Future Webinars. Future Webinars

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1 Continuing Education Webinar Series Future Webinars Future Webinars 24 May Donor Selection for Hematopoietic Stem Cell Transplantation featuring Dr Deborah Sage, NHSBT Tooting London, UK 27 June Transfusion Reactions, Part II TRALI, TACO, and GVHD featuring Dr Margo Rollins, Children s Healthcare of Atlanta Atlanta, Georgia, USA Link to register:

2 Handouts Program Handouts.aspx Continuing Education PACE, Florida and California DHS 1.0 Contact Hours Each attendee must register to receive CE at: Registration deadline is May 11, 2018 Certificates will be sent via only to those who have registered by May 25, 2018 Presentation Recording Session will be recorded and posted. Access information will be sent to each registrant when the recording becomes available CE credits will be issued to anyone who listens to the recording within one year of the original presentation date (today). Learn website: learn.immucor.com

3 learn.immucor.com Course content is for information and illustration purposes only. Immucor makes no representation or warranties about the accuracy or reliability of the information presented, and this information is not to be used for clinical or maintenance evaluations. The opinions contained in this presentation are those of the presenter and do not necessarily reflect those of Immucor. Interactive Webinar

4 Super Powers? Sue Johnson, MSTM, MT(ASCP)SBB Director, Clinical Education BloodCenter of Wisconsin, part of Versiti Milwaukee, WI Patient History 68 y/o Caucasian male No transfusion at your facility Just returned from vacation out of state Received 1 LRBC at local community hospital Admitted with 6.8 g/dl Hb Diagnosis CLL 2 u RBCs ordered for outpatient transfusion tomorrow Pretransfusion Testing Anti A Anti B Anti D A1 cells B cells Rh Kell Duffy Kidd Lewis P MNS Lu Xg Cell D C c E e f * V C w K k Kp a Kp b J s a J s b Fy a Fy b Jk a Jk b Le a Le b P1 M N S s Lu a Lu b Xg a Echo Pos Cntrl 4+ Neg Cntrl 0

5 Antibody Identification Panel SPRCA Rh Kell Duffy Kidd Lewis P MNS Lu Xg Cell D C c E e f * V C w K k Kp a Kp b J s a J s b Fy a Fy b Jk a Jk b Le a Le b P1 M N S s Lu a Lu b Xg a Echo Pos Cntrl 4+ Neg Cntrl 0 Antibody Identification Gel Rh K LE FY Kidd MNS Gel Fy b Jk a Jk b M N S s IAT D C E c e K Le a Le b Fy a I II III Auto 4+ What test would you perform next? 1. Direct Antiglobulin Test (DAT) 2. Autologous or Allogeneic Adsorption 3. Saline IAT in Tubes 4. LISS IAT in Tubes 5. Enzyme Treated RBC IAT

6 Direct Antiglobulin Test Results DAT Polyspecific AHG 4+ Anti IgG 4+ Anti C3 3+ Control 0 What test would you perform next? 1. Autologous or Allogeneic Adsorption 2. Saline IAT in Tubes 3. LISS IAT in Tubes 4. Enzyme Treated RBC IAT 5. Elution What should be done next? Adsorption Detect underlying antibodies Saline or LISS IAT Decreased sensitivity for autoantibodies, may not take Superman powers Enzyme Treated RBCs Aid in defining specificity evidence that adsorption will work if we go there Elution Confirm warm autoantibody vs. DIIHA

7 Antibody Identification Saline IAT Rh K LE FY Kidd MNS Saline Fy b Jk a Jk b M N S s IS 37C IAT D C E c e K Le a Le b Fy a I II III Auto What test would you perform next? 1. Test enzyme treated RBCs 2. Autologous or Allogeneic Adsorption 3. Prewarm & Cold Screen/Panel 4. Give least incompatible blood 5. Send it to the IRL 6. Genotype & give phenotypematched RBCs What next? Test Enzyme Treated RBCs Aid in defining specificity evidence that adsorption will work Autologous or Allogeneic Adsorption Detect underlying antibodies Prewarm & Cold Screen reactivity may be due to cold reactive antibody??

