The Duffy blood group system is an important blood group system that impacts susceptibility to malaria. It was discovered in the 1950s through studies of patients who developed antibodies. The Duffy antigens Fya and Fyb are carried on the Duffy glycoprotein, which acts as a receptor for malaria parasites and chemokines. People with the Fy(a-b-) phenotype, common in Africans, lack the Duffy glycoprotein and are resistant to malaria. The document provides details on the genetics and biochemistry of the Duffy glycoprotein, as well as the various Duffy antigens and antibodies that have been discovered.

1. IHBT

2. DUFFY BLOOD GROUP SYSTEM
 1950- cutbush, mollison & parkin- Mr.Duffy- a
hemophiliac patient with multiple transfusions- anti
fya
 1951- Ikin & co workers- fyb- in women of 3
pregnancies
 1955- sanger & colleagues- africans are Fy(a-b-)
 FyFy- common in blacks ; rare-in whites

3.  1975- Miller& co workers- Fy(a-b-) RBC resists
P.knowlesi & P.vivax
 Rare- Fy3,Fy5
 Fy(a-b-) are also Fy:-3,-5
 Fy-5 – seen in Rh null regardless of duffy status
 ISBT- OO8 , symbol- FY

4. Fya and Fyb antigens
 6 weeks gestational age
 well developed at birth.
 13,000 to 14,000 Fya or Fyb sites on Fy(a+b-)& Fy(a-b+)
RBCs, respectively
 there are half that number of Fya sites on Fy(ab) RBC(
6900 sites/cell)
 not seen on platelets, lymphocytes, monocytes, or
granulocytes

5.  identified in brain, colon, endothelium, lung, spleen, thyroid,
thymus, and kidney
 Inc expression – reticulocytes > mature RBC
 destroyed by ficin, papain, bromelin, and chymotrypsin, and by
ZZAP
 not affected by AET or glycine-acid-EDTA treatment.
 Neuraminidase -reduce the molecular weight of Fya and Fyb, it
does not destroy antigenic activity & purified trypsin

6.  Fya and Fyb do not store well in saline suspension -
tend to elute from RBCs stored in a medium with low
pH or low ionic strength.
 leads to inhibitory substances in the supernatant
fluid, which can weaken the reactivity of an anti-Fya or
anti-Fyb.
 changes are not seen in RBCs stored in licensed
anticoagulants, e.g., ACD, CPD, CPD-A1, or the reagent
solutions used by commercial manufacturers for
reagent RBCs

7. Phenotype India Whites Blacks
Fy(a+b-) 42.1 17 9
Fy(a+b+) 4.5 49 1
Fy(a-b+) 12.3 34 22
Fy(a-b-) 0.3 Very rare 68

8. ANTI Fya and ANTI Fyb
 6-10% of clinically significant antibodies
 Anti-Fya -common antibody; single specificity or in a
mixture
 Anti-Fya - three times less frequently than anti-K. Anti-
Fyb is 20 times less common than anti-Fya and often
occurs in combination
 Fya antigen -more immunogenic in Fy(a-b+) whites than
in Fy(a-b-) blacks

9.  usually IgG1 - react best at AHG phase.
 50% anti-Fya and anti-Fyb bind complement upto C3
 A few examples are saline agglutinins.
 Antibody activity is enhanced in a low ionic strength medium.
 Because anti-Fya and anti-Fyb do not react with enzyme treated
RBCs, this is a helpful technique when multiple antibodies are
present.
 Naturally occurring - rare

10.  show dosage- more obvious with saline agglutinins.
 some reagent RBCs appears to be from homozygotes (and
have a double dose of either Fya or Fyb) -may actually -
from heterozygotes if they are from black donors;
 a silent allele, Fy, is commonly found in blacks.
 For example, Fy(a+b-) RBCs =double dose of Fya if white
FyaFya donor but a single dose of Fya if from a black donor
who is genetically FyaFy.

11.  Additional phenotypic markers found in black donors
Ro, S-s-, V+VS+, Js(a+), Le(a-b-).
 Anti-Fya and anti-Fyb = acute & delayed HTR & mild
HDFN
 Once the antibody is identified, Fy(a) or Fy(b) blood
must be given, 1 in 3 is Fy(a-) and 1 in 5 is Fy(b-)
 auto antibodies with mimicking Fya and Fyb
specificity have been reported
 Fy- minor histo compatibility antigen in renal allograft

12. BIOCHEMISTRY
 336 AA
 relative mass of 36 kD ;2 N-glycosylation sites
 traverse the cell membrane seven times ( 7 trans
membrane alpha helical domains)and 2 disulfide bridges.
 Extracellular N terminus- 65AA,
 C terminus- intracellular
 Homologous to protein of G protein coupled receptor

