Nicole L. Draper, MD
Assistant Professor, Department of Pathology, University of Colorado
Associate Medical Director, Transfusion Services,
University of Colorado Hospital
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Identify categories of red cell and platelet
antibodies.
Discuss laboratory tests that are used to
identify antibodies, and tests that aid in
prediction of clinical significance.
Understand the clinical implications of
laboratory test interpretations in pregnancy.
42-year-old woman, G6P5003 at
13 weeks gestation
 First prenatal visit this pregnancy
 This baby is with a new partner
 Two previous pregnancies had heart
defects
 Standard prenatal testing today
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hCG
ABO and Rh type, red cell antibody screen
Hematocrit and MCV
Rubella and varicella immunity
Urine culture
Syphilis, HIV and hepatitis B antigen screening
Cervical cytology
If at risk
• Chlamydia, ghonorrhea, hepatitis C
• Thyroid function
• Cystic fibrosis, Down syndrome, fragile X,
hemoglobinopathies

Maternal IgG antibodies against paternal
antigens on fetal red cells
• Fetomaternal hemorrhage
• Transfusion

Increased likelihood in the third trimester
• Delivery
• Increased fetal blood
volume
• Cytotrophoblast no
longer consistently
present
Trends in Endocrinology & Metabolism, Volume 22, Issue 5, 2011, 164 - 170
Hemolysis
Anemia
Extramedullary
Hematopoiesis
Bilirubin
Ischemia
Kernicterus
Cardiac Failure
Decreased
oncotic pressure
Edema
Type: A, Rh-negative with anti-B
Rh
Kell
Duffy
Kidd
Lewis
MNS
Luth
SP
K k
Fya
Fyb
Jka
I
+ + 0 0 + 0 +
+
+
0
+
+
0
+ 0 + 0
0
+
+
II
+ 0 + + 0 0 +
+
0
+
0
0
+
0 + 0 +
0
+
+
III
0 0 + 0 + + +
0
+
+
0
0
+
0 + + +
0
+
0
D
AC
C c
E e
Jkb Lea
Leb
M
N S s Lua
Lub
0
• Common clinically significant antigens are represented
• Chosen because of potential for hemolysis
• Performed on first prenatal visit
Cross the placenta (IgG)
 Amount of antibody (titer)
 Antigen present on fetal RBC’s

• Paternal type
• I, Le(a), Le(b), P-system, Lu(b), Yt(a), VEL
not developed
• ABO not well
developed
Transfusion. 2010 Jul;50(7):1571-80
RBC
Antigens
A
B
AB
(AA, AO) (BB, BO) (AB)
Plasma
Anti-B
Antibodies
Anti-A
None
None
(OO)
Anti-A
Anti-B
Anti-A,B



Most common HDFN
O mom with A or B baby
DAT often negative
• ABO antigens few and
unbranched
• A and B on tissues
• Soluble A and B

Mild neonatal jaundice
Prevent alloantibody formation with
administration of passive anti-D (RhIg)
 Antepartum

• D-antigen negative (Rh-)
• Has not formed an anti-D
• Father of the baby is known to be Rh+ or
Rh type is unknown
• Full-dose 300 μg (1500 IU) administered
at 28 weeks gestation or with risk of fetalmaternal hemorrhage

Postpartum

Rh+ platelet transfusions

ITP treatment
• D-antigen negative (Rh-)
• Has not formed an anti-D
• Baby is Rh+
• Dose calculated
• Transfusion. 2014 Mar;54(3):650-4
Patient reports no recent
administration of RhIg
 Last pregnancy 9 years ago in Mexico

IgG antibody that can cross the
placenta
 Very severe HDFN possible
 Anti-D titer of 16 associated with
significant fetal anemia

Rh
Duffy
Kidd
Lewis
MNS
Luth
K k
Fya
Fyb
Jka
Jkb
Lea
Leb
M N S s Lua
Lub
+ 0 + 0 + 0 +
+
+
0
+
+
0
+ 0 + 0 0
+
D
Ror
Kell
C c
E e
• Anti-D currently too weak to titer
• Repeat in 1 month
LISS
0