8 What next? Give least incompatible Especially if no history of transfusion In consultation with medical director Send to IRL to confirm & rule out alloantibodies Let them do the work RBC Genotype & provide antigen matched blood Genotype Patient Transfusion request is urgent Perform adsorptions 1 st time Determine if any underlying alloantibodies are present Subsequent transfusions provide phenotype/genotype matched RBCs Antibody Identification Enzyme Treated RBCs Rh K LE FY Kidd MNS Saline Fy b Jk a Jk b M N S s IS 37C IAT D C E c e K Le a Le b Fy a I II III Auto

9 Autologous vs. Allogeneic Adsorption Transfusion history is key Recent in last 3 months Ever? Availability of patient s RBCs Patient s Hgb Reference Underlying Alloantibodies No. of Antibodies/ No. of Sera Tested % of Sera with Alloantibodies Morel 8/20 40 Branch and Petz 5/14 36 Wallhermfechtel et al 19/ Laine and Beattie 41/ James et al 13/41 38 Issitt et al 13/34 38 (alloadsorptions) Issitt et al 5/41 12 (autoadsorptions) Leger and Garratty 105/ Maley et al - UK 39/ Das and Chaudhary - 7/23 30 India TOTALS 255/796 32% Allogeneic Adsorption Patient s Phenotype is Unknown RBC Phenotype Antibodies Remaining R 1 R 1 ; Jk(a-); s- -c, -E, -Jk a, -s R 2 R 2 ; Jk(b-); S- -e, -C, -Jk b,-s rr; K- -D, -C, -E, -K

10 Allogeneic Adsorption Using Ficin Treated RBCs Pt Serum + R 1 R 1, Jk(a-) RBCs Pt Serum + R 2 R 2, Jk(a+) RBCs Pt Serum + rr, Jk(a-) RBCs = Anti-E = WAA = E+ RBCs Allogeneic Adsorption After 37C Incubation Pt Serum + R 1 R 1, Jk(a-) RBCs Pt Serum + R 2 R 2, Jk(a+) RBCs Pt Serum + rr, Jk(a-) RBCs = Anti-E = E+ RBCs Testing Serum Alloadsorbed x3 with Ficin Treated RBCs R 1R 1 R 2R 2 rr Rh K LE FY Kidd MNS Saline Fy b Jk a Jk b M N S s IAT IAT IAT D C E c e K Le a Le b Fy a I II III Phenotype of Adsorbing RBCs R 1 R 1, K, Jk(a ), s R 2 R 2 rr, K, Jk(b )

11 Alloadsorbed x3 with c E, K, Jk(a ), s Ficin Treated RBCs Rh K LE FY Kidd MNS Saline D C E c e K Le a Le b Fy a Fy b Jk a Jk b M N S s Adsorbing IAT RBCs I II III Alloadsorbed x3 with C e RBCs Ficin Treated RBCs Rh K LE FY Kidd MNS Saline D C E c e K Le a Le b Fy a Fy b Jk a Jk b M N S s Adsorbing IAT RBCs I II III Ruled out anti C, anti e, anti Le a & anti Fy b Alloadsorbed x3 with C E, K, Jk(b ) Ficin Treated RBCs Rh K LE FY Kidd MNS Saline D C E c e K Le a Le b Fy a Fy b Jk a Jk b M N S s Adsorbing IAT RBCs I II III Ruled out anti D, anti E, anti K, anti Le b & anti Jk b

12 Summary of Rule Outs on Alloadsorbed Serum Rh K LE FY Kidd MNS Saline Fy b Jk a Jk b M N S s IAT IAT IAT D C E c e K Le a Le b Fy a I II III Unable to rule out anti c, anti Jk a & anti s Selected Cells Tested with Alloadsorbed x3 with c E, K, Jk(a ), s RBCs Rh K LE FY Kidd MNS Saline D C E c e K Le a Le b Fy a Fy b Jk a Jk b M N S s Adsorbing IAT RBCs Underlying anti Jk a identified Anti c and anti s ruled out Would you do an eluate? 1. Yes, we always save or obtain RBCs 2. Yes, only if RBCs are left after adsorption 3. No 4. Other