14.  Fya/Fyb- SNP in exon 2- gly42asp on extra cellular N
terminus
 Fy3 - on the third extracellular loop
 Fy6 - amino acids 19 through 25
 a member of the superfamily of chemokine receptors
DARC (Duffy antigen receptor for chemokine)
 Receptor for P.vivax in N terminal domain( sulphation
of Tyr 42- increases affinity of binding by 1000 fold)

15. DARC
 binds CXC chemokines- (IL8) & melanocyte growth-
stimulating activity (MGSA)& CC chemokines
regulated on activation, normal T-cell expressed and
secreted (RANTES) & macrophage chemoattractant
protein-1 (MCP1).
 a scavenger for locally released chemokines.
 Decreases angiogenesis- ca prostate progression
decreases ( erythroid DARC is inc in ca prostate pts of
african ancestry)

16.  Present in capillaries & post capillary venules-
 Inhibits cancer metastasis & induction of cellular
senescence
 Facilitates movement of chemokines across
endothelium
 IL-8 binding – residues on EC domain 1 and 4 –
requires –S=S-
 Ligands for parasites- glycophorin A,B,C,D and NANA

17.  leucocyte recruitment to sites of inflammation
 regulate chemokine bioavailability between the
circulation and extravascular sites during
inflammation
 In Fy(a− b−) phenotype and many patients with sickle
cell disease have the Fy(a− b−) phenotype- elicit
markers of inflammation, & inc chances of allo
immunization
 more susceptible to chronic organ damage
&proteinuria & more susceptible to asthma, worst
outcomes for Acute lung injury

18. GENETICS
 chromosome 1
 the first human gene to be assigned to a specific
chromosome- DONAHUE
 2 exons- exon1 – encodes first 7 AA
 CO DOMINANT alleles
 Fy locus is syntenic to Rh
 Fya and Fyb allele- Fya and Fyb
 silent allele, Fy, that is the major allele in blacks

19. Fy allele
 found to be an Fyb variant - change in the promoter
region(GATA1) of the gene
 T-C substitution
 which disrupts the binding site for mRNA
transcription in the RBC
 do not express Fyb on their RBCs but express Fyb in
other tissues,
 no anti-Fyb is made by these individuals.
 in new guinea- GATA mutation seen in Fya allele

20.  In whites - 14 bp deletion in the Fy gene, resulting in a
reading frame shift and the introduction of a
translation stop codon.
 no Duffy protein on their RBCs or on other tissues and
--form anti-Fyb and anti-Fy3.
 Fy3 developed first, then Fyb, and that Fya arose
during human evolution.

21. DUFFY AND HIV
 Lachgar and co-worker- binds HIV and that this
binding is inhibited by the chemokine RANTES
 Here, red cells may be a reservoir for HIV and transmit
the virus to leucocytes.
 Individuals with the Fy(a− b−) phenotype are reported
to be more likely to acquire HIV.

22. Fyx
 1965 by Chown - new allele at the Fy locus.
 inherited weak form of Fyb
 Mis sense mutation- Arg89Cys in 1st EC loop of
cytosolic domain- reduced amount of Duffy
glycoprotein on the surface of RBCs
 reacts with some but not all examples of anti-Fyb.
 in white populations.

23.  Individuals with Fyx may type Fy(b-), but their RBCs
adsorb and elute anti-Fyb.
 depressed expression of Fy3 & Fy5 antigens.
 no anti-Fyx.

24. Fy3 antigen & anti Fy3
 In 1971 Albrey et al- anti-Fy3 in the serum of an Fy(a-b-
) white Australian female.
 It reacted with all RBCs tested except Fy(a-b-)
 an inseparable anti-FyaFyb,
 react with an antigenic determinant or precursor
common to both Fya and Fyb and was called Fy3.
 the Fy3 antigen is not destroyed by enzymes.

25. Anti fy 3
 rare antibody made by Fy(a-b-), causes HTR
 found in white, black, and Cree Indian families.
 Blacks with the Fy(a-b-) phenotype rarely make anti-Fy3.
 Examples of anti-Fy3 produced by non-blacks appear to react
with all Duffy positive cells equally well.
 made by blacks are similar, but they react weakly or not at all
with Duffy positive cord RBCs.
 Some patients who make anti-Fy3 initially make antiFya.

