Chapter 5 The Cross Matching

CHPTER -SEVEN
The Cross-Match
(compatibility testing)
CH
Content
 Cross matching
 Types of Cross Match
 Steps for compatibility testing

 Choice of Blood for cross-match
 Procedure for cross-match

Learning Objectives
At the end of this chapter, the student will be able

to:
 Explain the cross-match and its primary purpose.
 Describe the constituents of the major and minor
cross match.
 Select appropriate blood for cross-match.
 List the types of antibodies that can be
encountered at various phases of a cross-match.
 Perform cross matching
7.1 Cross-Matching
 It is a test to determine the compatibility between

recipient’s blood and donor’s blood.
 It is a procedure performed before transfusion to select
donor’s blood that will not cause any adverse reaction,
(hemolysis /agglutination)
 The test is important to see whether there is danger or
not if the donor’s blood is transfused to the recipient.
 helps the patient to receive maximum benefit from the
transfusion of red cells
Cross-Matching…
 Main purpose is to prevent a transfusion

reaction that may result due to:
 Unexpected antibodies in the patient’s /donor’s
serum
 Some ABO incompatibilities and
 Technical or labeling errors (clerical errors)

Cross Matching…
 Transfusion reaction – is the term used to

describe the clinical effects caused by the
destruction of red cells of recipient by the
donor’s antibodies or a donor’s red cell’s by the
recipient antibodies during an incompatible
transfusion.
Cross-matching…
 However, it will not:

 prevent immunization of the patient
 guarantee normal survival of transfused erythrocytes
 detect all unexpected antibodies in a patient’s serum.

7.2 Types of Cross Match
Major and Minor crossmatch

Major cross-match:
 involves mixing recipient’s serum with the donor’s red
cells.
 is much more critical for assuring safe transfusion

than the minor compatibility test.
 called major b/c the Abs in the recipient’s serum are

most likely to destroy the donor’s RBC
Types….
Minor cross match:
 Involves mixing the donor’s serum with patient’s
red cells
 Called minor because

 any Abin the donor’s serum will be diluted by the large
volume of the recipient’s blood
 thedestructed RBCs of the patient may be

compensated by the transfused RBC of the donors
7.3. Steps for compatibility testing
 Accurate Patient Identification
 Proper sample collection and handling
 Review of the recipient’s past blood bank records
 Careful ABO/Rh determination
 Antibody screening of the recipient (cross matching of the

donor unit).
Steps for…
 In cases when the recipient possess a clinically

significant Antibody, donor units must be:
 Screened for the corresponding Ag and should be

negative
 Cross- matched
Steps for….
 Finally, during the actual transfusion :
 careful observation of the recipient’s vital signs and
 posttransfusion hematocrit and Heamoglobin levels

must be considered.
7.4.Choice of Blood for cross-match
 The blood selected for cross-match should be of

the same ABO and Rh (D) group as that of the
recipient.
 However, Rh positive recipients may receive

either Rh positive or Rh negative blood.
Choice of…
 Whenever possible blood of the patients own

blood group should be given.
 Otherwise the following rules should be applied.

Group A patient.
- Should receive group A blood, if not available group O
Group B patient.
- Should receive group B blood, if not available group O
Choice of…
Group O patient.
 Can only receive group O blood
Group AB patient.
 Should receive from group AB, if not possible can
receive blood from group A,B, and O.
Choice of…
 When cross-matching is carried out, the serum

is tested against the cells.
 The serum should be fresh, that is not more than

48hours old.
Choice of…
 When deciding on methods for cross-matching,

the following conditions are required for Ag-Ab
reactions.
 The right Temperature.

