chapter 4

Question 1

what chromosome is the gene for ABO & the gene for Hh on

Answer 1

ABO is on chromosome 9

Hh is on chromosome 19

Question 2

A, B & O alleles

Answer 2

ABO system has 3 possible alleles A, B & O

A& B are co dominant only differing by 7 nucleotides( 4 differing amino acids)

O is a recessive amorph - from a single nucleotide deletion resulting in a premature stop codon = non functional group

Question 3

coding for A& B antigens

Answer 3

genes code only for proteins no carbohydrates

- A & B antigens are carbohydrates so the gene codes for an enzyme ( protein) that makes the antigen

Question 4

whats another name for the H gene

Answer 4

FUT1 ( fucosyltransferase I)
the action of the enzyme produced
H is a carbohydrate too

Question 5

The H antigen

Answer 5

a carbohydrate
an amorph
the building block for A & B antigens

only one H is needed to produce H antigen ; if a person is hh they will not produce the enzyme therefore no H will be made

Question 6

Bombay & para- bombay

Answer 6

hh genotypes 
very rare ( <0.1% worldwide)

this type of person will only be compatible with other bombay blood groups bc they have no H ( required for antigens to form on ) they will have no antigens & therefore develop antibodies to all antigens

ex. A/A, h/h
will have anti -A , anti-B , anti-A,B , & anti-H

Question 7

Immunodominant sugars

Answer 7

each A, B & H gene codes for an enzyme that adds an immunodominant sugar which characterizes the corresponding antigen

ANTIGEN; SUGAR ( ENZYME )

H ; L- fucose ( alpha-1,2-L-fucosyltransferase )

A; N-acetyl- D - galactosamine ( alpha-1,3-N-acetyl- galactosaminyltransferase )

B; D-galactose ( A-1,3-D-galactosyltransferase )

Question 8

Type 1 & Type 2 ABH precursors

Answer 8

Type 1 makes up the majority of precursors found in the gut, plasma & secretions(saliva, urine, milk)
-have a ß1-3 linkage

Type 2 precursor substances are found mainly on the RBCs & & come in the saliva
- have a ß1-4 linkage

Question 9

frequency of the 4 most dominant ABO phenotypes

Answer 9

O……………….46%
A………………..42%
B………………..9%
AB………………3%

Question 10

A1 & A2 % & similarities

Answer 10

variations in the A gene lead to subgroups of A or AB people; 80% are A1 or A1B with most of the remaining 20% being A2 or A2B

They both have the same immunodominant sugar A1 is dominant, A2 is recessive

Question 11

Qualitative & Quantitative differences in A1 & A2

Answer 11

A being more efficient at converting H to A means:

Quantitatively this leads to more A on A1 cells than A2

Qualitatively A1 have an additional epitope, therefore A2 can develop an anti- A ( no such thing as anti-A2)

~1-2% of A2 will develop an anti-A1
~20-25% of A2B2 will produce an anti-A1

Question 12

Anti - A1 significance

Answer 12

Anti-A1 is a naturally occurring, IgM ( cold) class antibody that is usually clinically insignificant 
however in some cases it can show reactivity at 37 degrees, leading to a reduced lifespan of transfused A1 cells 

note: anti-A1 can be found normally in O & B individuals

Question 13

other subgroups of A

Answer 13

most to least A antigen expressed:
A1> A2> A3> Ax> Aend> Ael

A3 will show Mixed field with A/A & A/A,B
Ax will show a weak reaction with A/A & A Rxn with A/A,B

Question 14

unexpected cold antibodies vs Anti-A1

Answer 14

reactions with A1 cells could be anti-A1 or an unexpected cold antibody

testing the plasma of a patient with a suspected anti-A1 against O cells as a negative control will help confirm or rule out Anti-A1 presence
if (+)= O person
if (-)= A2 person

Question 15

Reagents of Anti-A1 & Anti-H made from

Answer 15

some reagents are made form lectins ( extracts from plant seeds)
Anti- A1 is from the seed of Dolichos biflorus
Anti-H is from the seed of Ulex europaeus

