Mom_Baby Studies 2023 Crystal.html

PARENT/BABY STUDIESPREGNANCY RELATED SEROLOGIC TESTING Crystal Berger Fall 2023 OBJECTIVES Describe routine prenatal and postpartum testing used to diagnose, monitor and treat Hemolytic Disease of the Fetus and Newborn (HDFN) Compare and contrast ABO vs. Non-ABO HDFN Compare and contrast the tests...

PARENT/BABY STUDIESPREGNANCY RELATED SEROLOGIC TESTING Crystal Berger Fall 2023 OBJECTIVES Describe routine prenatal and postpartum testing used to diagnose, monitor and treat Hemolytic Disease of the Fetus and Newborn (HDFN) Compare and contrast ABO vs. Non-ABO HDFN Compare and contrast the tests used for the determination of fetomaternal hemorrhage Calculate RhIg dosage Describe process of pretransfusion testing in neonatal period WHAT IS THE CONCERN? Hemolytic Disease of the Fetus and Newborn (HDFN) ABO Non-ABO Can lead to miscarriage/stillbirth/early death OUTDATED TERMINOLOGY The terminology for prenatal and postnatal care is gendered. To be more inclusive and to reflect the patients that we serve, I will do my best to use non-gendered terms Transgender and non-binary people can and do get pregnant access to healthcare and acceptance increase we will see more of these patients For the ASCP and Older Providers the terminology is: Mom/Baby Studies Maternal antibody PRENATAL PRENATAL TYPE AND SCREEN (PREN) Performed early in the pregnancy ABO and Rh determination Policy for weak D testing varies by laboratory UWMC: test for weak D if there is an ABO/Rh discrepancy or the Anti-D is weakly positive Another lab: test person of childbearing potential for Weak D if the Anti-D is weakly positive or negative Screen Looking for unexpected RBC antibodies capable of causing HDFN RH NEG PARENT RhIg (Rhogam/Rhophylac/WinRho) Presumption that fetus is Rh POS until proven otherwise Administered: All pregnancies: Antepartum at 28 Weeks ne of the following occurs: miscarriage, abortion, trauma (fall or accident), or invasive procedure (examples:  IUT, amniocentesis, repositioning) Within 72 hours of delivery if: Baby is D pos or weak D pos or fetal Rh is unknown Patient is not already sensitized to D antigen RhIg will show up on antibody screen  Some labs may perform a Mini Panel for Anti-D vs a Full Panel are unable to distinguish between RhIg administration and allo Anti-D Need a titer performed now and again in several months  is preferred to have prenatal sample collected prior to RhIg administration ANTIBODY SCREEN POSITIVE Perform antibody ID Titer antibody if it is IgG (Clinically significant is capable of causing HDFN) Titer previously tested sample, if available, in parallel Standard practice to freeze prenatal samples for retesting later in pregnancy rule-out difference in technique (unnecessary if using gel) Serial Dilutions Use Homozygous cell - why? Significant if ≥16 using tube method anti-K is significant at ≥8 using tube method Gel method is more sensitive and will have different action levels Last time I checked, a paper has not been written comparing the two methodologies Repeated throughout pregnancy Watch for 2 fold increase Paternal phenotype – very uncommon (anecdotally, seems to be increasing) MONITOR THE FETUS Ultrasound – low risk Check for signs of hydrops Amniocentesis – increased risk of miscarriage or spontaneous delivery phenotype the baby Measure the amniotic fluid spectrophotmetrically and calculate the change in optical density (∆OD) at 450nm This measures the concentration of bilirubin pigment Cordocentesis – increased risk of miscarriage or spontaneous delivery Measure the hemoglobin in the cord blood INTRAUTERINE TRANSFUSION (IUT) Clinically indicated when: Change in optical density at 450nm is in Zone III or high Zone II of the Liliey graph Cordocentesis hemoglobin is less than 10 g/dL Fetal Hydrops is noted on ultrasound examination Also indicated for non-HDFN medical issues Twin to Twin transfusion The decision to start IUTs is a balancing act trying to give the fetus more time in case of spontaneous delivery without waiting too long and causing harm to fetus IUT PRODUCT REQUIREMENTS Blood needs to be: Reasoning: Irradiated (freshly irradiated is best) Prevent TA-GVHD (fresh IRR to reduce risk of hyperkalemia) Leukoreduced Prevent Cytomegalovirus infection Fresh (<5 days old or <7 facility dependent) Reduce risk of hyperkalemia and maximize 2,3 DPG levels O Neg (Parental blood can be used in case of high prevalence antibody) Reduce ABO HDFN risk Antigen Compatible with Parent Must test antigen negative for parental antibody Hgb S negative Remove chance of sickling in hypoxic and/or acidotic conditions  Possible Modification: Washing or Supernatant removed Remove diluent/anticoagulant to increase hematocrit A NOTE ON IUT PRODUCT REQUIREMENTS There is no standard practice for IUT unit preparation, there will be an article published soon in AABB's Transfusion  