Blood Banking forMommies

SEABB Annual Meeting 2008LeeAnn Prihoda, MEd, MT(ASCP)SBB

American Red Cross Reference LabDouglasville, GA

Disclaimer…• What you are about to see are some of

LeeAnn’s “pet peeves.”• She apologizes in advance for getting on her

soapbox!

Patient #1• 28 year old female• Admitted for delivery of first baby (38 weeks

gestation)• No known previous transfusions or pregnancies• T&S ordered prior to delivery• Initial results:

– ABO/Rh: B, Rh negative– DAT negative– Ab Screen –

• Negative at IS• Gel IAT: Cell I - 1+w, Cell II - 1+; Cell III - negative

• Your facility does not do any antibody ID, so you must send it to the Reference Lab– Sample received at Reference Lab– Let’s look at the request form and the sample…

D C c E e Cw K k Fya Fyb Jka Jkb Lea Leb P1 M N S s Gel1 + 0 + 0 + 0 0 + 0 0 + 0 0 0 + 0 + 0 + 1+w

2 + + 0 0 + 0 0 + + 0 + + + 0 + + + 0 0 1+3 + 0 + + 0 0 + + 0 + 0 + 0 + + + + + + 1+4 + + + 0 + 0 0 + 0 w + 0 0 + 0 + 0 + + 1+w

5 0 0 + + 0 0 0 0 0 + 0 + 0 + + + + + + 06 0 + 0 0 + 0 0 + + + + + 0 + + + + + 0 07 0 0 + + + 0 + + 0 + + 0 0 + + + 0 0 + 08 0 + + 0 + 0 0 + + 0 + + + 0 + 0 + 0 + 09 0 0 + 0 + 0 + + + 0 + + + 0 0 + 0 + 0 010 0 0 + 0 + 0 0 + 0 0 + 0 0 0 + 0 + + 0 011 0 0 + 0 + 0 0 + 0 + 0 + 0 + 0 + + + + 012 0 0 + 0 + 0 0 + + + + 0 0 + 0 0 + 0 + 0

Initial Panel at Reference Lab…

Rule in/Rule out…D C c E e Cw K k Fya Fyb Jka Jkb Lea Leb P1 M N S s Gel

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

2 + + 0 0 + 0 0 + + 0 + + + 0 + + + 0 0 1+3 + 0 + + 0 0 + + 0 + 0 + 0 + + + + + + 1+4 + + + 0 + 0 0 + 0 w + 0 0 + 0 + 0 + + 1+w

5 0 0 + + 0 0 0 0 0 + 0 + 0 + + + + + + 06 0 + 0 0 + 0 0 + + + + + 0 + + + + + 0 07 0 0 + + + 0 + + 0 + + 0 0 + + + 0 0 + 08 0 + + 0 + 0 0 + + 0 + + + 0 + 0 + 0 + 09 0 0 + 0 + 0 + + + 0 + + + 0 0 + 0 + 0 010 0 0 + 0 + 0 0 + 0 0 + 0 0 0 + 0 + + 0 011 0 0 + 0 + 0 0 + 0 + 0 + 0 + 0 + + + + 012 0 0 + 0 + 0 0 + + + + 0 0 + 0 0 + 0 + 0

Rule in/Rule out…D C c E e Cw K k Fya Fyb Jka Jkb Lea Leb P1 M N S s Gel

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

2 + + 0 0 + 0 0 + + 0 + + + 0 + + + 0 0 1+3 + 0 + + 0 0 + + 0 + 0 + 0 + + + + + + 1+4 + + + 0 + 0 0 + 0 w + 0 0 + 0 + 0 + + 1+w

5 0 0 + + 0 0 0 0 0 + 0 + 0 + + + + + + 06 0 + 0 0 + 0 0 + + + + + 0 + + + + + 0 07 0 0 + + + 0 + + 0 + + 0 0 + + + 0 0 + 08 0 + + 0 + 0 0 + + 0 + + + 0 + 0 + 0 + 09 0 0 + 0 + 0 + + + 0 + + + 0 0 + 0 + 0 010 0 0 + 0 + 0 0 + 0 0 + 0 0 0 + 0 + + 0 011 0 0 + 0 + 0 0 + 0 + 0 + 0 + 0 + + + + 012 0 0 + 0 + 0 0 + + + + 0 0 + 0 0 + 0 + 0

Conclusions – so far…• Patient has anti-D

– Weakly reactive in Gel

Do we need more info?• Did the patient receive antenatal RhIG?