Question 16

subgroups of B

Answer 16

extremely rare

sometimes patients/ donors with a very weak subgroup of B will produce a weak anti-B

Question 17

landsteiner’s lsw

Answer 17

in the ABO blood group system, unlike other blood group systems, everyone develops antibodies to antigens that aren’t on their own RBCs, without exposure to foreign RBCs.

they may result from environmental exposures, therefore they are called naturally occurring antibodies

• babies aren’t born with ABO antibodies, they develop them overtime ( tend to develop after ~3 months)
• ** this id why we don’t do reverse ABO typing on newborns
• ABO antigens are sufficiently developed at birth to be reliably detectable

Question 18

ABO antibodies class and thermal range

Answer 18

ABO antibodies are IgM ( react best at cold temps) , bind complement & are capable of intravascular hemolysis

ABO have a large thermal range & will also react at body temp.
ABO antibodies can also be IgG & IgA.
IgG antibodies are esspecually prevalent in group O people( in particular anti-A,B)

Question 19

Secretors

Answer 19

A, B & H antigens can be found in plasma of individuals with respective antigen, those antigens will only be found in secretions ( urine, saliva, milk) of about 80% of these people( if they have the secretor gene)

ABH antigens in secretions are primarily glycoproteins ( except for in milk & urine they are oligosaccharides)

Secretors have at least one inherited Se gene. The Se gene codes for a fucosyltransferase enzyme; similar to H gene, except Se gene product preferentially acts on type 1 precursors while H gene acts on the 2.

Se gene makes fucosyltransferase which converts type 1 precursor to Se

Se gene ( FUT2) is on chromosome 19, near H gene

Question 20

non secretors

Answer 20

the se gene is like the h gene; its an amorph

people who are se/se wont produce A, B or H in their secretions

~20% of populations are non secretors & don’t produce functional fucosyltransferase; the type 1 precursor substance secreted into their fluids is not converted to H substance & therefore cannot convert into A or B antigens, regardless of their ABO genotype

Question 21

ABH antigens in secretions on RBCs & in plasma

Answer 21

ABH antigens in secretions are primarily glycoproteins ( except for in milk & urine they are oligosaccharides; sugars not attached to either proteins or lipids)

ABH antigens can adsorb onto platelets & lymphocytes

granulocytes & monocytes do not possess ABH antigens

secretor or non secretor you will have ABH antigens on RBCs

Question 22

Lewis antigens

Answer 22

Lewis antigens are not intrinsic to RBC cell membrane but are adsorbed from plasma onto the RBC. This begins after birth, the full expression of lewis antigens isn’t reached until ~age 7
( Lewis antigens shouldn’t be phenotype on neonates or children)

Transfused cells acquire the lewis type of the recipient

during Pregnancy the production of lewis antigens can diminish, resulting in a transitory Le(a-b-) phenotype; lewis phenotyping shouldn’t be done on pregnant people
any antibodies produced during pregnancy to lewis will diminish after

Question 23

Lewis gene

Answer 23

Lewis antigen results form the presences of the lewis gene (FUT3) with or without secretor gene. if Le gene is present, an L-fucosyltransferase enzyme acts on type 1 chains, rsultingin Le^a antigen. Le^a is then absorbed onto RBC cell membrane; phenotype Le(a+b-)

Question 24

Secretor gene & Lewis gene

Answer 24

If the secretor gene is also present with the lewis gene, the secretor transferase enzyme adds fucose to type 1 chains & then the lewis transferase enzyme adds another fucose to this H type 1 chain, forming Le^b antigen

Le^b will be more abundant but with Le^a & Le^b will be present in body fluids. RBCs will preferentially absorb Le^b & the phenotype will be Le(a-b+)

RBC of people with no Le genes will type Le(a-b-)

Question 25

Lewis antibodies

Answer 25

lewis antibodies are usually IgM, react best at room temperature & occasionally bind compliment, which results in vitro hemolysis