 Suitable surrounding medium
 Antigen-Antibody ratio etc..
Choice of…
 The safe cross-matching of blood requires that

the donor’s cells be mixed with the patient’s
serum in three separate tubes, using :
1. Saline
2. Albumin
3. Anti-human globulin reagents
1. Saline tube
 The red cells from the donor are suspended in

saline and mixed with the patient’s serum .
 show the presence of any complete antibodies
 Agglutination in the saline tube is usually caused
by:
 anti-A or anti-B antibodies and
 Occasionally by Lewis, MNSs, Lutheran and kell
antibodies.
2. Albumin tube
 The red cells from the donor’s suspended in

saline, are mixed with the patient’s serum, and
albumin is added.
 The tube is incubated at 370C
 shows the presence of any incomplete
antibodies
 the antibodies react in albumin or any other
protein medium
Albumin tube…
 Agglutination in the albumin tube is often caused

by:
 the rhesus antibodies,

 Lewis, MNSs, Lutheran and P antibodies, and
 occasionally by anti-kell.
 Reaction caused by anti- A or anti- B antibodies

usually occur in albumin as well as in saline.
3. Anti-human globulin tube
 A more concentrated suspension of red cells is

mixed with the patient’s serum and incubated at
370C and then AHG is added.
 A postive test detects the presence of antibodies

of:
 rhesus, kell, kidd, S and Lewis
 Anti globulin is essential for detection anti-Duffy

7.5. Procedure for cross-match
7.5.1.Standard cross-match
 Is cross-match that is performed in three tubes
(Saline, albumin and AHG) within 45 to 60
minutes
Clinical significance
 detects unexpected (irregular) antibodies in the
recipient/ donor serum
Cross match (Standard)…
Principle
 Serum of the recipient / donor is tested against

the red cells of the donor/ recipient under
different conditions in order to establish their
compatibility
Cross match….
Type of specimen
 Serum (plasma) not older than 48 hrs

 Washed cells (20-30% and 2-5%)
Equipments and reagents
 Test tubes
 Centrifuge
 Microscope
 Microscopic slide
 Normal saline
 20% albumin
 AHG (Coombs reagents)
Procedure
1. Take 3 small tubes mark them 1,2 and 3, and add to

each the following
Tube 1 1volume of patient’s serum

1volume of 3-5% donor’s red cells
Tube 2 1 volume of 20% bovine albumin
1volume of patient’s serum
Tube 3 3 volume of patent’s serum
1volume of 20-30% suspension of
donor’s cells
2. Incubate tube 1at RT for 30 min and Tube 2 for 30

minutes at 370C. Incubate tube 3 for 15 minutes at
370C.
3. After incubation, remove tube 3 and wash the cells

three times with clean saline to make sure that
all the globulins are removed from the cells.
4. And make a 3% saline suspension of the washed

cells in a tube.
5. To one volume of red cell deposit add 2 volumes

of fresh diluted antiglobulin (coombs) Reagent.
6. Remove tube 1 and 2 and centrifuge with tube 3

for one minute at 1000 rpm
7. Examine the tube for heamolysis

macroscopically and microscopically for
agglutination.
Results
 No hemolysis or agglutination is seen in tube 1,
2 or 3
 the blood is compatible and can be issued with the
completed cross-match label.
 If there is agglutination or hemolysis in any of
the tubes
 the blood is incompatible, and must not be issued for
the patient.
Incompatibility investigation
ABO incompatibility (anti A and Anti-B.)

 Saline tube ………………….Shows strong agglutination
 Albumin tube ………………. Shows agglutination
 Anti- globulin tube ………… show no agglutination
Rhesus incompatibility (anti-D ,c, e)

 Saline tube …………………….does not usually show
agglutination
 Albumin tube …………………. Shows agglutination
 Anti- globulin tube ……………. Shows agglutination
Cross match (Incomp. Invest.)…
Anti- Duffy and anti- kidd incompatibilities

 Saline tube …… Does not usually show agglutination
 Albumin tube … does not usually show agglutination
Anti- Lewis incompatibility

 Saline tube …………………….. Shows agglutination
 Albumin tube ………………….. Shows agglutination
7.5.2 Emergency cross match
 Performed when there is no enough time to

perform the standard cross match
 Takes about 25 to 30 minutes and
 Does not include antiglobulin test.

