Serological Cases: What keeps us up at NightStephanie Goe, MT (AAB-P)

June 6, 2013

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Solving Problem Workups

• Unusual and rare antibodies present complex and challenging serological workups for the blood banker. This can be due to lack of experience, exposure and appropriate reagents.

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Dilemma.. “How do you know what you don’t know”

• How does one identify an antibody they have never seen ???

• Some antibodies react similarly. However, they can be completely benign, capable of producing a mild transfusion reaction or death.

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Goals

• The objectives of this case study are:

1. Presentation of a workup involving a clinically significant high incidence antibody Patient Information Serological techniques used for identification Challenges of finding compatible units

2. Compare and contrast a clinically significant and nonsignificant antibody

3. Discuss and show that “Antibodies don’t react by the book”

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Case Study

• Patient Information– 54 year old Caucasian female– Diagnosis: acute kidney injury, rhabdomylosis, multiple

traumas– Pregnancy and transfusion history unknown– Referring hospital transfused 6 rbcs as follows: * 2 rbcs on 2/26/13- DAT negative; no antibodies identified * 2 rbcs on 2/28/13 * 2 rbs on 03/15/2013- DAT negative; anti-c,-E, cold autoantibody

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What now? Specimen sent to IRL

• We rec’d both pre and post transfusion samples 10 days after patient initially presented to referring hospital

• All samples had substantial hemolysis ranging from 600 to 3000 mg % free hemoglobin

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Hmmm…How to proceed?

• Pretransfusion sample:* ABO/Rh: O Positive

* DAT: Negative with polyspecific AHG * Used to phenotype for common antigens *Patient phenotype:

C+,E-,c-,e+,M+,N+,S-,s+,K+,k+,Fy(a-b+),Jk(a+b+)

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Initial Testing

• Post Transfusion Specimen: * ABO/Rh: O Positive * DAT: w+ with polyspecific AHG & complement * Specimen grossly hemolyzed

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Initial Testing

Initial selected cell panel run in PEG which included a phenotypically similar cell.

IS results all 3+ except for the autocontrol

Reactions at IgG ranged from w+ to 1+

Panel tested in LISS yielded similar results

Initial Panels   

 LISS

  

PEGRh system Kell Duffy Kidd MNSs   IS 3

7o

C

IgG

IgG

  D C c E e K k Fya Fyb Jka Jkb M N S s          

1 + + 0 0 + + + 0 + + 0 + + + +   3+ 2+ 1+ 3+

2 + + 0 0 + 0 + + 0 0 + 0 + + 0   3+ 2+ 1+ 3+

3 + + 0 0 + 0 + 0 + + 0 + 0 0 +   3+ 2+ 1+ 3+

4 + + 0 0 + + + 0 + + 0 0 + 0 + P.S. 3+ 2+ 1+ 3+

AC                                 0 0 0 0✓ 

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Now what?

• Similar strong IS reactivity with IgG carryover in panel cells

• Phenotypically similar cell reactive• Hx of cold autoantibody; however,

negative autocontrol! • Cold screen performed

Cold Screen Results

4C 37C

SC1 4+ 3+

SC2 4+ 3+

AC 0 0

I Neg 4+ 3+

I Neg 4+ 3+

Information Gained

• A cold reacting antibody present. However, not a cold autoantibody. Autocontrol is negative.

Panel with 0.01M DTT Treated Serum

    

LISSTest

Rh system Kell Duffy Kidd MNSs   IS

37o

C

IgG

  D C c E e K k Fya Fyb Jka Jkb M N S s        

1 + + 0 0 + + + 0 + + 0 + + + +   0 0 1+

2 + + 0 0 + 0 + + 0 0 + 0 + + 0   0 0 1+

3 + + 0 0 + 0 + 0 + + 0 + 0 0 +   0 0 1+

4 + + 0 0 + + + 0 + + 0 0 + 0 + P.S. 0 0 1+

AC                                 0 0 0✓ 

Panel with Control Serum

    

LISSTest

Rh system Kell Duffy Kidd MNSs   IS 37o

C

IgG

  D C c E e K k Fya Fyb Jka Jkb M N S s        

1 + + 0 0 + + + 0 + + 0 + + + +   3+ 2+ 2+

2 + + 0 0 + 0 + + 0 0 + 0 + + 0   3+ 2+ 2+

3 + + 0 0 + 0 + 0 + + 0 + 0 0 +   3+ 2+ 2+

4 + + 0 0 + + + 0 + + 0 0 + 0 + P.S. 3+ 2+ 2+

AC                                 0 0 0✓ 

Panel with 0.2M DTT Treated Cells

    

LISSTest

Rh system Kell Duffy Kidd MNSs   IS 37o

C

IgG

  D C c E e K k Fya Fyb Jka Jkb M N S s        

1 + + 0 0 + + + 0 + + 0 + + + +   1+ 1+ 1+

2 + + 0 0 + 0 + + 0 0 + 0 + + 0   1+ 1+ w+

3 + + 0 0 + 0 + 0 + + 0 + 0 0 +   1+ 1+ 1+

4 + + 0 0 + + + 0 + + 0 0 + 0 + P.S. 1+ 1+ 1+

AC                                 0 0 0✓ 

Recap

• IgM reactivity abolished with 0.01M DTT treated serum (IgG component still present)

• Diminished reactivity with 0.2M DTT treated cells • Reactive phenotypically similar cell

• Negative autocontrol ………Possible high??