– No indication on request form.– Reference Lab must call hospital for more info.

What we learned…• Patient received RhIg on 12/17/2008.• At that time, antibody screen was negative.

How do we report?• “The presence of anti-D in the patient’s serum is most likely

due to the recent administration of Rh Immune Globulin (RhIG). In serologic tests, it is not possible to distinguish between anti-D actively produced by a patient and anti-D passively acquired through administration of RhIG. Passively acquired anti-D can be expected to disappear from the circulation 5-6 months after the last injection. The presence of anti-D due to antenatal injection at the time of delivery cannot be considered protective; it is essential that the patient receive at least one standard dose of RhIG following delivery of an Rh-positive infant.”

Communication to patient’s physician

• Must be clear and complete– Must not indicate that patient has immune

stimulated anti-D– Must ensure that patient receives proper post-

partum RhIg.

How can you help?• Make sure that request form is properly

completed.• Make sure that all pertinent patient historical

information is provided.• Make sure that sample label EXACTLY

matches request form information.

Patient #2• 35 year old Caucasian female• G3P2, admitted for delivery (41 weeks gestation)• Possible previous transfusion, but not confirmed• 2 units RBC requested for C-section• Initial results:

– ABO/Rh: O, Rh negative– DAT negative– Ab Screen –

• Negative at IS• PeG IAT: Cell I - 3+, Cell II - 2+; Cell III - negative

– Phenotype: rr, kk, Fy(a+b+), Jk(a+b-), ss

D C c E e Cw K k Fya Fyb Jka Jkb Lea Leb P1 M N S s PeG Gel1 + 0 + + 0 0 0 + 0 + + + 0 + + 0 + 0 + 2+ 3+2 0 + 0 0 + 0 0 + 0 0 + + 0 0 + + + 0 0 3+ 3+3 0 + + 0 + 0 0 + 0 + 0 + 0 + + + + + + 3+ 3+4 0 0 + + + 0 0 + 0 w + 0 0 + + + 0 + + 0√ 05 0 0 + + 0 0 0 0 0 + 0 + 0 + + + + + + 0√ 06 0 0 + 0 + 0 + + + + + + 0 + + + + + 0 1+ 1+7 0 0 + 0 + 0 + + 0 + + 0 0 + + + 0 0 + 1+ 1+8 0 0 + 0 + 0 0 + + 0 + + + 0 + 0 + 0 + 0√ 09 0 0 + 0 + 0 0 + + 0 + + + 0 0 + 0 + 0 0√ 010 0 0 + 0 + 0 0 + 0 0 + 0 0 0 + 0 + + 0 0√ 011 0 0 + 0 + 0 0 + + 0 0 + 0 + 0 + + + + 0√ 012 0 0 + 0 + 0 0 + + + + 0 0 + 0 0 + 0 + 0√ 0

