Automation to PreciseType and Everything in Between Jessie Singer MT(ASCP) Transfusion Medicine Children s Hospital Los Angeles
None Disclosures
Objectives Describe the application of molecular testing in the hospital donor center and transfusion service. Discuss the impact of molecular testing on serologic antibody identification workups through case studies. List the benefits of leveraging software to simultaneously view serology and molecular testing.
CHLA Overview 374 Inpatient Beds Level 1 Trauma Center CHLA has the largest pediatric hematology, oncology, and blood and marrow transplant program in the western United States Continuously ranked in top 10 in each category Rank in top 10 for pediatric cardiology Average 10 cardiac surgeries weekly Patient breakdown by ethnicity: African-American 5 % Asian 5 % Caucasian 19 % Latino 61 % Native American 0.08 % Other 9 %
CHLA Blood Donor Program Average whole blood collections >10,000/year 50% Mobile and 50% In-House collection sites Collect ~90% of blood required at CHLA All RBC units are serologically typed for C,E,K antigen
Targeted Blood Donation We are one of the few hospital-based transfusion services practicing Personalized Transfusion Medicine. By testing our patients and donors at the molecular level, we are able to Target our Collections (Donor Buddy Program) and meet our patients antigen specific transfusion needs. Member of Society for the Advancement of Blood Management Targeted Blood Donation program includes molecular testing on: CHLA Employees Donors within 5 mile radius of campus Frequent donors >2 times a year Minorities Improve blood collection efficiency by maintaining an optimal inventory of fresh blood products with minimal wastage Reduce total purchases of blood products
Need for Matched RBCs at CHLA Provide antigen matched blood to Infusion Center for chronically transfused patients (sickle cell, thalassemia) CHLA is the largest Sickle Cell center in California All patients matched for CEK at minimum Infusion Center requires ~5000 RBC units yearly Common Antibodies: c, Kpa, Wra, Cw, Jsa, Fyb, Jkb, e Growing RBC Exchange program (approximately 12 patients currently) Increases demand for CEK= and other specially phenotyped RBC units Additional Oncology patients Other surgical patients
Transfusion Service Methodologies Utilized Solid Phase (primary method) Echo (Immucor) NEO (Immucor) Gel LISS Reference lab for adsorption studies and incompatible crossmatches PreciseType RBC Phenotype
PreciseType HEA Multiplexed molecular assay that rapidly predicts genotype of 35 Human Erythrocyte antigens 24 polymorphisms associated with 35 RBC antigens Immucor, 2014
PreciseType BeadChip Array Overview Immucor, 2014
PreciseType Immucor, 2014
PreciseType Immucor, 2014
PreciseType Immucor, 2014
PreciseType Immucor, 2014
PreciseType Immucor, 2014
PreciseType Immucor, 2014
PreciseType Generated Report Kpa+ in 2% of population Lu(a+b+) 7.5 %
Molecular Phenotyping Patients Phenotype may be used to predict antigens most likely to cause alloimmunization Assist in antibody ID identification. Molecular phenotype provides whole picture on antibody ID and can be used to support or rule out suspected antibodies PreciseType performed on: Hematology patients (Sickle cell, Thalassemia) Recently transfused patients Patients with warm autoantibodies, positive DAT Patients with complicated antibody ID Multiple antibodies Antibodies to high or low frequency antigens Non-specific antibodies Bone marrow recipients
Molecular Phenotyping Donors Grow pool of phenotyped donors Closely match blood required for transfusion Expand pool of rare donors Reduce need to purchase special units from outside facility Screen for units in which no commercial antisera is available Increase recruitment efficiency Decrease need to perform sickle screen Investigate discrepancies in serological testing (patient phenotype, donor CEK typing)
c C e E V VS K k Kpa Kpb Jsa Jsb Fya Fyb Jka Jkb M N S s U Lua Lub Dia Dib Coa Cob Doa Dob Hy Joa LWa LWb Sc1 Sc2 HbS 121 Patients Typed 38 Gata Silencing Mutations 1 Fyb weak expression 1 Possible Hybrid C Allele CHLA Patient Population Snapshot (negative antigen expression) 100% 90% 80% 84% 83% 79% 97% 100% 92% 97% 96% 98% 100% 100% 70% 60% 60% 60% 50% 40% 30% 20% 10% 0% 46% 36% 36% 36% 36% 30% 21% 19% 17% 10% 1% 4% 0% 0% 0% 1% 0% 0% 0% 2% 2% 0% 0%
CHLA Donor Population Snapshot (negative antigen expression) >200 donors phenotyped 10/2016 to 12/2017 15 Gata Silencing Mutations detected 2 Possible Hybrid C Allele 100% 96% 95% 94% 98% 97% 99% 95% 95% 100% 100% 90% 80% 78% 70% 60% 52% 50% 40% 30% 20% 19% 41% 35% 31% 27% 27% 26% 23% 40% 17% 10% 0% 1% 0% 0% 0% 6% 0% 0% 0% 0% 0% 0% 0% 0%
