Case # 2

• Clinical progress: 2009• HLA typing: no sibling match• Hb S: 27-40% anti-S, anti-Jk(b)• TCD velocities conditional range: < 200 cm/s• Liver iron content (MRI): 6.7 mg Fe/g• RBC transfusion # 16• Commenced Deferasirox (oral Fe chelator)

• 2010• Hb S: 16-28%• Brain MRI/MRA: moderate narrowing of A1 segment of ACA• Blood bank: DAT weakly POS anti-IgG (probable autoAb)

Transfusion in Patients with Haemoglobinopathies(Sick Kids approach)

• Thalassemia (including congenital anemia like Diamond Blackfan Syndrome) and sickle cell disease

• Extended red cell phenotyping before first transfusion:– Rh (C, E, c, e), K1(Kell), Fya, Jka, (Jkb, S in SCD patients)

• Patient already transfused– Hypotonic saline (0.3% NaCl will lyse normal cells but not

sickle cells)AABB Technical Manual 15th edition Method 2.16

– Molecular genotyping– Family studies

• Prophylactic antigen matching:• Thalassemia: K negative• SCD: Rh and K matched, Fya, Jk, (S) if alloimmunized (No

need for prophylactic Fyb matching• Why not Fyb?

Fy(a-b-) Phenotype• Frequency

– Very rare (0%) in Caucasians and Asians– 68% in Africans

• Mutation in the promoter region of FYB (–33 T>C), which disrupts a binding site for the erythroid transcription factor GATA-1 and results in the loss of Fy expression on RBCs.

• Because the erythroid promoter controls expression only in erythroid cells, expression of Fy proteins on endothelium is normal in people with Fy(a–b–) RBCs.

• To date, all blacks with a mutated GATA box have been shown to carry FYB, therefore Fyb is expressed on their nonerythroid tissues.

• This explains why Fy(a–b–) individuals make anti-Fya but not anti-Fyb.

Immunohematology 2004;20:37-49

The Duffy Antigen/Receptor Chemokine locus

• Comprised of 2 exons, spanning ~2 kbp in the region of chromosome 1q22-23

• The single gene is responsible for the expression of Duffy antigens on RBCs and other non-erythroid tissue

• Promoter GATA-1 box mutation is responsible for the Fy(a-b-) RBC phenotype (pseudo-null)

• The gene product has no obvious function on the surface of RBCs; null RBCs appear to function normally

• FY affords protection from malarial invasion• It has been proposed that the gpFy modulates chemokine

levels in the blood; regulation of inflammation by scavenging chemokines

Allelic variants of Allelic variants of DARCDARCGATA-1 FY*A/FY*B

-33C^ ^

125A

-33T^ ^125A

(amorph) FY

FYX

FY*X

265T^

265C^

-33T^ ^

125A

(wt) FYB

265C^

-33T^ ^

125G

FYA

265C^

(null) FY0-33T

^ ^125A 265C

^

Transfusion in Patients with Haemoglobinopathies(Sick Kids approach)

• Start with SAGM units (prestorage leukoreduced)• 15 to 20 ml/kg, SAGM red cells have hct approx 0.6• SCD: Units tested by sickle test (sickle negative for

exchange transfusions)• Repeated allergic reactions: pre-med• Recurrent allergic reactions: plasma-reduce• Frequent allergic reactions: Washed red cells from

CBS: 24 hours shelf-life• Currently, no Sick Kids patients on washed cells

Case #3

• Female, born in 1980, sickle cell disease• B pos, C-E-c+e+, K-, Fy(a-b-), Jka(a+b+)• 1998: no antibody, transfused 3 units• 1999: anti-K, anti-C, anti-E, autoantibody• Sept 1, 2004, transfused 2 units, O neg, C- E-

K- S- Fya-, crossmatch compatible• Sept 14, 2004, 3 units B pos, C- E- K- S- Fya-,

crossmatch incompatible

Case #3 (2004)

• Anti-IH– 4+ with group O cells by Sal I.S., 370C and IAT– 1-2+with group B cells– Negative to weak with Oh (Bombay) cells

• Testing with Oh cells

Cell IS 370C IAT

H-, M- 0 w 1

H-, M+N+ w 2 3

H-, M+N - 2 4 3

Anti-IH• I antigen: subterminal portion of the oligosaccarides that are

eventually converted to H, A, and B antigens• Most normal adult RBCs are I-positive • H antigen is the substrate for A and B antigens• H antigen expression: O > A2 > B > A1B > A1 > A1B

• The most common cold autoagglutinins are directed against the Ii blood group

• The most commonly encountered cold autoagglutinin recognizing complex ABH-Ii antigens is anti-IH

• Anti-IH does not generally interfere in pretransfusion testing done at 370C, but may be picked up in MTS-gel

• Anti-IH is usually not clinically significant, anti-IH causing hemolytic transfusion reaction is very rare but has been reported.

Transfusion 2000:40;828

Case #3 (2005-2006)

• Anti-IH not detectable• Autoantibody• Recommended for transfusion: group B, C- E- K-

M- Fya-• May 5, 2006, transfused 2 units O pos, C- E- K-

Fy(a-b-) S-, MTS compatible, Hb 70 to 99 g/L• May 15, 2006, Hb to 50g/L• Strongly reactive with all cells tested, except for 2

Group B Rhnull cells• Episode of hyperhemolysis, eventually recovered

Case #3 (2008)

Antibodies to high prevalence antibodies

• Red cell alloantibodies– k, Kpb, Lub, Jsb

– Jk3, U – African ethnicity: SsU, Jsb, Ata (Augustine), Hy (Holley),

Joa (Joseph)1. Phenotype patient’s RBCs (Rh and others)2. Antigen negative cells3. Enzymes and chemicals

• Antibodies to reagent/preservative• ABH antibodies

– Group O patient, think Bombay or para Bombay– Non group O patient, think IH (more common in A1, A1B,

less common A2, B, A2B)

Case #3 (2008)

Case #3 (2008)

Summary

• Phenotype patient before 1st transfusion• If transfused, hypotonic saline for SCD, molecular

genotyping and family studies• Autoantibodies are common• SCD patients can make unusual alloantibodies• When investigating for high incidence antibodies,

do not forget ABO and reagents• Clinical information including patient’s ethnic

background

References

• Guidelines for Antibody Investigation. AABB 2010

• Judd’s Methods in Immunohematology, 3rd edition, AABB Press 2008.

• The Blood Group Antigen Facts Book, 2nd edition, 2004, Reid & Lomas-Francis, NYBC

• Applied Blood Group Serology 4th edition. Issitt 1998.