Initial Panel

D C c E e Cw K k Fya Fyb Jka Jkb Lea Leb P1 M N S s PeG Gel1 + 0 + + 0 0 0 + 0 + + + 0 + + 0 + 0 + 2+ 3+2 0 + 0 0 + 0 0 + 0 0 + + 0 0 + + + 0 0 3+ 3+3 0 + + 0 + 0 0 + 0 + 0 + 0 + + + + + + 3+ 3+4 0 0 + + + 0 0 + 0 w + 0 0 + + + 0 + + 0√ 05 0 0 + + 0 0 0 0 0 + 0 + 0 + + + + + + 0√ 06 0 0 + 0 + 0 + + + + + + 0 + + + + + 0 1+ 1+7 0 0 + 0 + 0 + + 0 + + 0 0 + + + 0 0 + 1+ 1+8 0 0 + 0 + 0 0 + + 0 + + + 0 + 0 + 0 + 0√ 09 0 0 + 0 + 0 0 + + 0 + + + 0 0 + 0 + 0 0√ 010 0 0 + 0 + 0 0 + 0 0 + 0 0 0 + 0 + + 0 0√ 011 0 0 + 0 + 0 0 + + 0 0 + 0 + 0 + + + + 0√ 012 0 0 + 0 + 0 0 + + + + 0 0 + 0 0 + 0 + 0√ 0

Initial Panel – what antibodies?

D C c E e Cw K k Fya Fyb Jka Jkb Lea Leb P1 M N S s PeG Gel1 + 0 + + 0 0 0 + 0 + + + 0 + + 0 + 0 + 2+ 3+2 0 + 0 0 + 0 0 + 0 0 + + 0 0 + + + 0 0 3+ 3+3 0 + + 0 + 0 0 + 0 + 0 + 0 + + + + + + 3+ 3+4 0 0 + + + 0 0 + 0 w + 0 0 + + + 0 + + 0√ 05 0 0 + + 0 0 0 0 0 + 0 + 0 + + + + + + 0√ 06 0 0 + 0 + 0 + + + + + + 0 + + + + + 0 1+ 1+7 0 0 + 0 + 0 + + 0 + + 0 0 + + + 0 0 + 1+ 1+8 0 0 + 0 + 0 0 + + 0 + + + 0 + 0 + 0 + 0√ 09 0 0 + 0 + 0 0 + + 0 + + + 0 0 + 0 + 0 0√ 010 0 0 + 0 + 0 0 + 0 0 + 0 0 0 + 0 + + 0 0√ 011 0 0 + 0 + 0 0 + + 0 0 + 0 + 0 + + + + 0√ 012 0 0 + 0 + 0 0 + + + + 0 0 + 0 0 + 0 + 0√ 0

Initial Panel – rule in / rule out

Conclusions so far…• What antibodies does the patient have?

– Anti-D– Anti-C– Anti-K

• Do we need to do anything else?– We need more information!!

Additional History• This is 3rd pregnancy:

– First in 05/2003• No known problems• Negative antibody screen throughout• RhIg given both antenatal and post-partum

– Second in 02/2006• No problems, negative antibody screen• Antenatal RhIg given• Infant Rh negative

– Third in 01/2008• Antibody screen at 10 weeks negative• Antenatal RhIg given in OB office at 28 weeks• No antibody screen done at that time• Uneventful pregnancy

Why does this patient have anti-D?• Previous transfusion of D positive blood?• Incorrect history?• RhIg failure?• Unknown FMH during this pregnancy?

• Or maybe it isn’t really anti-D???– If not anti-D, then what is it?– What about anti-G??

Anti-G – What is it?

• Antibody that recognizes an antigen present on almost all C positive and D positive red cells.– Antigen is distinct from C or D.

• Mimics anti-D+C.• Often reacts more strongly with C positive cells

than D positive.– Note: Anti-LW + anti-C can give a similar

appearance.

Anti-G - Significance• Relevance of identification depends on the

situation.• In routine transfusion:

– Complete characterization is not required.– Transfuse donor units negative for D and C, so

identity of anti-G is of academic interest only.• In prenatal and post-partum studies:

– RhIg administration required to protect against immunization to D antigen

– If anti-G is misidentified as anti-D+C, proper RhIg will not be given.

Anti-G – Significance, cont.

• Often found in combination with anti-D, anti-C or both.

• Can cause Hemolytic Disease of the Fetus and Newborn (HDFN) – Severity less than anti-D or anti-C.– Most often HDFN has been reported in conjunction

with a more potent anti-C.• Often not fully evaluated, so incidence may be

higher than reported.