c C e E V VS K k Kpa Kpb Jsa Jsb Fya Fyb Jka Jkb M N S s U Lua Lub Dia Dib Coa Cob Doa Dob Hy Joa LWa LWb Sc1 Sc2 c C e E V VS K k Kpa Kpb Jsa Jsb Fya Fyb Jka Jkb M N S s U Lua Lub Dia Dib Coa Cob Doa Dob Hy Joa LWa LWb Sc1 Sc2 HbS Patients 100% 90% 80% 70% 60% 50% 40% 30% 20% 10% 0% 21% 36% 1% Tested Populations Compared 79% 84% 83% 97% 0% 100% 0% 92% 0% 36% 30% 19% 36% 17% 36% 60% 4% 1% 97% 0% 96% 0% 0% 98% 46% 10% 2% 2% 0% 100% 0% 100% 60% Donors 100% 90% 80% 70% 60% 50% 40% 30% 20% 10% 0% 96% 95% 98% 94% 97% 99% 95% 95% 100% 100% 78% 52% 41% 40% 27% 31% 26% 27% 35% 23% 19% 17% 1% 0% 0% 0% 6% 0% 0% 0% 0% 0% 0% 0% 0% Patient Population: 19 % Caucasian, 61% Latino, 5% African American Donor Population: 45 % Caucasian, 35% Latino, 3% African American
Example of Impact to Donor Recruitment Program 19 year old sickle cell patient added to RBC Exchange program November 2017 B+, E Negative Anti-Fya (~19% population is type B or O and negative for Fya) Requires 6 units every 4 weeks Compatible donors Historical pool of 70 B and O donors with visits since 2015 Since live with PreciseType (10/2016), added 55 Fya= B or O donors to recruitment pool
Patient Workups: Case Studies
Case Study 1 IP 10 year old male Presented to CHLA with scleral icterus No known transfusion history Hemoglobin 9.6
Case Study 1 IP (Continued) Initial Presentation: Unable to type ABORH due to panagglutination DTT not routinely performed at CHLA Solid Phase: All cells 3+ Gel Screen/ Panel: All cells 4+ LISS Screen: All cells 2+ AHG DAT: Strong Positive IGG, Poly, C3b/C3d
Case Study 1 IP (Continued) Sent to Reference Lab Patient typed as O Positive Probable Auto anti-e detected via warm and cold adsorbed serum IAT methodology Diagnosed with Warm and Cold Autoimmune Hemolytic Anemia Discharged without transfusion
Case Study 1 IP (Continued) 3 months later Presentation: Hemoglobin 7.2 Presents with increased jaundice and back pain Workup Gel screen/panel: All 3+ and 4+ LISS screen: All cells 2+ AHG, autocontrol 3+ DAT: IgG, Poly, C3b/C3d all 3+ Reference Lab: Rouleax and cold autoagglutinins @ RT. Anti-D detected (presumed autoantibody) Recommended to transfuse Rh Negative No transfusion required
Case Study 1 IP (Continued) 3rd presentation: Autoanti-E and Autoanti-D not demonstrable Gel Panel: all cells 3+ Liss Screen: all cells 2+ AHG PreciseType performed: Decision made to crossmatch AHGcompatible units negative for K, Jka, Fyb until they can be ruled out in LISS Outcome/ Follow-up: Patient diagnosed with Evans Syndrome, controlled predominately by rituximab Has received 5 RBCs at CHLA LISS Screen negative 2 years following initial presentation
Case Study 2 AM 16 year old male History of autoimmune hemolytic anemia and warm autoantibodies Presented with fatigue and darkened urine No known transfusion history Hemoglobin 9.0
Initial Workup Case Study 2 AM (Continued) ABO/RH: O Positive Solid Phase Screen: All cells 4+ Solid Phase Panel: All cells reactive (varying reactions) LISS Screen: All cells 2+ AHG, autocontrol 2+ DAT: Poly 4+, IgG 3+, C3b/C3d 2+ Patient discharged without transfusion, specimen not sent to reference lab for adsorption study
Case Study 2 AM (Continued) 6 months post initial workup Presentation: Presents with scleral icterus, fatigue, lightheadedness, and dark urine Hemoglobin: 4.2 Workup: Probable warm autoantibody Solid phase: all cells reactive LISS: all cells 2+ AHG DAT: 4+ Poly, 4+ IgG, 3+ C3b/C3d 2 units urgently required Crossmatched two least incompatible CEK negative RBCs
Case Study 2 AM (Continued) Reference Lab: Acid eluate of RBCs treated with chloroquine reacted strongly with all RBCs by PEG IAT Adsorbed Serum contained: Anti-E, reactive at RT, 37C, and LISS AGT Anti-c reactive by LISS AGT Anti-S reactive by LISS AGT Probable warm autoantibodies
Case Study 2 AM (Continued) PreciseType Performed Confirmed negative for E, c, S Conclusion: Crossmatch least incompatible E-, c-, S-, K- RBCs Follow up: Patient s warm Idiopathic AIHA controlled with steroids
Case Study 3 ES ES is a 30 year old Asian male with Thalassemia major Followed and transfused at CHLA since birth Transfused with 2-3 RBCs every 3 weeks ABO/RH: O Positive Special Needs: E negative, K negative RBCs No known antibodies
Workup Case Study 3 ES (continued) Solid Phase screen: Cells 1 and 2 are 2+ reactive Gel Screen: all cells negative Solid Phase Panel: Jkb suspected (known to react earlier in solid-phase)
Case Study 3 ES (continued) Specimen sent to reference lab DAT positive in IGG and complement Acid elution non reactive Serum contained no unexpected RBC antibodies Anti-Jkb not confirmed, reference did not test by solid phase at the time Recommended to conservatively select Jkb negative units for crossmatch
Case Study 3 PreciseType predicts sample to be Jkb+ Rules out suspected anti- Jkb ; variant testing could be considered Follow up: Patient has since shown other non-specific reactivity on occasional visits. Warm-autoantibody suspected.