13 Yes! Should you do an eluate? RBC sample size Acid eluate requires just drops Current draw is preferred Confirms warm autoantibody Identifies possible DIIHA 1 st time patient is seen Doesn t need to be repeated on subsequent samples ELUATE Eluate Last Wash D C c E e K Fy a Fy b Jk a Jk b S s IAT IAT Auto NT Positive with all panel cells, consistent with warm autoantibody coating the patient RBCs Super Powers Case Summary Warm autoantibody reactive by SPRCA, Gel, Saline IAT, Ficin IAT Warm autoantibody showed broad specificity reacting with all RBCs tested DAT positive due to IgG and C3 coating Eluate positive with all cells tested Consistent with WAIHA, need to correlate with patient s clinical condition

14 Something Just Like This by Coldplay, The Chainsmokers I've been reading books of old The legends and the myths The testaments they told The moon and its eclipse And Superman unrolls A suit before he lifts But I'm not the kind of person that it fits Super Powers Case Summary The forces of ruling out on alloadsorbed serum did not stop us from identifying an underlying anti Jk a! Questions? sue.johnson@bcw.edu

15 Justice for Anti-M Monica Kalvelage, aka Green Lantern Our oath In brightest day and blackest night no evil shall escape my sight. Let those who worship evil s might beware my power green lantern s light. In brightest day and blackest night no antibody shall escape my sight. Let those who defy serology s light beware the power blood banker s might. Initial BB Testing 26 year old female Routine clinic visit, pregnant H&H = 12%, 36g/dl Positive Antibody Detection Test

16 ABO/Rh tube testing Forward Reverse Anti-A Anti-B Anti-D Rh Control A1 A2 B pt Antibody Detection Test - solid phase D C E c e M N S s K k Fy a Fy b Jk a Jk b IAT R 1R R 2R What do the ABO/Rh and antibody screen indicate? 1. Looks like warm reactive antibody. 2. Looks like cold reactive antibody. 3. Possible combination of things. 4. I ve seen enough. Send the sample to the IRL. Antibody Identification Test - solid phase Rh MNS LU P Lewis Kell Duffy Kidd SP Lot 567 D C E c e V M N S s Lu a Lu b P 1 Le a Le b K k Fy a Fy b Jk a Jk b IAT RzR R1Wr R2R Ror r r R2r rr rr rr r r rr rr rr R1R Pos Ctrl 4+ Neg Ctrl 0

17 Antibody Identification Test - solid phase Rh MNS LU P Lewis Kell Duffy Kidd SP Lot 567 D C E c e V M N S s Lu a Lu b P 1 Le a Le b K k Fy a Fy b Jk a Jk b IAT RzR R1Wr R2R Ror r r R2r rr rr rr r r rr rr rr R1R Pos Ctrl 4+ Neg Ctrl 0 Based on the panel results what would your facility do next? 1. Switch to tubes to evaluate all phases. 2. Phenotype for M and N. 3. Perform an antibody titration. 4. Run a selected cell panel in another method. 5. Other Antibody Identification Test, tube testingube testing Rh MNS P Lewis Kell Duffy Kidd Lot 123 D C E c e V M N S s P 1 Le a Le b K k Fy a Fy b Jk a Jk b I.S. 37C IAT Prewarm RzR R1Wr R2R Ror r r R1r rr rr R2r R0r rr A/C NOTE: Run additional M- selected cells to complete rule-outs of other antibodies.

18 What guides your next decision? 1. Facility SOP and policy 2. Technical Manual 3. Other reference material 4. Ask your coworker 5. Call the supervisor 6. Call your Medical Director Do you have these books? What do thiol reagents do? e.g. 2-ME (mercaptoethanol) or DTT (dithiothreitol) 1. Disrupt Antibody-Antigen binding. 2. Destroy IgM by breaking disulfide bonds. 3. Destroy IgG by breaking trisulfide bonds. 4. Inhibit the power of green lantern s ring.