When to suspect anti-G• Patient appears to have anti-D+C.• Reactions with C+ cells are stronger than with D+

cells.• Testing with various D+ cells may not give expected

reactions.– Reaction with R2R2 (DcE) cells may be weaker than reaction

with Ror (Dce) cells• Patient has had previous transfusion of only D negative

red cells, and/or

• Patient has had appropriate RhIg prophylaxis and no known pregnancy complications that would support immunization during pregnancy.

Anti-G - Investigation• First test with appropriate cells

– D-, C+: positive reaction– D+, C-: positive reaction– D-, C-: negative reaction

• Testing with rare G positive/negative red cells– rG (D-, C-, G+): positive reaction– R2r DIIIb (D+, C-, G-): negative reaction

• Adsorption/Elution studies– Issitt PD. Serology and genetics of the Rhesus

blood group system. Cincinnati: Montgomery Scientific Publications, 1979.

Adsorption/Elution for anti-GPlasma/Serum mixed 1:1 with Ro (Dce, G+) cells

Incubate at 37C – 60 min.

Centrifuge and remove adsorbed plasma/serum

Prepare eluate from adsorbed Ro cells

Mix eluate 1:1 with r’r (Ce, G+) cells

Incubate at 37C – 60 min.

Centrifuge and remove adsorbed eluate

Prepare eluate from adsorbed r’r cells

Test adsorbed serum/plasma to identify anti-C

Test adsorbed eluate to identify anti-D

Test eluate to detect presence of anti-G

Anti-G – Investigation

• Time-consuming and tedious!• Requires rare resources.

– Most often will require Reference Lab expertise.• Not necessary for routine transfusion therapy.

– Transfuse D-, C- red blood cells.• Adsorption/elution procedure can be done to

definitively differentiate anti-D+C from anti-G.

Back to our patient…• Additional rare cells tested:

– 3 of 3 rG (D-, C-, G+)* cells reacted 3+.– 1 of 1 R2r DIIIb (D+, C-, G-)# cells negative.

• Patient’s plasma was adsorbed with r’r cell (D-, C+, G+).– Remove anti-C and/or anti-G, but not anti-D.– Adsorbed plasma testing: 2 of 2 R2R2 (D+, C-) cells negative

D C c E e K k Fya Fyb Jka Jkb Lea Leb P1 M N S sPeG

Neat

PeG

Ads

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

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

3 0 0 + 0 + 0 + + + 0 + 0 + 0 + + 0 + * 3+ 0

4 0 + + 0 + 0 + 0 + + 0 0 0 + 0 + + 0 3+ 0

5 + 0 + + + 0 + 0 + + + 0 + + 0 + 0 + # 0 0

6 + 0 + + 0 0 + + 0 + + 0 + + + 0 0 + 2+ 0

7 + 0 + + 0 0 + 0 0 + 0 + 0 0 + + + 0 2+ 0

What can we conclude?• Does our patient have anti-D?

– No apparent anti-D• Neat plasma non-reactive with R2r DIIIb cells.• Adsorbed plasma non-reactive with other D positive cells.

– Anti-G present• Neat plasma reactive with rG cells .• Adsorbed plasma non-reactive with same cells.

– Anti-G (and possible anti-C) removed by adsorption.• Don’t forget…

– Patient also has anti-K

Significance of anti-G

• Should be investigated in female of child-bearing age, especially if known to have gotten proper RhIg in past.

• When to suspect anti-G– Apparent RhIg failure.– Reaction with C+ cells stronger than that with D+

cells.– Variable reactions with D+ cells.

Proper reporting of anti-G• Ensure that physician understands when no anti-

D is detected.• Report antibodies that are confirmed.• Provide information for physician on antibodies

identified.• Give physician guidance on proper treatment.• If no anti-D, patient must get RhIg prophylaxis.

Thanks for your attention!!