12 year old female Case Study 4 TA History of ITP and family history of thalassemia No transfusion history Presented at outside hospital with anemia, lethargy, jaundice, and dark urine Hgb 4.1
Case Study 4 TA (Continued) Initial workup ABO/ Rh: A Positive Gel Screen/Panel: All Cells 3+ Solid Phase Screen (Echo): All cells 4+ LISS Screen: All Cells 3+ Eluate: All cells 3+ in gel DAT: Poly 3+, IgG +, c3b/c3d negative Reference Lab Serum contained warm autoantibodies reactive by LISS, PEG, and ficin AGT
Case Study 4 TA (continued) PreciseType Patient is negative for K, Fya, Jkb Decision made to conservatively crossmatch units that are K, Fya, and Jkb negative and least incompatible with patient s unadsorbed serum at AHG Received 11 RBCs during stay in 2015 Discharged with hemoglobin 7.3
Case Study 4 TA (continued) Patient returns 2018 (almost 3 years post- initial presentation) Hgb 6.5 Solid Phase Screen (NEO) : DAT: Poly 3+, IgG 3+, C3b/C3d negative
Case Study 4 TA (continued) NEO Autocontrol 3+ E, c not ruled out Anti-S ruled out on additional panel
Eluate: All cells 3+ or 4+ Strong warm autoantibody Case Study 4 TA (continued) Only antigens not ruled out in multiple panels: E, c Patient phenotype E+ c+ Add more panels/ liss/gel Conclusion: autoanti-e and autoanti-c Transfusion requirements: AHG compatible E-neg, c-neg (while Auto-E and Auto-c presenting) and K-neg RBCs Follow Up:
7 month old male Case Study 5 JM History of biliary atresia admitted to CHLA with increased abdominal growth Lab results consistent with liver failure and added to liver transplant list Hgb: 7.0 Patient had received one aliquot of RBCs at outside hospital 3 weeks prior
Workup Case Study 5 JM (continued) ABO/RH: O Positive Solid Phase Screen (NEO): cell 3 is 1+ need to rule out K, Jkb, Leb Gel Screen: All cells negative
Case Study 5 JM (continued) Autocontrol and DAT Negative Solid phase panel: All antigens ruled out, anti-jkb still suspected. Added special instruction to crossmatch Jkb negative RBCs (1 RBC transfused)
Case Study 5 JM (Continued) Anti-Jkb prediction supported with PreciseType
Case Study 5 JM (Continued) Follow up visit Solid Phase panel:? on almost all homozygous positive Jkb cells Solid Phase Panel
Case Study 5 JM (continued) Gel Screen: w+ in 3 cell Gel Panel: w+ in all homozygous positive Jkb cells Autocontrol: 2+, no antibody eluted
Case Study 5 JM (continued) Conclusion: Anti-Jkb predicted earlier through solid phase and supported by PreciseType Follow Up: Patient received liver transplant 2 Jkb negative units transfused in OR, 3 aliquots post-op Transfusion required 4 months after transplant, anti-jkb not demonstrable
Leveraging software to simultaneously view serology and molecular testing
Blood Bank LIS Our blood bank LIS, SafeTrace TX displays antibodies and antigens collectively in the patient profile. PreciseType/BASIS does not interface directly with our Blood Bank LIS and Donor Services LIS Molecular phenotype antigens must be manually resulted as a test batch for 35 antigens in both LIS software. Manual entry eliminated with leveraging software, ImmuLINK
Addition of ImmuLINK to workflow Benefits of a Leveraging Software Transmit automated CEK and molecular antigen testing results via automation to donor and transfusion service LIS Reduce transcription error from manual entry Reduce tech time of antigen result entry and review Easily view all serology and molecular testing results for patients to provide full picture of patient workup and history Print reports with panel images and reactions for all ordered tests on a specimen (including Antibody ID and Molecular testing)
Report Example #1
Molecular and Serology displayed on one report Report Example #2
Future plan: Continue to grow inventory of donors with historical molecular phenotype Grow targeted blood donor program Build dashboards for snapshot of screened units in inventory Further match patient transfusion needs to proactively prevent alloimmunization in hypertransfused patients
Questions?