Download ABO IgM antibodies

1 Human Physiology-I (PHSL 205) Lab
4th Lab
Hakami,Hana A- 2010/2011
Blood groups, erythrocyte
sedimentation rate and
osmotic fragility of red blood
cells
Prepared by;
Hakami, Hana A
Viewed by;
Dr.Naseem Siddiqui
2 Human Physiology-I (PHSL 205) Lab
4th Lab
Hakami,Hana A- 2010/2011
First: Blood Groups (Blood Systems)
Introduction :
A total of 30 human blood group systems are now recognized by
the International Society of Blood Transfusion (ISBT). But there are 2
blood group systems are famous and we will study them, the ABO and
the Rh blood group systems.
ABO blood group system
The ABO system is the most important blood-group system in
human-blood transfusion. The associated anti-A antibodies and anti-B
antibodies are usually Immunoglobulin M, abbreviated IgM, antibodies.
ABO IgM antibodies are produced in the first years of life by
sensitization to environmental substances such as food, bacteria, and
viruses. The O in ABO is often called 0 (zero, or null) in other languages.
Blood Group Antigen
Antibody
A
A
Anti – B
B
B
Anti – A
AB
A,B
Absent
O
Absent Anti-A , Anti-B
The human ABO blood groups are an example of the multiple
allelic series. There are 3 alleles: allele A (IA) which produces antigen A ,
allele B (IB) which produces antigen B and allele i (Ii) which can not
produce any antigen .
Allele i is recessive to both alleles A and B, but allele A and B are
co-dominant to each other.
Blood Group Type (Phenotype)
A
B
AB
O
Rh blood group system
Possible Genotypes
IAIA or IAIi
IBIB or IBIi
IAIB
IiIi
The Rh system is the second most significant blood-group system
in human-blood transfusion.
3 Human Physiology-I (PHSL 205) Lab
4th Lab
Hakami,Hana A- 2010/2011
This blood group may be the most complex genetically of all blood
type systems since it involves 45 different antigens on the surface of red
cells. The most significant Rh antigen is the D antigen. It is common for
D-negative individuals not to have any anti-D IgG or IgM antibodies,
because anti-D antibodies are not usually produced by sensitization
against environmental substances.
However, D-negative individuals can produce IgG anti-D
antibodies following a sensitizing event: possibly a fetomaternal
transfusion of blood from a fetus in pregnancy or occasionally a blood
transfusion with D positive RBCs. Rh disease can develop in these cases.
Blood Group
Antigen
Antibody
Rh+
Present (D)
Absent
Rh
Absent
Produce anti-D (mainly absent)
This blood group is determined by only two alleles. The dominant
allele Rh+ (D) produces the Rh antigens, while the recessive allele Rh(d) does not produce the Rh antigen.
Blood Group Type (Phenotype)
Rh+ (positive)
Rh- (negative)
Detection of blood group type
Possible Genotypes
Rh Rh+ (DD) or Rh+Rh- (Dd)
Rh-Rh- (dd)
+
Detection of blood group types depends on the antigen-antibody
reaction (agglutination) occurring between the antigen on the red blood
cells and the antibodies in the serum.
Anti - A
Anti - B
= agglutination
Anti - D
= un-agglutination
Blood group
A+
AB+
BAB+
ABO+
O-
4 Human Physiology-I (PHSL 205) Lab
4th Lab
Hakami,Hana A- 2010/2011
ABO and Rh blood group types Detection :
Purpose :
To determine your ABO and Rh blood group types depends on the
antigen-antibody reaction (agglutination) by using :
1. Anti-A (containing antibody A)
2. Anti-B (containing antibody B)
3. Anti-D (containing antibody D)
Materials :
Alcohol swab , Lancet , Gloves , Anti-A , Anti-B , Anti-D , Blood Group
slide , Wooden sticks , Sharp container
Procedure :
In clean aseptic work area :
1. Sterilize your finger with alcohol swab and prick it by lancet .
2. Put drop on each hole in ABO & D slid.
3. Then, place drop of each antibody in special hole and mix it with
blood drop. Waite and observe occur any agglutination.
Results :
Write result of sample and study it :
Sample Name
Hana
Jawaher
Salha
Hind
Manal
Blood Group Types
O+
Sample Name
Areeg
Amal
Nada
Mariam
Mashael
Blood Group Types
OO+
5 Human Physiology-I (PHSL 205) Lab
4th Lab
Hakami,Hana A- 2010/2011
Second: Erythrocyte Sedimentation Rate
Introduction :
The erythrocyte sedimentation rate (ESR), also called a
sedimentation rate or Biernacki Reaction, is the rate at which red blood
cells sediment in a period of 1 hour. It is a common hematology test that
is a non-specific measure of inflammation. To perform the test,
anticoagulated blood is placed in an upright tube and the rate at which
the red blood cells fall is measured and reported in mm/h.