No lewis antibodies are produced by people whose RBC type is Le(a-b+) because their body fluids contain both lewis structures
People who type Le(a+b-) can produce anti-Le^b
people who are Le( a-b-) can produce anti-Le^a &/or anti-Le^b

anti- Le^bH can be produced by people whose RBCs lack Le^b & have a small amount of H antigens
- A1 cells Le( a+b-)

Question 26

Saliva inhibition test

Answer 26

the presence or absence of A, B , H , Le^a &/or Le^b antigens in saliva can be determined using the saliva inhibition test

saliva is collected from the patient and incubated with suitable diluted anti-A, anti-b, anti-H , anti-Le^a & anti-Le^b, each in separate tubes
if present it will bind to the antibody

then cells with the corresponding antigens are added to each tube
if there is no agglutination, the antibodies added have already bound to the antigens in the persons saliva ( meaning they are positive for that antigen )

if there is agglutination, the antibodies didn’t have an antigen to bind to originally so they can bind to the regent cell added in the last step

if agglutination occurs in all tube of ABO( negative results )
this person is a non secretor & therefore ABO is unknown

if agglutination overs in both lewis cells tubes ( negative results)
this person is lewis negative & therefore secretor status is unknown

Question 27

controls for saliva inhibition test

Answer 27

controls must be run to ensure the procedure worked as expected, the antisera worked as expected & the dilution didn’t cause ant of the negative results
this usually includes a Dilution control tube *
- containing the diluted regent, appropriate RBCs & saline

• a positive control tube should contain diluted regent, appropriate RBCs & the saliva of a known secretor

the dilution control tube should be neg( agglutinate )
the positive control tube should be pos ( no agglutination)

Question 28

True Bombay

Answer 28

bombay phenotype results from hh & sese genotypes.
These people lack H substance both on their RBCs & in their secretions
they will lack A& B antigens, regardless of their ABO genotype, in both as well since there is no H precursor

during ABO testing a bombay will appear as a group O
during antibody testing bombay will produce an anti-H that reacts with all RBCs except those of bombay phenotype

therefore the plasma of a bombay person will react with screening cells that are typically group O

Question 29

para-bombay

Answer 29

some people appear bombay but have very small amounts if A &/or B antigen on their RBCs
their genotype is hh but they possess at least one Se gene
the Se gene produces fucosyltransferase that acts on type 1 precursor substance in secretions, this type 1 H substance formed from the Se gene can convert to A or B antigens

para-bombay will have ABH antigens in their secretions & plasma
sometimes they will have small amounts of AB &/or H antigens that have absorbed onto their RBCs from their plasma

they will possess a weak reaction & they only have type 1 antigens (absorbed) not intrinsic type 2 antigens
type 2 requires the H gene

Question 30

anti-H

Answer 30

Bombay will make a naturally occurring anti-H as well as anti-A, anti-B & anti-A,B

Anti-H is an IgM that reacts strongly at 37 degrees
can cause intravascular hemolysis

Parabombay will make H substances in they secretions & wont produce anti-H

Question 31

I system

Answer 31

there are 2 carbohydrate antigens I & i
at both i antigen is expressed on RBCs
most is converted to I in the firth 18 months

antibodies in this system are usually IgM autoantibodies
auto anti-I is a common cold autoantibody
- reactivity can increase due to Mycoplasma pneumoniae
- clinically significant anti -I may be detected in adults who have not converted i antigen

detection of anti-i has been associated with infectious mononucleosis

Question 32

Cold auto agglutinins

Answer 32

Cold auto agglutinins that react at room temp or lower & react with H ( anti-H ) or anti I&H ( anti-IH) are relatively common & insignificant
- these are different than the Bombay anti-H which are highly significant

anti-H or anti-IH an be found in the plasma of para-bombay people but much more commonly in patients of the A1 or A1B phenotype
bc A1 have the least amount of H on their cells

anti-IH will exhibit little or no reaction with cord cells bc they have i and no I antigen.
anti -IH will react strongly with O cord cells ( bc of H antigen)but not at all with A cord cells