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What are Some Candidates?

• Patient is Caucasian• IS reactions• Diminished reactivity with 0.2M DTT

treated cells

Possible PP1Pk, I, H, Vel ?

Panel Reflecting R2R2 2x PEG Adsorbed Sera Results

    

Rh system Kell Duffy Kidd MNSs   R2R2

  D C c E e K k Fya Fyb Jka Jkb M N S s    

1 + + 0 0 + + + 0 + + 0 + + + +   0✓ 

2 + + 0 0 + 0 + + 0 0 + 0 + + 0   0✓

3 + + 0 0 + 0 + 0 + + 0 + 0 0 +   0✓

4 + + 0 0 + + + 0 + + 0 0 + 0 + P.S. 0✓

Differential Cells

R1R1

P1+; M+N-S+s-;K-; Fy(a+b-); Jk(a-b+)

R2R2

P1+; M-N+S-s+;K-; Fy(a-b+); Jk(a+b+)

rrP1-; M+N+S-s+;K-; Fy(a-b+); Jk(a+b-)

Additional Phenotyping

• Patient typed with the following antisera:Anti-H -P1 -I -Vel

Pos Pos Pos Neg

Antigen Negative Cells Tested

    

LISS 

  

PEG

Rh system Kell Duffy Kidd MNSs   IS

37o

C

IgG

IgG

  D C c E e K k Fya

Fyb

Jka Jkb M N S s Vel        

1 + + 0 0 + + + + + + 0 + + + + 0 0 0 0  0✓

2 + + 0 0 + 0 + + 0 0 + 0 + + 0 0 0 0 0 0✓

Prewarm Panel   

Prewarm 

 Rh system Kell Duffy Kidd MNSs  

IgG 

  D C c E e K k Fya Fyb Jka Jkb M N S s    

1 + + 0 0 + + + + + + 0 + + + +   2+

2 + + 0 0 + 0 + + 0 0 + 0 + + 0   2+

3 + + 0 0 + 0 + 0 + + 0 + 0 0 +   2+

4 + + 0 0 + + + 0 + + 0 0 + 0 + P.S. 2+

AC                                 0✓

REStTM Adsorbed Sera 1X

    

LISS 

Rh system Kell Duffy Kidd MNSs   IS 37o

C

IgG

  D C c E e K k Fya Fyb Jka Jkb M N S s        

1 + + 0 0 + + + + + + 0 + + + +   3+ 3+ 3+

2 + + 0 0 + 0 + + 0 0 + 0 + + 0   3+ 3+ 3+

3 + + 0 0 + 0 + 0 + + 0 + 0 0 +   3+ 3+ 3+

4 + + 0 0 + + + 0 + + 0 0 + 0 + P.S. 3+ 3+ 3+

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Anti-Vel

• Antibody to the high incidence antigen that is present

in > 99 % of the population• Can be both IgM and IgG in nature• IgM complement binding anti-Vel can sometimes

present as hemolysins • Anti-Vel can cause severe hemolytic transfusion

reactions• Anecdotal reports of successful Vel+ transfusions.

Not recommended.

A Tale of Two Antibodies…..

• “It was the very best of times. It was the very worst of times” -Charles Dickens

• Let’s compare a similarly reacting nonsignificant antibody to our anti-Vel

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Ex. Nonsignifcant anti-I and cold autoantibody

    

PEG

  Kell Duffy Kidd MNSs   IS

IgG

  D C c E e K k Fya Fyb Jka Jkb M N S s   3+ 1+1 0 0 + 0 + 0 + 0 + + 0 + + 0 +   3+ 1+2 0 0 + 0 + 0 + 0 0 0 + 0 + 0 +   3+ 1+3 + 0 + 0 + 0 + 0 0 + 0 0 + 0 +   3+ 1+4 0 0 + 0 + 0 + 0 0 + 0 0 + 0 + I-  3+ 0✓

AC                                 1+ 0✓

Comparisons

Anti-Vel

• IS reactivity• IgM carryover• Negative autocontrol

Anti-I and cold autoantibody• IS Reactivity• IgM carryover• Negative autocontrol at IgG

Contrast 

Anti-Vel

• Cord cells reactivity not diminished

• Reactivity not diminished by prewarm technique

Anti-I and Cold autoantibody• Cord cell reactivity 

diminished• Reactions eliminated by 

prewarm technique

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Cautionary Tale

• Exercise caution using the prewarm technique as significant alloantibodies can be eliminated.

• If you have IS reactions and a negative autocontrol, proceed with caution

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…But it’s Not What the Textbook Said

• Not all antibodies behave in textbook fashion. Our anti-Vel behaved differently in several ways:

• (1) Cord cells did not react weaker• (2) REStTM did not adsorb out the

anti-Vel

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..and Our Patient?

• We believe our patient exhibited an anamnestic response to the initial transfusion and was undergoing a transfusion reaction.

• We initiated an ARDP search and imported 2 rbcs.

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Works Cited

• Lomas-Francis C, Reid M, The Blood Antigens Fact Book 2nd ed. New York City, New York: New York Blood Center; 2004

• Anstee J, Issitt Peter, Applied Blood Group Serology 4th ed. Durham, North Carolina: Montgomery Scientific Publications; 1998