgG antibodies bind to the RBCs(sensitized cells) → processed by reticuloendothelial system→ RBCs removed by spleenIncompletely removed cells result in spherocytesMain causes Kidd (Jk) and Duffy (Fy) systems
124Signs and Symptoms Fever/chills (mild) Mild jaundice Anemia/pallor SplenomegalyRarely death
125Laboratory Findings DAT positive Hyperbilirubinemia Usually no free hemoglobin (versus AHTR)“New” antibodySpherocytes on peripheral smear
126Transfusion-Associated Graft versus Host Disease Highly (> 90%) fatalDonor WBC recognize pt’s HLA antigens → WBC activation → destruction of host tissuesSigns and symptoms: Fever, diarrhea, skin rash, bone marrow suppressionTreatment/PreventionSupportive careIrradiation of blood productsWhy???
128Post-transfusion Purpura Patient abs against donor platelet antigens (usually PL-A1)PL-A1 negative patients (RARE)Signs/symptoms: Profound thrombocytopenia, purpura, bleedingPatient’s own platelets undergo destruction as well (unknown mechanism)Treatment: supportive/plasmapheresis
129Iron OverloadTransfusion dependent anemia (i.e. sickle cell and thalassemia patients)Iron accumulates with multiple transfusionsResults in end-organ damageTreatment/preventionChelation therapyExchange transfusions
130Risks of Blood Product Transfusion Hives: 1 in 30 to 100Febrile reactions: unknown quantity, but commonTransfusion Associated Circulatory Overload (TACO): 1 in 3,000 to 12,000Transfusion-Related Acute Lung Injury (TRALI): 1 in 5,000Acute Hemolytic Reactions: 1 in 15,600 to 35,700Bacterial Infection: 1 in 20,000Hepatitis B: 1 in 200,000HTLV-1: 1 in 641,000Hepatitis C: 1 in 1.2 millionHIV: 1 in 1.4 millionOther infectious diseases such as West Nile virus: less than 1 in millionOther infections such as Babesiosis or Chagas: very rareAnaphylaxis: 1 in 20,000Indianapolis Coalition for Patient Safety, Transfusion consent form.
131What To Do For Suspected Transfusion Reaction? Stop TransfusionKeep line open with salineReport to physicianOrder transfusion work upClerical check at bedsideUnit and tubing need to be returned to blood bankPatient blood sample sent to lab
132Laboratory Work-Up Examine blood for hemolysis Perform DAT Repeat type and crossIf greater then >2°C, send retention segment and bag for cultureAlso, blood cultures on patientIf suspect HTRCollect first voided urineCheck bilirubin 6 hours post transfusionCheck haptoglobinCheck CBC
133References References available upon request. Thank you: Judi Seidel, MT (ASCP) SBBDr. Steven Gregurek M.D.
135MS2 Pathology Lab Blood Bank Lab 10/21 & 10/22 Ted Kieffer PGY3 Case PresentationMS2 Pathology LabBlood Bank Lab 10/21 & 10/22Ted KiefferPGY3
136Case PresentationA 25 year old pregnant Chinese female is brought to the emergency department following a motor vehicle accident. She does not speak English but appears to be in significant pain and grasping her lower abdomen.What do you do?
137Case PresentationHistory and review of systems cannot be obtained due to language barrier and no translator readily available.VitalsHR 110 beats per minuteResp 20 breaths per minuteBP 135/90Sats 97% on room air
138What else might you want to know? Case PresentationPhysical examRelatively unremarkable aside fromAbdominal exam which reveals a band-like contusion across the lower abdomen and a firm, tender uterus with a fundal height of 26 cmVaginal exam remarkable for bloodWhat else might you want to know?(i.e. who else might you want to evaluate?)
