Anti N antibody belongs to the MNS blood group system. Usually, anti N antibodies are naturally occurring, cold agglutinins. Clinically significant anti N antibodies have also been reported. We report here a case of autoantibody with anti N specificity presenting with severe autoimmune haemolytic anaemia.

1. A case of severe autoimmune haemolytic anaemia due to
clinically significant auto anti-N antibody

2. Case Report
A case of severe autoimmune haemolytic anaemia
due to clinically significant auto anti-N antibody
R.N. Makroo a,
*, Dhaval Fadadu b
, Mohit Chowdhry c
, Aakanksha Bhatia d
,
Prashant Karna e
a
Director & Sr. Consultant, Department of Transfusion Medicine, Molecular Biology and Transplant Immunology,
Indraprastha Apollo Hospitals, India
b
DNB Resident, Department of Transfusion Medicine, Molecular Biology and Transplant Immunology,
Indraprastha Apollo Hospitals, India
c
Consultant, Department of Transfusion Medicine, Molecular Biology and Transplant Immunology,
Indraprastha Apollo Hospitals, India
d
Junior Consultant, Department of Transfusion Medicine, Molecular Biology and Transplant Immunology,
Indraprastha Apollo Hospitals, India
e
Department of Transfusion Medicine, Indraprastha Apollo Hospitals, India
1. Introduction
MNS blood group system is the second largest blood group
system after ABO system, containing approximately 46
antigens. Antibodies of M and N blood group are associated
with variable clinical significance as both IgG and IgM types of
antibodies are encountered. Frequency of M+N+ phenotype is
54.1% in Indian population.1
Among antibodies of MNS blood
group system, anti M is a relatively common, naturally
occurring antibody,2
whereas anti N is quite rare as compared
to anti M.2,3
. Generally anti M and anti N are naturally
occurring, cold reactive IgM saline agglutinins, and are usually
not active at 37 8C; therefore, they are not clinically signifi-
cant.4
However, reports of clinically significant antibodies of
MNS blood group are also available in literature.5
Anti N has
been implicated as the cause of haemolytic transfusion
reactions (HTRs).6
We report a case of autoantibody with anti
N specificity which is reactive at 37 8C.
2. Case report
A 45-year-old male presented to Emergency department with
the chief complaint of fever and haematuria for 4 days, sore
a p o l l o m e d i c i n e 1 2 ( 2 0 1 5 ) 1 3 8 – 1 4 0
a r t i c l e i n f o
Article history:
Received 10 April 2015
Accepted 1 May 2015
Available online 15 June 2015
Keywords:
Haemoglobin
Immucor
Galilio
a b s t r a c t
Anti N antibody belongs to the MNS blood group system. Usually, anti N antibodies are
naturally occurring, cold agglutinins. Clinically significant anti N antibodies have also been
reported. We report here a case of autoantibody with anti N specificity presenting with
severe autoimmune haemolytic anaemia.
# 2015 Published by Elsevier B.V. on behalf of Indraprastha Medical Corporation Ltd.
* Corresponding author. Tel.: +91 9810170969.
E-mail address: makroo@apollohospitals.com (R.N. Makroo).
Available online at www.sciencedirect.com
ScienceDirect
journal homepage: www.elsevier.com/locate/apme
http://dx.doi.org/10.1016/j.apme.2015.05.004
0976-0016/# 2015 Published by Elsevier B.V. on behalf of Indraprastha Medical Corporation Ltd.

