HLA Antigens and other Platelet and Leucocyte Antigens and their Role in Blood Transfusion

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Antigens expressed on leucocytes and platelets
These include:
• HLA antigens
• Blood group antigens on other cells
• Other antigens
Role of HLA in Blood Banking
The human leucocyte antigen (HLA) system is the name of the
major histocompatibility complex (MHC) in humans. The locus
contains a large number of genes related to immune function.
HLA antigens are major histocompatibility complex (MHC)
molecules and have the following functions and characteristics:
• The HLA system is highly polymorphic, there are many
alleles of each MHC gene in the population.
• This complex contains 35–40 genes and is clustered on
chromosome 6p21.3.
• MHC molecules have key roles in regulating T cell mediated
immune response.
• Display peptide fragments of proteins for recognition by
antigen-specific T cells.
HLA Antigens
• Class I A, B and C antigens are expressed on all the cells
except spermatozoa, trophoblastic cells and RBCs. They are
expressed on platelets and leucocytes also. On RBC mem-
brane, these can be adsorbed from the plasma.
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• Platelets possess mRNA encoding HLA class I molecule and
are capable of synthesizing them.
The proteins encoded by certain genes are also known as
antigens, the major HLA antigens are essential elements for
immune function. HLA antigens corresponding to MHC class I
(A, B, C) present peptides from inside the cell (including viral
peptides, if present). These peptides are produced from
digested proteins that are broken down in the proteosomes. In
general, the peptides are small polymers about 9 amino acids
in length. These antigens attract CD8 positive cytotoxic T cells
that destroy them.
Class II antigens are on lymphocytes, monocytes, macro-
phages, activated T cells and Langerhan’s cells.
HLA antigens corresponding to MHC class II (DP, DM,
DOA, DOB, DQ and DR) present antigens from outside of the
cell to T-lymphocytes. These particular antigens stimulate
the multiplication of T-helper cells, which in turn stimulate
antibody-producing B-cells to produce antibodies to that
specific antigen. Self-antigens are suppressed by suppressor
T-cells.
HLA Antibodies
HLA antibodies are typically not naturally occurring. They are
formed as a result of an immunologic challenge of a foreign
material containing non-self HLAs via blood transfusion,
pregnancy (paternally-inherited antigens), or organ or tissue
transplant.
The HLA system has important implications in transfusion
therapy. Sensitization of recipients with consecutive formation
of HLA antibodies and is the cause for thrombocytopenia and
may cause febrile transfusion reactions.
Blood Group Antigens on Other Cells
• A, B antigens are present on lymphocytes and are detected
by lymphocytotoxicity tests. Lewis antigens are acquired
from plasma.
• A, B and H antigens are on platelets.

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Essentials of Blood Banking and Transfusion Medicine
• Epidermal cells, cells of amniotic fluid, sinusoidal cells of
spleen, spermatozoa, endothelial cells have A, B and
H antigens.
Other Antigens
• Human neutrophil antigens (HNA)
• Platelet specific antigens
Human Neutrophil/Granulocyte Antigens (HNA)
• Neutrophil specific antigens like HNA1 (1a and 1b), HNA2
and SH are present on the neutrophils.
Other antigens on neutrophils are HNA 2a, HNA 3a, HNA
4a and HNA 5a.
• Antibodies develop when the person lacks these antigens,
antibodies to HNA1-1a and 1b are often implicated.
These are recognised by Fc portion of the IgG.
Granulocyte specific antibodies (HNA-1a and 1b) and HLA
class I antibodies are responsible for:
• Neonatal allo and isoimmune neutropenia
• Febrile non-haemolytic transfusion reactions
• Transfusion related acute lung injury (TRALI)
• Auto-immune neutropenia
In platelet packs and RBC packs, the granulocytes release
cytokines with storage, hence pre-storage leuco-reduction is
preferred.
Alloimmunization
Leucocytes in blood products are the major stimulus for allo-
immunization. The number of leucocytes in different blood is
given below.
PRBCs: 2–5 × 109 WBCs.
Random platelets: 0.5–2.5 × 108 WBCs.
Leucoreduced: Less than 5.0 × 106 WBCs.
Neonatal alloimmune neutropenia is similar to haemolytic
disease of newborn with following possibilities:
• Passively transmitted maternal antibodies.