The ESR is increased by any cause or focus of inflammation. The
ESR is increased in pregnancy or rheumatoid arthritis, and decreased in
polycythemia, sickle cell anemia, hereditary spherocytosis, and
congestive heart failure. The basal ESR is slightly higher in females.
ESR can be done by two, methods;
I. Wintrobe Method : Requirements
1. Wintrobe tube with following features: Length 110 mm , Diameter 3
mm and Graduation of lower 100 rom from 0 to-100.
2. Anticoagulant: Double oxalate.
*Normal Values : Men o to 9 mm in first hour Women : a to 20 mm in
first hour.
II. Westergreen Method : Requirements
1. Westergreen tube. with following features : Length 300 mm ,
Diameter 2.5 mm , Graduation in mm from 0 to 200
2. Anticoagulant: 0.5 ml (3.13% sodium citrate solution).
* Normal Values : Men a to 5 mm in first hour Women : a to 7 mm in first
hour.
6 Human Physiology-I (PHSL 205) Lab
4th Lab
Hakami,Hana A- 2010/2011
Principle
When anticoagulant is added to the blood sample and then
allowed to stand in a tube, the red cells slowly settle down to the
bottom of tube leaving clear plasma above; The rate of sedimentation,
estimated under standard conditions, is called erythrocyte
sedimentation rate (ESR).
Erythrocyte Sedimentation Rate (ESR) Detection by use Westergreen
Method
Purpose :
To determine your ESR depends on Westergreen Method.
Materials :
Alcohol swab , Gloves , Syringes , Royal Blue Top Tube, Westergren
tubes, Stand , Sharp container
Procedure :
In clean aseptic work area :
1. The blood mixture (blood + anti-coagulant) drawn into
Westergreen tube up to the zero mark.
2. Tube is set upright in a stand with a spring clip on top and rubber
at bottom.
3. The level of the top red cell is read at the end of one hour.
Results :
Write result of sample and study it :
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7 Human Physiology-I (PHSL 205) Lab
4th Lab
Hakami,Hana A- 2010/2011
Third: Osmotic Fragility of Red Blood Cells
Introduction :
This test is based on the limit of the hypotonicity that red cells can
withstand. In some heamatological conditions the resistance of red
blood cells to hypotonic solution is reduced or increased. Spherocytes
are less resistant while hypochromic cells are more resistant.
Temperature and pH of salt solution greatly influence the test.
Osmotic fragility test (Dacie's Method)
Purpose :
To determine your Osmotic Fragility of Red Blood Cells
Materials :
Alcohol swab , Lancet , Gloves , Syringes , EDTA Top Blood Tube(or
Heparin blood tube) , Test tubes (small) , Graduated Pipettes,
Centrifuge, Photoelectric Colorimeter, Sensitive balance, Sodium
Chloride , Disodium Phosphate, Distilled Water, Sharp container
Procedure :
In clean aseptic work area :
1. Prepare Buffered Saline as flowing;
 Stock: Dissolved 90 g Sodium Chloride and 15 g Disodium
Phosphate in 1 liter of d.H2O (Distilled Water)
 Dilute 'stock' solution 10 times before use, which is equivalent
to 1 per cent saline.
 Make saline equivalent to 0.85, 0.80, 0.75, 0.60, 0.50, 0.45,
0.40, 0.35, 0.30, 0.20 and 0.10 per cent in 5 ml quantities.

2. Mix well again and centrifuge at 1500 rpm for 10 minutes.
3. Read the color of supernatant at 450 nm (or equivalent filter) in a
photoelectric colorimeter using saline as blank.
8 Human Physiology-I (PHSL 205) Lab
4th Lab
Hakami,Hana A- 2010/2011
4. Prepare a curve plotting per cent of heamolysis against salt
concentration.
Results :
Write result of sample and study it :
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Interpretation
It is useful to record the concentration of saline causing 50 per cent lysis,
i.e. Median Corpuscular Fragility (Normal = 0.4to 0.45 NaCl). For field
studies of beta thalassaemia, a single tube with 0.4 per cent can be
used. Decreased fragility may be observed in other condition also
including iron deficiency anaemia. Increased heamolysis at high
concentration is a positive test for hereditary Spherocytosis and
Hereditary Elliptocytosis.