3. throat with cough for 2 weeks and urinary hesitancy for 3
weeks. He was earlier admitted to a local hospital where he
was found to have elevated blood sugar level and severe
anaemia Coombs positive. He was brought to our hospital for
further management.
On first day, his haemogram revealed, haemoglobin level
6.1 g/dl, TLC was 45,300/mm3
and reticulocyte count was
0.1%, LDH level 2800 IU/L, total bilirubin 2.7 mg/dl in the lab
reports. During further work up, both ANA and anti dsDNA
were negative, ruling out SLE. Any significant drug history
which could lead to haemolysis was elicited. Haemoglobin
level continued to decline, and at the level of 5.2 g/dl, 2 units
PRC transfusion was required, and a sample for Group and
Screen was received by the department of Transfusion
Medicine.
As per the departmental protocol, blood grouping was
performed on an automated SPRCA platform (Immucor), and
when a group discrepancy was encountered, blood grouping
was done with tube technique, and results are shown in Table 1.
Forward grouping suggested blood group B, Rh (D) positive
but reverse grouping was giving agglutination (4+) with all the
3 cells (A1 cells, B cells and O cells). The reverse grouping was
repeated with incubating the test tubes at different tempera-
tures (room temperature, 4 8C and at 37 8C) and the result is
demonstrated in Table 2.
Meanwhile, antibody screening was done on Capture-R
Ready Screen using SPRCA technique on fully automated IH
analyser (Galilio, Immucor, US) using a 4-cell screen panel.
Results are shown in Fig. 1.
Auto control was performed on conventional tube tech-
nique with immediate spin (IS) method which gave positive
reaction (2+). Direct Coombs test using monospecific (IgG)
Coombs sera gave weak (1+) reaction and with monospecific
C3d, Coombs sera gave (2+) reaction.
As per the protocol, 4 EDTA blood samples for antibody
identification were received. Patient did not have any
transfusion history or operative history before. Using 16-cell
(Capture-R Ready-ID) and 10-cell panels (Pano Cell-10; Immu-
cor), antibody identification was performed.
As it is evident that the 16-cell panel points towards a
possible anti N, however, the 10-cell panel is pan positive both
at IS as well as at 37 8C (Fig. 2).
In view of positive auto control and pan positive 10-cell
panel, adsorption was attempted. Due to extreme haemolysis
in the sample, sufficient intact red cells could not be retrieved
for performance of auto-adsorption. Multiple allo-adsorptions
were done using allogenic cells (R1R1, R2R2 and rr) and antibody
screening was performed with adsorbed plasma. Antibody
screen with adsorbed plasma was negative.
According to screening and 16-cell panel, it was concluded
that patient had auto-antibodies reacting at wide thermal
range (4–37 8C) with possible anti-N like specificity.
Antigen typing of the patient revealed N positive pheno-
type. The patient had no history of blood transfusion and any
operative history, which supports the absence of alloantibody.
It was, therefore, concluded that the antibody in question
was an autoantibody possibly auto anti N.
Approximately 43 units were typed for n antigen, of which,
4 units were found as N negative and were compatible with
patient's serum in AHG cross matching.
The patient was transfused 1 unit initially followed by 2
units after 3 days. There was no evidence of HTR or aggravated
haemolysis in the patient. He was also given 2 doses of
rituximab. After 3 units of PRC transfusion his haemoglobin
level reached 8.9 g/dl.
3. Discussion
Anti N antibody belongs to the MNS blood group system, and
have been reported with variable frequency in literature.
Majority of anti-N are naturally occurring cold reactive IgM
Table 1 – Forward grouping.
Anti A Anti B Anti AB Anti D
Reaction – 4+ 4+ 4+
Table 2 – Reverse grouping.
Temperature (8C) A B O Autocontrol
Room temperature 4+ 4+ 4+ 2+
4 4+ 4+ 4+ 2+
37 4+ 4+ 4+ 2+
[(Fig._1)TD$FIG]
Fig. 1 – Results of screening cell panel (Capture-R Ready Screen).
a p o l l o m e d i c i n e 1 2 ( 2 0 1 5 ) 1 3 8 – 1 4 0 139

4. antibodies. However, IgG type anti N,7
reactive at 37 8C has also
been reported. There have also been reports of clinically
significant anti N antibodies resulting in HTRs8
as well as
haemolytic disease of foetus and new born.
This anti N antibody detected in our case was reactive over
a wide thermal range and in AHG phase with IgG component,
and therefore can be considered as potentially clinically
significant.5
Inaddition tothisand inviewofthefact thatthepatient gave
no history of previous transfusions, dialysis and his phenotyp-
ing showed an N positive phenotype. The antibody in question
can be considered to be a clinically significant auto anti N.
Auto anti N antibodies have been reported earlier, and have
also shown to be associated with severe autoimmune
haemolytic anemias,8
like in our case where the patient
presented with clinical and laboratory features of autoim-
mune haemolysis.
4. Conclusion
Auto antibodies with anti N specificity are uncommon, but
may be clinically significant; also they result in severe
autoimmune haemolytic anaemia. Immunohaematology lab-
oratories especially in higher and reference centres should be
equipped to identify such antibodies, and provide necessary
transfusion support.
Conflicts of interest
The authors have none to declare.
r e f e r e n c e s
1. Makroo RN, Bhatia A, Gupta R, Phillip J. Prevalence of Rh, Duffy,
Kell, Kidd and MNSs blood group antigens in the Indian blood
donor population. Indian J Med Res. 2013;137:521–526.
2. Perrault R. Naturally-occurring anti-M and anti-N with special
case: IgM anti-N in NN donor. Vox Sang. 1973;24:134–149.
3. Race RR, Sanger R. Blood Groups in Man. 6th ed. Oxford:
Blackwell Scientific Publications; 1975.
4. Mollison PL, Engelfriet CP, Contreras M, et al. Blood Transfusion
in Clinical Medicine. 10th ed. Oxford: Blackwell Science; 1997.
5. Makroo RN, Arora B, Bhatia A, Chowdhry M, Luka RN. Clinical
significance of antibody specificities to M, N and Lewis blood
group system. Asian J Transfus Sci. 2014;8:96–99.
6. Ballas SK, Dignam C, Harris M, Marcolina MJ. A clinically
significant anti-N in a patient whose red cells were negative
for N and U antigens. Transfusion. 1985;25:377–380.
7. Leger RM, Calhoun L. Other major blood group system. In:
Harmening DM, ed. In: Modern Blood Banking and Transfusion
Practices 5th ed. New Delhi: Jaypee Publisher; 2008:165–167.
8. Immel CC, McPherson M, Hay SN, Braddy LR, Brecher ME.
Severe hemolytic anemia due to auto anti-N.
Immunohematology. 2005;21(2):63–65.
[(Fig._2)TD$FIG]
Fig. 2 – Results of 16-cell panel (Capture-R Ready-ID).
a p o l l o m e d i c i n e 1 2 ( 2 0 1 5 ) 1 3 8 – 1 4 0140

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