Question 33

most to least H antigens on cells

Answer 33

O> A2 > B > A2B > A1 > A1B

Question 34

ABO grouping

Answer 34

consist of 2 parts :

1. testing for antigens on RBCs ( forward grouping )
   - combining unknown patients cells with know reagent antibody
2. testing for antibodies in plasma ( reverse grouping )
   - combining unknown patients plasma with known ABO cells

Question 35

ABO discrepancies

Answer 35

usually involve forward & reverse reactions not agreeing.
mixed field, or obtaining different results from past results are also discrepancies

when identifying discrepancies, weak reactions are more likely to be aberrant than strong ones ( look at weak reactions to find answers)

Question 36

missing or weak reactions in the forward group may indicate :

Answer 36

the patient is a subgroup of A or, less frequently, a subgroup of B . The very rare cis AB genotype ( both A& B gene are on the same chromosome with no ABO gene on the other) may also result in a weak expression

leukaemia or other malignancies may result in alterations of th A or B antigen on the RBCs. they wont react normally with antisera

Some people have excessive amounts of soluble A &/or B antigen in their plasma. if plasma suspended cells are used the antigens in the plasma can neutralize the reagent antiserum, resulting in days - neg reaction ( using washed cells avoids this )

Question 37

enhancing a weak ABO reaction

Answer 37

modifying test; extended room temp or 4 degree incubation, enzyme rating the patients cells ( as ABO antigens are enhanced by enzymes)

saliva inhibition test may be used ( if patient is a secretor )

genetic analysis can be performed

Question 38

mixed field agglutination

Answer 38

when sample has 2 different cell populations

most often caused by transfusion with non-ABO identical cells
most commonly a non group O patent is given an emergency transfusion of group O cells ( to avoid intravascular hemolysis)

other causes:
ABO non identical bone marrow or hematopoietic progenitor transplant

weak ABO subgroups such as A2

Chimersim - fraternal twins exchange blood- forming cells in utero or when 2 different populations of cells ( donor & recipient ) reside together in the bone marrow after transplantation

mosaicism -occurs when 2 sperm fertilize 1 egg ( dispermy) resulting in 2 distinct cell populations

mixed field due to chimerism & mosaicism exist throughout patients life & are considered self, so ABO antibodies aren’t made towards them

Question 39

missing or weak reactions in the reverse group

Answer 39

discrepancies in the reverse group are less common
the first step when seeing a discrepancy in the reverse is to check the patients age
-if the patient is younger than 4 month the patient shouldn’t be tested fro reverse bc they haven’t developed antibodies yet
- elderly also exhibit decreased antibody concentrations

other reasons for discrepancies :

• ABO antibodies may be diluted to a non detectable level if the patient has received therapeutic plasma exchange therapy
• patients who are on aggressive immunosuppressant therapy can have decreased expression of there antibodies
• patient who has a non identical ABO bone marrow transplant
• certain diseases such as agammaglobulinemia, hypogammaglobulinemia,& some leukemias & lymphomas

Question 40

resolution techniques for discrepancies

Answer 40

serum/cell ration : in a tube testing, add 1-2 additional drops of patient/donor plasma can sometimes enhance a weak reaction

temperature : ABO antibodies are IgM & are generally cold -reactive. the reverse can be incubated for 15-30mins at room temp. if that doesnt resolve it , the temp can be lowered to 4 degrees, where group O cells & an auto control should be tested as well to control for the presence of other cold antibodies

enzymes: proteolytic enzyme treatment of the reverse grouping cells can soemtimes enhance a weakly reactive ABO antibody. can be done using ficin ( fig), papain ( papaya ) bromelin ( pineapple) or trypsin(hogs stomach)
controls of group O & autologous cells should be enzyme-treated & tested at the same time to ensure any reactivity is due to ABO antibodies

Question 41

unexpected positive reactions in the forward group