26. Fy4 and anti Fy4
 In 1973 Behzad et al- anti-Fy4 in a Fy(a+b+) black female
with sickle cell anemia.
 reacted with RBCs from all Fy(a-b-) blacks, many Fy(a+b-)
and Fy(a-b+) blacks
 but not with Fy(a+b+) blacks, and not with whites of any
Duffy type.
 most Fy(a-b-) blacks carry an Fy4 - genetically Fy4Fy4.
 is not destroyed by enzymes.
 No other example of anti-Fy4 has been reported

27. Fy5 antigen & anti Fy5
 Anti-Fy5 - Colledge et al- in 1973 in the serum of an Fy(a-b-)
black child who later died of leukemia.
 thought as second example of anti-Fy3 bcoz- reacted with all
Fy(a+) or Fy(b+) RBCs but not with Fy(a-b-) cells.
 differed in that it reacted with a Fy(a-b-)Fy:-3 white female,
 but it did not react with Fy(a+) or Fy(b+) Rhnull RBCs
 reacted only weakly with Fy(a+) or Fy(b+) D- RBCs.
 Sometimes, sera with anti-Fy5 also contain anti-Fya.

28.  anti-Fy5 have been reported in multiply transfused
Fy(a-b-) sickle cell patients with a mixture of other
antibodies.
 result of interaction between the Rh complex and the
Duffy glycoprotein.
 Fy5 is not destroyed by enzymes

29. Fy6 antigen
 In 1987 Nichols et al -a murine monoclonal ab
 reacted much like anti-Fy3, except that its reactivity was
destroyed by ficin, papain, and chymotrypsin.
 Trypsin enhanced its reactivity.
 The antibody appeared to define the Duffy receptor used by P.
vivax to penetrate RBCs
 Fy6 involves amino acids 19 to 25 on the extracellular domain of
the Duffy glycoprotein.
 No human examples of anti-Fy6 have been reported to date.

31. LUTHERAN
 1945, anti-Lua was discovered in SLE patient
following the transfusion with LPA- ( also made anti-c,
anti-N, anti-CW, and anti-Levay, now known as Kpc!)
 In 1956 Cutbush & Chanarin - anti-Lub
 1961- Crawford- Lu(a-b-) with dominant inheritance.
 In 1963 Darnborough et al Lu(a-b-)phenotype
inherited as a recessive silent allele.
 22 antigens- Lu 10,LU15 = obsolete
 4 anti thetical ag

32.  HPA- Lub,
Lu3,lu4,lu5,lu6,lu7,lu8,lu11,lu12,lu13,lu16,lu17,lu20,lu21,
lu22
 Lua-8% in whites& 5 in blacks
 Lu9- 2%
 Lu14-2.4%
 Aua,lu18=80% whites
 Aub,lu19=50% whites

33. Phenotype Prevalence in most population
Lu(a+b-) 0.15
Lu(a+b+) 7.5
Lu(a-b+) 92.35
Lu(a—b-) Very rare

34. Basic concepts
 either very HPA/LPA.
 10 to 12 weeks
 poorly developed at birth and do not reach adult
levels until age 15 years.
 not been detected on platelets, lymphocytes,
monocytes, or granulocytes
 seen in brain, lung, pancreas, placenta, skeletal
muscle, & hepatocytes (especially fetal hepatic
epithelial cells

35.  Lu on placenta- adsorption of maternal Ab to lu Ag-
low risk of HDFN
 Resistant to papain/ficin/glycine acid EDTA
 Destroyed by trypsin,chymotrypsin,pronase
 Do not react with DTT/AET treated RBC
 4 antithetical Ag- lu1/lu2,lu6/lu9(
ser275phe),lu8/lu14,lu18/lu19

36. Lua and Lub antigens
 Co dominant
 Most- lu(b+)
 8% of whites & 5 % in blacks- lu(a+)
 Lua/lub= his77arg
 expression is variable from one individual to another; one
individual’s RBCs can also vary.
 Lub sites= 1640 -4070 on lu(a-b+)
 850-1820 on lu( a+b+)

37.  Transient loss of lua- in AITP
 Shows dosage effect
 Lub- more immunogenic, HPA

38. Anti Lua
 IgM, naturally occurring saline agglutinins that react better
at room temperature.
 A few react at 37*C by IAT.
 Some -binds complement, but invitro hemolysis has not
been reported.
 may be IgA as well as IgM and IgG
 Anti-Lua often goes undetected in routine testing because
most reagent cells are Lu(a-).