Some facilities wash the RBCs to remove anticoagulant and raise the hematocrit with others perform supernatant reduction Some facilities only use RBCs in CP2D anticoagulant Many blood centers are consolidating their product offerings due to decreased staffing and cost Studies show that it is safe to use other anticoagulants (see sources for white sheet and list of studies if interested) Some facilities use the "Belt and Suspenders" approach and use CMV Neg and Leukoreduced Desired Hematocrit also varies by facility UWMC's target is 72-75%  POSTNATAL TSCR ON PARENT May need to provide blood to parent Establishing baseline for care What is your hospital population? Parent may have been alloimmunized since prenatal testing was performed ABO/RH ON BABY Perform the forward type only Weak D testing varies by testing lab UWMC policy: perform if baby is D negative AND parent is a potential candidate for RhIg Other labs: perform on all D negative babies Reverse type not performed antibodies YET Any antibodies detected would be from the parent Sample source: Cord blood Heel stick Line draw CHALLENGES FOR TESTING NEWBORNS Cord Blood: Risk of parental contamination Wharton’s Jelly Cannot be blood type of record Newborns in general Mixed field in ABO – is it Parental contamination or incomplete expression of A or B antigen (not fully expressed until 4 years old)? Mixed field is even more pronounced in pre-term infants Did the parent undergo IUT during pregnancy? Interference from Parental antibody – Parental antibody coating cells so much that the control is positive resulting in an invalid ABO Perform EGA treatment to strip bound antibody from the red cells so that the red cells can be tested DAT ON BABY Only IgG is a concern May detect ABO HDFN Especially if Parent is group O and baby is A, B, AB* DAT can be Weak Positive or negative - Why? DAT is positive for IgG: perform an elution and test the eluate Eluate testing policy varies by lab: UWMC will not perform an elution if the positive DAT can be explained by ABO incompatibility * How could this happen? ELUTION Typically acid elution or Lui Freeze/Thaw elution Performed on baby's red blood cells Test elution with antibody screen and panel Add type specific cells if ABO incompatibility is suspected Why wouldn’t a panel detect A or B Antibody? careful with cell selection to conserve eluate sample PROBLEM SOLVING FOR FETUS/NEWBORN Severe signs of anemia Antibody screen negative DAT positive for IgG Eluate negative Causes? Further testing? Choice of RBC unit? FETAL BLEED TESTING RH NEG PARENT AND RH POS BABY Determine candidacy for RhIg Therapy Assess amount of bleed Optimal collection time is 1 hour post delivery – Why? Screening test Enumeration test Used to determine RhIg dosage ROSETTE TEST (FETAL BLEED SCREEN) Will detect >10mL of fetal blood Sample of parent’s blood Add CHEMICALLY MODIFIED ANTI-D NOT use your blood typing Anti-D Incubate Wash Add indicator cells (Immucor uses R2r, Ortho uses R2R2) Evaluate for rosettes using microscope FETAL BLEED SCREEN RESULTS Positive Perform enumeration test Negative dose of RhIg indicated Cannot use test if parent or baby is weak D pos Why? FETAL BLEED SCREENS ON THE MARKET Immucor: FMH RapidScreen detects 30mL of fetal blood Ortho: FetalScreen II Detects 7.5mL fetal red cells when ABO compatible IMMUCOR FMH RAPID SCREEN IMAGE ALTERED FOR SCREEN PROJECTION Original Settings altered KLEIHAUER BETKE (KB) Make a blood smear Acid bath destroys adult hemoglobin but not fetal hemoglobin Stain Fetal cells appear pink, adult cells appear as “ghosts” Count a total of 2000 cells Resulted as % of fetal cells Image credit: Pathology Student (https://www.pathologystudent.com/?s=kleihauer) IMAGE ALTERED FOR SCREEN PROJECTION Original Settings altered KB STAIN LIMITATIONS Not very precise Time consuming Staining issues Error between techs Fetal cells and lymphocytes could be confused with this stain technique parent has increased Hgb F levels, this test will be falsely elevated Can be used on Rh positive and Weak D positive parents RHIG ADMINISTRATION FLOW CYTOMETRY Anti-Hgb F Very precise Reproducible Expensive, requires special testing equipment Can be used for Rh positive and Weak D positive parents RHIG dose of RhIg can address 30mL whole blood bleed 15mL packed cells Needs to be given within 72 hours of sensitizing event and should not exceed 5 doses Can be given via IV or intramuscular injection Should be cleared within 6 months RHIG How does it work? Blocking/hindering Only 20% of sites can be blocked This theory has been debunked but may work in conjunction Immune Clearance RBCs are cleared out instead of stimulating immune system Not great since wherever RBCs are destroyed B cells are monitoring Immunostat Body thinks it already made antibody All three CALCULATING DOSE (# of fetal cells / # of Parent’s cells) * 100 * 50 = fetal