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• Mother’s serum has potent IgG antibodies against granulo-
cyte specific antigens.
• Usually 1st child is affected.
Neonatal Iso-immune Neutropenia
This develops in subjects in whom the glycoprotein is absent
on the granulocytes due to deletion of the gene, may form
strong antibodies.
Infants born to these mothers will suffer from iso-immune
neutropenia.
Febrile Non-Haemolytic Transfusion Reactions (FNHTRs)
For details refer to transfusion reactions.
As a remedy for these reactions, leucocyte reduction is
preferred.
Pre-storage leucocyte reduction is preferred with less than
5 × 106 leucocytes and 85% RBCs to be retained.
Leukofilters with
• Barrier filtration: Pore size smaller than WBCs.
• Cell adhesion are used.
Note: For details refer to transfusion reactions.
Transfusion Related Lung Injury (TRALI)
For details refer to transfusion reactions.
Transfusion-Associated Graft-Versus-Host Disease (TA-GVHD)
For details refer to transfusion reactions.
Tests done to detect antibodies:
• Granulocyte agglutination
• Immunofluroscence (IF)
• Chemiluminescence
• Monoclonal antibody specific immobilisation of granulocyte
assays.
Granulocyte agglutination technique:
• Granulocyte suspension is prepared by dextran sedimenta-
tion followed by centrifugation on ficoll-hypaque.

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Essentials of Blood Banking and Transfusion Medicine
• The RBCs are lysed with ammonium chloride or distilled
water or WBCs are obtained by double density gradient
centrifugation.
• Agglutination carried out on microplates.
• They are agglutinated by cross-linking of cells by IgM or
IgG antibodies which adhere to Fc receptors on granulo-
cytes.
• Granulocyte specific antibodies and HLA A, B and C
antibodies can be detected. HLA A, B and C antibodies can
be better detected with lymphotoxicity tests.
Platelet Antigens
HLA antibodies to HLA class I antigen after transfusion are
the major cause of antibody mediated refractoriness to platelet
transfusion, febrile transfusion reactions and occur in preg-
nancy.
Allo-antibodies to HLA class I antigen on platelets and
antibodies to platelet specific antigens occur in neonatal allo-
immune thrombocytopenia (NAT/NAIT), post-transfusion
purpura (PTP), passive alloimmune transfusion (PAT) and
transplantation associated purpura. Anti-platelet specific
antibodies are detected in 8% of recipients of multiple platelet
transfusions.
Post-transfusion Purpura
• Rare complication.
• Rapid onset of thrombocytopenia as a result of anamnestic
production of platelet allo-antibodies.
• Usually occurs in multi-parous women who do not have the
antigen.
• Multiple transfusion recipients.
• Allo-antibodies in the recipient clear the donor platelets.
Alloimmunisation to Platelet Specific Antigens
Post-transfusion purpura is a mystery of alloimmunisation.
• Sudden decrease in platelet counts 7–10 days post-trans-
fusion.

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• Recipient is negative for the platelet antigen (usually PLA1
or Bra). Usually under diagnosed or diagnosed after severe
bleed.
Mechanism
Majority of population is PLA1 positive and thus majority of
donors are PLA1 positive. Pl-A1 is a soluble antigen. When Pl-A1
negative individuals receive PLA1 positive products, Pl-A1
antigens bind to PLA1 negative platelets. Pl-A1 is recognized
as a foreign and antibodies are produced. Antibody binds to
PLA1 antigen. Pl-A1 antigen is removed from circulation.