139Fetal heart rate by doppler – 160 bpm Reasurring? Case PresentationFetal heart rate by doppler – 160 bpmReasurring?Current Differential Diagnosis?Placental abruptionPreterm LaborPlacenta previaUterine ruptureSubchorionic hematoma
140Case PresentationCurrent Differential Diagnosis: Placental abruption – acute onset, tender & firm abdomen/uterus, maternal hypotension Preterm Labor – gradual onset, progressive, Mucus plug may be mistaken for vaginal bleeding Placenta previa – characteristically painless vaginal bleeding at >20 weeks gestation Uterine rupture – sudden onset FHR abnormalities, vaginal bleeding, abdominal pain, and maternal hypotension Subchorionic hematoma – light vaginal bleeding, generally no abdominal pain, <20 weeks gestation
141Case Presentation Lab tests? Imaging? Anything else? Orders? CBC BMP + Ca, Mg, PhosType and crossCoag studiesIncluding: Fibrinogen, fibrinogen degradation products (FDP), D-dimer*other testsAlpha fetal protein (AFP), human chorionic gonadotropin (hCG)**Kleihauer-Betke test**Imaging?UltrasoundAnything else? Orders?Fetal heart rate monitorPain medication for mom
142Case Presentation Diagnosis!!! Placental Abruption Abdominal US/FAST scanDiagnosis!!!Placental AbruptionRadiologist eventually reads sonogram and interprets findings as small abruption
143Treat Patient – Tocolysis Case PresentationWhat do you do now?Vitals still stableLab resultsCBC –8.9>------<34731BMP – 142/105/10 Ca – 4.9 mg/dL (wnl)<95 Mg – 2.0 mg/dL (wnl)4.0/24/1.1 Phos – 3.5 mg/dL (wnl)Anything else?Treat Patient – TocolysisMagnesium Sulfate
144What do you do now? Case Presentation Lab results Coag tests PT – 9 seconds (normal sec)aPTT – 21seconds (normal sec)Fibrinogen – 350 mg/dL (wnl)FDP – 5 mcg/mL (wnl)D-dimer - <5μg/mL FEU (wnl)AFP & hCG wnlKleinhauer–Betke test – 0.2%Hypercoag?Why did you order these?
145What do you do now? Case Presentation More lab results DAT B-cells A-cellsAnti-D Anti-A Anti-BWhat do you do now?More lab resultsType and crossABO type ARh negativeScreen positiveDAT positive with mixed fieldDATAntibody screen4+
146Case Presentation Antibody Panel 4+ 4+ 4+ M 4+4+4+MAnti-D with a mixed field on patient sample
147Any other questions you would like to ask? Prior birth history? Case PresentationAt that moment the translator arrives and informs you the patient is feeling much better, abdominal pain has decreased, and that her contractions appear to have ceased.Any other questions you would like to ask?Prior birth history?
148Case PresentationThe translator informs you that the patient has had two children prior to this pregnancy, all with the same father. The first was born without complications but the second was jaundiced for several weeks. All prenatal and postnatal care was received in rural China.