bleed volume (Fetal Bleed Volume / 30) + 1 = Vials of RhIg to give Example: baby cells / 1980 Parent cells) * 100 * 50 = 50.5 30 = 1.7 + 1 = 3 vials of RhIg NOTE:  Mini Dose is indicated for <12 weeks pregnant and a single vial is sufficient for <20 weeks pregnant HEMOLYTIC DISEASE OF THE FETUS AND NEWBORN ABO Non-ABO Occurs on first pregnancy Yes No (unless antibody was developed prior) Immunoglobulin IgG IgG Antibodies of concern Anti-A Anti-B Anti-A,B Anti-D Anti-c Anti-E Anti-K (most significant after Rh) DAT Weak Pos or Neg Pos Microscopic findings Spherocytes – increased Reticulocytes – increased Nucleated RBCs – Increased Spherocytes – occasional Reticulocytes – increased Nucleated RBCs – Increased Sedimentation Rate (ESR) Increased Normal or slightly increased Bilirubin Slightly increased Greatly increased COMPARISON MONITORING OF BABY Anemia Bilirubin Prevent kernicterus Total bilirubin >25 mg/dl Bili-lights Bili-lights may not be effective if bilirubin rises at a rate of 0.5-2.0 mg/dL per hour Exchange transfusion Last resort when bili-lights fail Must be performed when serum bilirubin reaches 18-20mg/dL TRANSFUSING BABY Same blood requirements as IUT Pedi-packs or aliquots of extremely low birth weight infants and 58% of preterm infants < 32 weeks of gestational age receive red blood cell transfusions, mainly due to iatrogenic phlebotomy losses and ventilatory requirements. Typically performed as exchange transfusion – reconstituted RBC Not all facilities reconstitute the RBCs and Plasma due to waste this case, they issue a unit of RBCs and a unit of Plasma and it is up to the clinician to transfuse the correct ratio non-ABO HDFN, units must be tested crossmatch compatible Can use parent plasma for crossmatching if unable to obtain plasma from the baby IN SUMMARY Routine Prenatal and Postpartum testing on the mother for HDFN consist of ABO/Rh, Antibody Screen, and antibody identification if indicated. Titers are monitored throughout pregnancy if there is a clinically significant antibody. ABO HDFN and Non-ABO HDFN have different lab findings but the same effect on the baby There is a qualitative Fetomaternal Hemorrhage screen that can only be used on Rh Negative parents and there are two methods for quantifying the amount of the hemorrhage that can be used on any Rh. The number of RhIg of vials is calculated based on volume of Rh positive blood exposure Newborns are tested for ABORh forward type and IgG DAT. Units must be crossmatch compatible for non-HDFN. CASE STUDIES EXERCISE 1: IS PARENT A RHIG CANDIDATE? Parent: O NEG Antibody Screen: POS 1+ Antibody ID: anti-D (RhIg given at 28 weeks) Baby: A POS DAT: NEG Should FBS/FMH testing be performed: Yes FBS result: NEG Does the parent need RhIg? Yes EXERCISE 2: IS PARENT A RHIG CANDIDATE? Parent: O NEG Antibody Screen: NEG Baby: A NEG Weak D: POS 3+ DAT: NEG Should FBS/FMH testing be performed: No What testing should be done? Kleihauer Betke/Flow Cytometry Does the parent need RhIg? Yes EXERCISE 3: IS PARENT A RHIG CANDIDATE? Parent: O NEG Antibody Screen: POS Antibody ID: anti-E, -K Baby: A NEG Weak D: Not Tested DAT: 3+ Should FBS/FMH testing be performed: Unknown at this time What testing should be done? Weak D on EGA treated cells Does the parent need RhIg? Unknown pending Weak D results STORIES FROM THE BLOOD BANK are only as good as the information we get Mother Neg Positive antibody screen at delivery, RhIg administered at 28 weeks Baby Pos Neg DAT days old, baby’s bilirubin starts to go up Doctors start calling the blood bank asking about the work and for repeat testing Everything was done correctly; all results the same Doctor goes back to the mother for more information Mother fell down the stairs at 34w but didn’t notify her provider Developed anti-D late in pregnancy so there were no signs of fetal distress, mother's liver was filtering the bilirubin. DAT was negative because the cells were being lysed and cleared. Bilirubin started going up after delivery when baby no longer had mother's liver doing the work. SOURCES Howarth Claire, Banerjee Jayanta, Aladangady Narendra  “Red Blood Cell Transfusion in Preterm Infants: Current Evidence and Controversies” Neonatology 2018; 114:7-16 Quinley, Eva Immunohematology: Principles and Practice Lippencott Williams & Wilkins, 1998 Harmening,  Modern Blood Banking and Transfusion Practices F.A. Davis Company, 2005 Post, Ginell & Lazarchick, John & Singh, Zeba “Clinically Silent Massive Fetomaternal Hemorrhage: Important Lessons from an Illustrative Case” Laboratory hematology : official publication of the International Society for Laboratory Hematology 2012 18. 11-3. 10.1532/LH96.12002 Leon "Small Volume Transfusions in Neonates: Red Cells in Additive Solution are an Acceptable Choice" Vitalant (n.d.) https://www.vitalanthealth.org/getattachment/49628ee2-0dcd-40c1-9de4-3b090efb06d1/Neonate Transfusion.pdf