19 Review disulfide bonds and thiol agents IgM is held together by S-S bonds S-S also found in some RBC Ag and Ab Thiol agents break the bonds Abozenadah, H., Bishop, A., Bittner, S., Lopez, O., Wiley, C., and Flatt, P.M. (2017) Consumer Chemistry: How Organic Chemistry Impacts Our Lives. Accessed at ME treatment and interpretation Rh MNS Lot 886 D C E c e V M N S s Control, untreated (IAT) 2-ME treated control (IAT) Patient, untreated (IAT) 2-ME treated patient serum (IAT) Ror r r wk R2r rr rr What are some options for additional testing if your facility does not have chemicals? 1. No enhancement, 37C IAT only 2. Pre-warm IAT 3. Titration 4. Other

20 Titration studies of anti-m Titration RBC: M+N+ Neat Pt serum The case is not finished! We must resolve the ABO discrepancy. How? Some ideas: Pre-warm It looks like a group A, so report as group A Reverse type with M Negative cells Shake the A1 and A2 reverse cells harder Reverse type with Ficin-treated cells Considerations on reporting these case findings: Why is Solid Phase 4+ if the Ab is mostly IgM? Why does pre-warm not remove reactivity? Is it really reactive at 37 or just has such a high titer that it overwhelms the system? Perhaps IgM component > IgG component. Can clinical significance be accurately assessed?

21 Final reporting Testing indicates this anti-m has both IgG and IgM components. Recommend continued monitoring of this pregnant patient. Wonderful Wonder Woman Jayanna Slayten, MS, MT(ASCP)SBB cm Blood Bank Supervisor, Indiana University Health Adjunct Faculty, University of Texas Medical Branch SBB Program Patient History A 35 year old female patient has arrived in preadmission testing for labs to be drawn anticipating a knee surgery in two weeks. She has never been transfused She has 8 children.. Which makes her a Physician has ordered CBC BMP Type and Screen

22 Current Labs CBC Normal BMP Normal, no flags Type and Screen A B D A1Cell B Cell Interpretation NTD Rh Kell Duffy Kidd Lewis P MNS Lu Xg Cell D C c E e f * V Cw K k Kpa Kpb J sa J sb Fya Fyb Jka Jkb Lea Leb P1 M N S s Lua Lub Xga Echo Pos Cntrl 4+ Neg Cntrl 0 What is your next step? Option 1: Run a Panel in Echo Option 2: Change method with Antibody Screen (ABS) Option 3: Change method with ABS and run Echo Panel Option 4: Send to Reference Lab Initial Troubleshooting Review Option 1: Run a Panel in Echo ABS positive would reflex to a Panel testing Option 2: Change method with ABS Good possibility. However, what is in Echo? Option 3: Change method with ABS and run Echo Panel Good choice as it covers both troubleshooting options Option 4: Send to Reference Lab Always a good option

23 Wonder Woman Capture-R Ready-ID *= presumptive Rh Kell Duffy Kidd Lewis P MNS Lu Xg Cell D C c E e f * V Cw K k Kpa Kpb J sa J sb Fya Fyb Jka Jkb Lea Leb P1 M N S s Lua Lub Xga Echo Pos 4+ Neg Cntrl 0 Case B Cntrl 4/16/ Wonder Woman Panoscreen *= presumptive Rh Kell Duffy Kidd Lewis P MNS Lu Xg Cell D C c E e f * V Cw K k Kpa Kpb Jsa Jsb Fya Fyb Jka Jkb Lea Leb P1 M N S s Lua Lub Xga IS PEG Autocontrol 2+ 0 IgG-DAT is negative Complement DAT is weakly positive 4/16/ Wondering What Next? Option 1: Another Echo Panel (I or II) Option 2: Panel in PEG-AHG Option 3: Panel in LISS-AHG Option 4: ABS with saline-no enhancement (37C-AHG) Option 5: Cold - ABS with autocontrol (IS, Room Temperature and 4C) Option 6: Send to Reference Lab