39.  encountered as CXM incompatible or during an antibody
workup for another specificity.
 mixed-field reactivity in a test tube.
 not altered with enzymes ficin and papain, destroyed with
trypsin, chymotrypsin, pronase, AET, and DTT.
 Most Lua antibodies are clinically insignificant in
transfusion.
 no immediate but DHTR

40. Anti Lub
 IgM and IgA but most anti-Lub is IgG4 , reactive at 37*C
at AHG phase.
 It is made in response to pregnancy or transfusion.
 Alloanti-Lub reacts with all cells tested except the
autocontrol
 reactions are often weaker with Lu(a+b+) RBCs and cord
RBCs.
 Ficin or papain does not significantly alter reactivity.but
AET or DTT destroys Lub antigen
 Dec survival of transfused cells, post transfusion jaundice
noted

41. BIOCHEMISTRY
 Located on type I transmembrane protein
 Protein in 2 forms – d/t alternate RNA spliceosomes
 Lutheran glycoproteins.
 85kD , 597 AA, 5 potential N glycosylation sites as EC
IgSF domain
 It traverses the cell membrane just once – hydrophobic
transmembrane domain -19AA
 cytoplasmic domain of 59 AA- interacts with spectrin
 Lua/lub- on 1st domain, Aua/Aub – on domain 5

43.  Smaller isoform-BCAM
 78 kD
 lacks cytoplasmic domain
 external portion 5 disulfide-bonded domains (three are
constant, and two are variable).
 belong to the immunoglobulin superfamily of proteins.
 Binds laminin in asp312 on domain 3

44.  Laminin- has alpha 5 chains, La-Lu complex migrates
mature RBC from marrow to circulation
 Upregulation of laminin – in sickle cell patients-
vascular crisis
 In PCV, increased adhesion- d/t la- lu on endothelial
cells

45. Molecular basis of HPA &
antithetical antigens
 through the creation of Lutheran glycoprotein
mutants.
 Subsequent sequencing of exons encoding on EC
domains
 For most- it is SNP

46. GENETICS
 Chr 19
 (H, Se, Le, LW, Oka) and genes for C3, apolipoprotein
C-II (APO), and myotonic dystrophy
 Lu and Se gene (FUT2) was the first example of
autosomal linkage described

47. Lu(a-b-) phenotype- dominant
 Reported by crawford
 phenotype was seen in successive generations as 50% of
members & null individuals passed normal genes-
expression was suppressed by a rare dominant regulator
gene later called In(Lu) for “inhibitor of Lutheran.” or
SYN1B
 In(Lu) segregates independently from Lutheran.
 prevalence- 0.03%

48.  Unsuitable as donor- lysis invitro after 7 days when
stored in alsever solution- 5 times than normal
 Mutations in gene for EKLF
 Caused by inheritance of loss of function mutation on
1 allele of EKLF
 abnormal poikilocytosis and acanthocytosis
 On incubation with plasma> 24 hrs- decreased
osmotic fragility , in vivo survival – normal
 Carries trace Lu antigens – detected ONLY by elution
& adsorption

49.  No anti Lu3
 Decreased expression of CD44
 Weak expression of P1,i,AnWj.MER2,Inb
 Also Kna,Sla,McCa,Csa,Yka
 SYN-1A= gene for nirmal expression of inv Ag
 SYN 1B- allele causing depression
 SYN1C= luw
 Para lutheran- lu2,lu5,lu7,11,12,13,16,17,20( ab reacts
with all except lu(a-b-)

50. Recessive lu(a-b-)
 d/t inheritance of 2 rare silent alleles LuLu
 Parents & off spring- Lu(a-b-), single dose of lub
 Lacks all lu antigens
 Makes anti Lu3
 Normal expression of P1,i
 Inactivating mutation- truncated glycoprotein- no
integration in RBC

51. Recessive X linked inhibitor type
 By normal et al
 In australian family
 Male, X borne, locus- XS
 Trace Lub detected by adsorption & elution
 Mutation in GATA1- ter 414 arg- 41 aa extra @ c
terminal
 XS1 – common allele, XS2- rare inhibitor that
suppresses in hemi azygous state

52. Mode of inh Gene Lu Others Antilu3
Dominant EKLF Extremely
weak
Decreased
P1,i,Inb,Anwj,
MER2,CD44
No
Recessive Lu None Not affected Yes
X linked XS2 Extremely
weak
Not affected no

53. Anti lu3
 reacts with all RBCs except Lu(a-b-).
 looks inseparable anti-Luab
 recognizes a common antigen, Lu3
 Anti-Lu3 is usually antiglobulin reactive.
 This antibody is made only by LuLu individuals, i.e.,
the recessive type of Lu(ab)
 . RBCs from dominant and X linked type Lu(a-b-)
individuals can be safely transfused to patients with
anti-Lu3.