Circulating cell-free fetal DNA testing
• Fetus is predicted to be female
• Fetus is predicted to be Rh(D) positive

FOC type
• Heterozygous
• ~2% have
incorrect
knowledge
of paternity
http://www.ariosadx.com/healthcare-professionals/technology/
Rh
Duffy
Kidd
Lewis
MNS
Luth
K k
Fya
Fyb
Jka
Jkb
Lea
Leb
M N S s Lua
Lub
+ 0 + 0 + 0 +
+
+
0
+
+
0
+ 0 + 0 0
+
D
Ror
Kell
C c
E e
• Anti-D currently titer of 4
• Repeat in 1 month
LISS
3+
Rh
Kell
Duffy
Kidd
Lewis
MNS
Luth
SP
K k
Fya
Fyb
Jka
I
+ + 0 0 + 0 +
+
+
0
+
+
0
+ 0 + 0
0
+
+
II
+ 0 + + 0 0 +
+
0
+
0
0
+
0 + 0 +
0
+
+
III
0 0 + 0 + + +
0
+
+
0
0
+
0 + + +
0
+
0
D
AC
C c
E e
Jkb Lea
Leb
M
N S s Lua
Lub
0
Rh
Kell
Duffy
Kidd
Lewis
k
Fya
Fyb
Jka
Jkb
Lea
Leb
1
+ + 0 0 + 0 +
0
+
+
+
0
2
+ + 0 0 + + +
+
0
+
+
3
+ 0 + + 0 0 +
0
+
0
4
+ 0 + + 0 0 +
0
0
5
0 + + 0 + 0 +
0
6
0 0 + + + 0 +
7
MNS
Luth
s
Lua
Lub
SP
+
0 + + +
0
+
4+
+
0
0 + 0 +
0
+
4+
+
+
0
+ + 0 +
0
+
3+
+
0
+
0
+ 0 0 +
0
+
3+
+
+
0
0
+
+ + 0 +
0
+
3+
+
+
+
+
0
+
+ 0 0 +
0
+
0
0 0 + + + 0 +
0
+
+
0
0
+
+ + 0 +
0
+
0
8
0 0 + 0 + + +
+
+
0
+
+
0
0 + + 0
0
+
0
9
+ 0 + 0 + 0 +
0
0
0
+
0
0
+ + 0 +
+
+
3+
D
A
C
C
c
E
e
K
M N S
0
Rh
D
C c
Kell
E e
K k
Duffy
Kidd
Lewis
MNS
Luth
Fya
Fyb
Jka
Jkb
Lea
Leb
M N S s Lua
Lub
LISS
r’r
0 + + 0 + 0 +
0
+
+
0
0
+
+ + 0 + 0
+
0
Ror
+ 0 + 0 + 0 +
+
+
0
+
+
0
+ 0 + 0 0
+
3+
• Anti-C too weak to titer
• Anti-D currently titer of 256
• MCA doppler ultrasound needed
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IgG, but not typically clinically significant
Appears to be anti-D and anti-C
• Shared epitope
• 103Ser
Explains D- exposed to D-(C+) blood
develops apparent anti-D
Almost all anti-G with anti-D or anti-C
RhIg?
Semin Hematol. 2007 Jan;44(1):42-50
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Not likely anti-G
• Formed anti-D months before anti-C
• Very different titer results
• No RhIg indicated
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Likely RBC exposure during this
pregnancy
• Anamnestic response for anti-D
• New formation or anamnestic response for
anti-C
AJUM November 2010; 13 (4):24-27
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Intrauterine transfusion of RBC’s
ABO compatible with both mother & fetus (usually O-neg)
CMV-safe (LR & CMV-seronegative)
Irradiated
Lacking any antigen for which mom has the corresponding
antibody
• Hyperpacked (Hct target 85%)
• Fresh (<5days)
• Hgb S-negative
•
•
•
•
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Delivery
• >33weeks
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Baby girl delivered by C-section at 34
weeks
Type A, Rh(D)-positive
C-antigen negative
IgG DAT 3+
Hemoglobin 5.1 g/dL (14.5-22.5)
MCV 153.2 fL (95.0-121.0)
RDW 132.0 fL (37.1-48.8)
Max bilirubin 12.3 mg/dL at day 1
Required RBC transfusion, not exchange
32-year-old woman, G3P2002 at
27 weeks
 Transfer to maternal fetal medicine for
anti-K
 Titer at outside institution of 64
 MCA doppler ultrasound with PSV 1.7
MoM
 Planned IUT
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Discrepancy between titer and bilirubin
level in amniotic fluid (Berkowitz et al.
1982)
Anti-K titer of 2048
Decreased bilirubin
between weeks 21
and
24 with development
of hydrops
Inhibits fetal erythropoiesis
 Causes peripheral hemolysis
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http://ben-may.uchicago.edu/faculty/macleod/_img/research_3.jpg
Titer of 8 considered critical
 MCA Doppler preferred
 20% of severely affected infants type
as K- with IgM reagent anti-K