24 Wondering.. Option 1: Another Echo Panel (I or II) Would look the same as the initial panel Option 2: Panel in PEG-AHG Option 3: Panel in LISS-AHG Would look the same as the initial antibody screen Option 4: ABS with saline-no enhancement (37C-AHG) Good possibility due to the IS reactions Option 5: Cold - ABS with autocontrol (IS, Room Temperature and 4C) Good possibility due to the IS reactions Option 6: Send to Reference Lab Always a possibility! Wonderful Wonder Woman Panoscreen *= presumptive Rh Kell Duffy Kidd Lewis P MNS Lu Xg IS RT 4C Cell D C c E e f * V Cw K k KpaKpb Jsa Jsb FyaFybJkaJkbLeaLeb P1 M N S s Lua Lub Xga Autocontrol Confirmed as a cold autoantibody IgM reactivity verified IgG reactivity has not been identified yet 4/16/ Wonderful Wonder Woman Panoscreen *= presumptive No Rh Kell Duffy Kidd Lewis P MNS Lu Xg Enhancement Cell D C c E e f * V Cw K k KpaKpb Jsa Jsb FyaFybJkaJkbLeaLeb P1 M N S s Lua Lub Xga IS 37C AHG Unable to rule out D, C, E, K, Fy a, Jk a, Jk b, Le b, P 1, N and s The cold autoantibody detected at IS but not at 37C An underlying alloantibody is detected on Screen Cell 1 Autocontrol /16/

25 Wonder Woman 37C-AHG Panel Cell D C c E e f V C w Rh Kell Duffy Kidd MNS P Lewis No Enhancement K K k F y a F J J M N S s P L L 37C AHG y k k 1 e e b a b a b Anti K at AHG 4/16/ The patient ABO Red Cell Types A, but ABO Serum Types O. How do you resolve? Option 1: Report as No Type Determined (NTD) and give group O Option 2: Test with C antigen negative A1 cells and B cells, due to anti C Option 3: Repeat ABO Serum Typing with prewarm AHG method Option 4: Repeat ABO Serum Typing with with 37C AHG The patient ABO Red Cell Types A, but ABO Serum Types O. How do you resolve? Option 1: NTD Possible in the short term, but try and resolve Option 2: Test with C antigen negative Not necessary, anti C is IgG and ABO serum typing is direct/igm testing Option 3: Repeat ABO Serum Typing with prewarm AHG method Option 4: Repeat ABO Serum Typing with with 37C AHG Both good possibilities

26 Wonderful Warm ABO Serum Type Prewarm AHG ABO Serum Typing ABO Red Cell Typing A1 Cells B Cells Interpretation Group A NTD Not wonderful but woeful. IgG gets in the way 37C testing ABO Serum Typing ABO Red Cell Typing A1 Cells B Cells Interpretation Group A 0 4+ Group A ABO now Wonderful! Wonder Woman Finale The forces of IgG and the bitter COLD did not stop this case from being cracked nor the patient from getting her knee surgery and get back to fighting the Evil in the World Anti K at IgG Cold Autoantibody ABO Resolution with 37C Testing We like you! Like us on social media!

27 Continuing Education PACE, Florida and California DHS 1.0 Contact Hours Each attendee must register to receive CE at: Registration deadline is May 11, 2018 Certificates will be sent via only to those who have registered by May 25, 2018 Future Webinars 24 May Donor Selection for Hematopoietic Stem Cell Transplantation featuring Dr Deborah Sage, NHSBT Tooting London, UK 27 June Transfusion Reactions, Part II TRALI, TACO, and GVHD featuring Dr Margo Rollins, Children s Healthcare of Atlanta Atlanta, Georgia, USA Link to register: Future Webinars

28 Thank you!