149Lets bring it all together Case PresentationSo what is happening? What must you be concerned about? What are your next steps?Lets bring it all togetherDiagnosis of placental abruptionBlood tests indicate two different RBC inmom’s bloodMother is Rh negativeAnti-D antibody identified on RBC panelHistory of complicated pregnancyFMHHDFN
150Case Presentation Causes of Fetal Maternal Hemorrhage (FMH) Indications for administering RhIG if mother is Rh- and fetus Rh+/unkDeliveryInduced abortionSpontaneous abortionEctopic pregnancyPartial molar pregnancyChorionic villus samplingCordocentesisPercutaneous fetal procedures (eg, fetoscopy)AmniocentesisExternal cephalic versionAbruptio placentaAntenatal hemorrhageMaternal abdominal traumaSpontaneousManual removal of the placentaAt 28 weeks of gestationSpontaneous abortion, threatened abortion, induced abortionEctopic pregnancyInvasive procedures: genetic amniocentesis; chorionic villus sampling; multi-fetal reduction; fetal blood samplingHydatidiform moleFetal death in the second or third trimesterBlunt trauma to the abdomenAntepartum hemorrhage in the second or third trimester (eg, placenta previa or abruption)External cephalic version
151Management of FMH – RhIG Case PresentationManagement of FMH – RhIGKleihauer-Betke teststandard method of quantitating fetal-maternal hemorrhage for calculating RhIG doseMaternal blood smear exposed to acid pH which dissolves adult hemoglobin and leaves fetal hemoglobin intact – special stain applied (Shepard’s) which stains hemoglobin and percentage of fetal cells is calculated out of 2000
152Management of FMH – RhIG Case PresentationManagement of FMH – RhIGUsing Kleihauer-Betke testCalculate maternal blood volume:prepregnant wt kg x 70ml/kg x [1.0 + (0.5 x wks gestation/36)] - est blood lossOr estimate 5LCalculate fetal blood volume in maternal circulation:(% from KBT) x maternal blood volumeCalculate dose of RhIG1 vial (300μg RhIG)/30mL fetal bloodfudge factor – round up one vial for values of n.0 to n.4; round up two vials for values of n.5 to n.9
153Management of FMH – RhIG Case PresentationManagement of FMH – RhIGUsing Kleihauer-Betke test in our patientCalculate maternal blood volume:No info given – using 5L or 5000mLCalculate fetal blood volume in maternal circulation:(0.2%) x 5000mL = x 5000mL = 10mLCalculate dose of RhIG1 vial/30mL fetal blood x 10mL = 0.33 vials = 1 vial (rounded up)You administer 1 vial of RhIG to the patientWhy did you go through all of that effort?
154Hemolytic Disease of the Fetus/Newborn (HDFN) Defined as the destruction of fetal and neonatal red blood cells by maternal antibodies (termed alloantibodies) – Historically IgG Rh/anti-D antibodies
155Pathophysiology of HDN Maternal IgG antibodies cross the placenta and attach to fetal red blood cellsRed blood cells hemolyzed or removed via reticulo-endothelial systemResultant anemia causes accelerated production of RBCs by bone marrow, termed erythroblastosis fetalisIn severe disease, bone morrow inevitably fall short of necessary RBC productionBody responds with extramedullary hematopoesis in the spleen and liverHepato-splenomegaly causes portal hypertension and hepatocellular damageAnemia coupled with hypoproteinemia leads to massive, diffuse edema and high cardiac output heart failure
157Sequelae of HDN Hyperbilirubinemia RBC destruction does not cease with deliveryIgG due to its small size as a monomer distributes throughout tissues (intravascular and extravascular)IgG has a half life of 25 daysPrior to delivery bilirubin is transported across placenta, conjugated by maternal liverBilirubin conjugation system in a neonate is immatureWithout therapy unconjugated/indirect bilirubin can reach toxic levels (18-20 mg/dL) and diffuse into the brain causing kernicterus and acute bilirubin encephalopathy
159Development of alloantibodies Mother must be antigen negativeAntigen positive individuals will not form antibodiesMother must be exposed to antigenFeto-maternal hemorrhageTransfusion with ABO compatible, minor group incompatible bloodInjection with needles contaminated with minor group antigen positive bloodMinor blood group mismatch allogeneic stem cell transplantAntigen exposure induces antibody formationExact volume unknown (varies between individuals) but as little as 0.1 mL of Rh+ RBC have been shown to stimulate antibody productionLarger volume of exposure tends to produce a more robust responseFollowing antibody production mother must become pregnant with an antigen positive fetusThe initial response will be IgM which will not cross the placentaIn most cases, the first minor group incompatibility between mother and fetus will not be affected, with exceptionsSubsequent exposure will induce memory cells to produce IgG antibodies which will in turn cross the placenta and cause hemolysis or clearance