K
K
K
IUT at 27, 28, 30 and 31 weeks
gestation
 MCA doppler PSV normalized
 Delivered baby at 33 weeks
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Usually IgM antibody
 Conversion to an IgG occurs rarely –
6 reports in English literature
 DTT treatment to determine IgG
component
 IgG anti-M titer of 32 critical

IgG
 Autoantibodies often exacerbated in
pregnancy
 Hemolysis in mother?
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DAT
• Poor predictive value
• 23% of DAT+ infants required treatment
for hyperbilirubinemia
• RhIg
• ABO incompatibility
A 19-year-old woman
 Diagnosis of immune
thrombocytopenic purpura (ITP)
 Post splenectomy at age 13
 Platelet count typically 30-50x109/L
 Wants to conceive, but concerned
about how her disease could affect
pregnancy
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Acute
• Typically children
• Post viral infection,
upper respiratory
• Platelets 5-20

>3mm purpura
usually
<3mm petechiae
Chronic
•
•
•
•
20-50 years old
More common in women
Other associated autoimmune diseases
Platelets 5-75
http://www.dermnet.com/images/Henoch-Schonlein-Purpura/picture/14794
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Test for antibodies to platelet specific
antigens to confirm the diagnosis
www.lookfordiagnosis.com
http://www.hematology.org/Clinicians/Guidelines-Quality/Quick-Reference.aspx
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Transplacental transfer of maternal
antibodies (anti-HPA-1a most common)
against paternal antigens
Mother formed platelet specific antibody
after
• Exposure to baby blood
• Transfusion
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Can have moderate (50) to severe (10)
thrombocytopenia, risk of intracranial
hemorrhage
Previous pregnancy outcome good predictor
for future pregnancies
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The mother of the baby
The father of the baby
The baby’s paternal grandfather
The baby’s maternal grandfather
A random, unrelated donor
Transfusion of washed, maternal platelets or
platelet-antigen matched platelets
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Tested sera from 187 pregnant women for
the presence of anti-HLA-A, -B, and -C and
anti-B lymphocytes or anti-HLA-DR).
Patients were studied before 20 weeks,
between 21 and 30 weeks, and between 31
and 40 weeks' gestation.
No correlation was found between the
presence of such antibodies and obstetric
complications, fetal wastage, placental
weight, or infant birth weight.
Obstet Gynecol. 1981 Apr;57(4):444-6.
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Severe fetal hemolysis with anti-D (85%),
anti-c (3.5%), anti-K (10%) and anti-E.
IgM antibodies do not cross the placenta A, B,
P1, Le(a,b), M, I
Antigens poorly expressed on fetal cells
Lu(b), Yt(a), VEL
Titers aid in prediction of fetal hemolysis
Platelet-specific alloantibodies are likely to
cause severe thrombocytopenia in fetuses
and neonates
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Moise KJ. Fetal anemia due to non-Rhesus-D redcell alloimmunization. Seminars in Fetal and
Neonatal Medicine (2008) 13, 207-214.
Rossi’s Principles of Transfusion Medicine, fourth
ed.
Transfusion therapy: clinical principles and practice
third ed. AABB 2011.
Technical manual, eithteenth ed. AABB 2014.