Cross match (Emergency)…
Principle
 Serum of the recipient / donor is tested against

the red cells of the donor/ recipient in saline and
albumin medium in order to establish their
compatibility
Type of specimen
 Serum(plasma) not older than 48 hrs

 Washed cells (2-5%)
Equipments and reagents
 Test tubes
 Centrifuge
 Microscope
 Microscopic slide
 Normal saline
 20% albumin
Procedure:
1. Take 2 small tubes, mark them 1 and 2 and

add to each the following

1volume of 3-5% donor’s red cells.
1volume of 20% bovine albumin
2. Leave tube 1 at room temp for 15 minutes

incubate tube 2 for 15 minutes at 37 0C.
3. Centrifuge both tubes for one minute at 1000

rpm/min
4. Examine the tubes macroscopically for

hemolysis and microscopically for agglutination.
Results
 If no hemolysis or agglutination is seen in either tube 1
or 2
 the
blood is compatible and can be issued with the
emergency cross match.
 If agglutination or hemolysis is seen in either of the

tubes
 the blood is incompatible and must not be issued for the
patient.
7.5.3. Rapid direct slide cross match
(Request for un cross matched blood )
 Takes only 3 or 4 minutes
 Plasma is used instead or serum.
 Not safe and must only used in extreme emergencies
 Standard cross match should be carried out while the

transfusion is in progress.
Procedure
1. Take 2 volume of patient’s plasma on a slide
2. Add 1 volume of donor’s whole blood of the

same group as the patient and mix.
3. Leave for 2 minutes and examine

microscopically for agglutination
Cross match (Rapid)…
Results
 If the cells show agglutination the blood must not

be given and will usually indicate that the wrong
ABO group blood is being cross marched.
Cross match (Rapid)…
Sources of errors in cross-matching

 Rouleaux
 Auto agglutinins
 Infected donor cells
 Anti- A1
 Over centrifugation
 Dirty glass wares etc..
Review Questions
1. What is cross-matching?
2. What is the purpose of cross-matching?
3. List the types of cross-match with their constituents
4. List the stages of cross-match and their respective

importance in antibody detection.

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CHPTER -SEVEN

 Types of Cross Match

 Steps for compatibility testing

 Procedure for cross-match

At the end of this chapter, the student will be able

 It is a test to determine the compatibility between

 Main purpose is to prevent a transfusion

 Some ABO incompatibilities and

 Technical or labeling errors (clerical errors)

 Transfusion reaction – is the term used to

 However, it will not:

 guarantee normal survival of transfused erythrocytes

 detect all unexpected antibodies in a patient’s serum.

Major and Minor crossmatch

 is much more critical for assuring safe transfusion

 called major b/c the Abs in the recipient’s serum are

 Called minor because

 thedestructed RBCs of the patient may be

 Proper sample collection and handling

 Review of the recipient’s past blood bank records

 Careful ABO/Rh determination

 Antibody screening of the recipient (cross matching of the

 In cases when the recipient possess a clinically

 Screened for the corresponding Ag and should be

 Finally, during the actual transfusion :

 careful observation of the recipient’s vital signs and

 posttransfusion hematocrit and Heamoglobin levels

 The blood selected for cross-match should be of

 However, Rh positive recipients may receive

 Whenever possible blood of the patients own

 Otherwise the following rules should be applied.

 When cross-matching is carried out, the serum

 The serum should be fresh, that is not more than

 When deciding on methods for cross-matching,

 The right Temperature.

 The safe cross-matching of blood requires that

 The red cells from the donor are suspended in

 The red cells from the donor’s suspended in

 Agglutination in the albumin tube is often caused

 the rhesus antibodies,

 Reaction caused by anti- A or anti- B antibodies

 A more concentrated suspension of red cells is

 A postive test detects the presence of antibodies

 Anti globulin is essential for detection anti-Duffy

 Serum of the recipient / donor is tested against

 Serum (plasma) not older than 48 hrs

Equipments and reagents

1. Take 3 small tubes mark them 1,2 and 3, and add to

Tube 1 1volume of patient’s serum

2. Incubate tube 1at RT for 30 min and Tube 2 for 30

3. After incubation, remove tube 3 and wash the cells

4. And make a 3% saline suspension of the washed

5. To one volume of red cell deposit add 2 volumes

6. Remove tube 1 and 2 and centrifuge with tube 3

7. Examine the tube for heamolysis

ABO incompatibility (anti A and Anti-B.)

Rhesus incompatibility (anti-D ,c, e)

Anti- Duffy and anti- kidd incompatibilities

Anti- Lewis incompatibility