160Development of fetal alloantibodies Mendelian geneticsGenes code for an enzyme (ABO system), surface protein (Rh system), or nothing (silent/amorph)Blood group genes major (ABO)and minor (all others) are inherited in a Co-dominant patternEach parent contributes half of the inheritanceIndividual traits are inherited independent of each other
162Development of fetal alloantibodies Rh +Rh -RhUnlike A and B antibodies,many minor antibodies arenot naturally occurringD dd dFirst Rh + child sensitizes MotherMother develops Anti-D antibodiesSecond child conceivedIgG Alloantibodiescross placentaSecond ChildD dFirst ChildD dd dd dRh +Rh +Rh -Rh -
163Why only IgG antibodies? Only IgG type antibodies can cross the placentaActively transported via a receptor specific to IgG Fc regionStarts in the second trimester and continues until birth
164Antigens associated with HDFN Major incompatibilityABO antigens – usually mild HDFN, A and B antigens less well developed in neonate (actually protects from minor incompatibility)Minor incompatibilityFrequently associated with severe HDFNRh (D, c) and Kell (K)Infrequently associated with severe HDFNSee next slide
165Antigens infrequently associated with severe HDFN Infrequently associated with severe diseaseColtonCoaMNSMtaRhesusHOFMCo3MUTLOCRDiegoELOMurRivDiaMvRh29DibsRh32WrasDRh42WrbSRh46DuffyFyaUSTEMKellJsaVwTarJsbBeaOther antigensHJKk (K2)CJFVKpaCeJONESKpbCwKgK11CxMAMK22ceREITKuDwRdUlaEKiddJkaEwEnaEvansFareHilGHutGoaMHrMiaHr0MitJALAntigens infrequently associated with severe HDFN
166Rh Alloimmunization: diagnosis Rh(D) typing and screenPerformed at first prenatal visitIf Rh(D)-, screen negative, and prenatal course uncomplicatedRepeat at 28 weeksRepeat at deliveryIf Rh(D)-, screen positive diagnostic testsIndirect antiglobulin test on maternal serum with titerDirect antiglobulin test on fetal red blood cells with titerPositive agglutination in saline or albumin indicate Anti-D IgM production
167Prevention of Rh Alloimmunization Anti-D immune globulin/RhIGIgG anti-D manufactured from human plasma (primarily male who undergo repeat injections of Rh+ blood)Preparation methodsHyperRho S/D®, RhoGAM®Cohn cold ethanol fractionation followed by viral-clearance ultrafiltration – intramuscular only as IgA and other plasma proteins have the potential to produce anaphlyaxisRhophylac®, WinRho SDF®Ion-exchange chromatography isolation – intramuscular or IV
168Prevention of Rh Alloimmunization Anti-D immune globulinMechanism of action: epitope masking?, rapid clearance?GuidelinesWeak D positive managed as Rh-Mother Rh-, fetus confirmed or suspected Rh+300 micrograms early in third trimester300 micrograms if there is increased risk of feto-maternal hemorrhageRepeat doses if risk is ongoing, guided by titers300 micrograms within 72 hrs after delivery of a Rh+ infantIf inadvertently not administered, give ASAP – partial protection has been seen up to 13 days after birth
169Diagnosis and Treatment of Intrauterine fetal anemia Diagnostic techniques:UltrasoundDoppler assessment of MCA peak velocityPercutaneous umbilical cord samplingAllele-specific PCR on fetal cells in amniotic fluidItrauterine fetal transfusionTransfuse when fetal hematocrit falls below 2 standard deviations of mean hematocrit for gestational ageCan be performed between 18 and 35 weeksIntraperitoneal transfusions not as effective as intravascular transfusions in hydropic fetus due to congested lymphaticsABO type O, Rh- packed RBC utilizedFetal loss 1-2% with overall survival of 85% after transfusion
170Case Presentation Finalizing the case What do you order? Maternal Anti-D titer1:64US of fetus and MCA peak velocitySlightly hydroptic fetusMCA peak velocity elevatedFetal CBCModerate anemiaIntrauterine fetal transfusion performed by IRMom transferred to high risk OB service where last you heard she was doing very well and expecting to be discharged