identifying antibodies
DESCRIPTION
Identifying Antibodies. The Antibody Panel. CLS 422 Clinical Immunohematology I. Objectives. Discuss clinical situations when it is appropriate to perform antibody identification. Define a panel of cells. Explain how the following factors aid in the interpretation of antibody panels: - PowerPoint PPT PresentationTRANSCRIPT
Identifying AntibodiesIdentifying Antibodies
The Antibody Panel
CLS 422
Clinical Immunohematology I
ObjectivesObjectives Discuss clinical situations when it is appropriate to
perform antibody identification. Define a panel of cells. Explain how the following factors aid in the
interpretation of antibody panels:a. Cross-out techniqueb. Variation in strengths of reactionc. Phases of reactiond. Autocontrole. Red blood cell antigen typing
ObjectivesObjectives
List testing that can be performed to confirm the identification of antibodies.
Identify the antibodies present, when given panel results.
When is an When is an antibody antibody
identification panel identification panel performed?performed?
When the antibody screen is positive.
When to perform testWhen to perform test
The panel red blood cells (RBCs) are tested against the patient’s serum or plasma in order to identify the unexpected antibody or antibodies present.
IdentificationIdentification
Antibody screen – positive Run antibody panel to identify antibody (-ies). If original panel does not provide a clear-cut
ID, test additional RBCs. Selected cells Alternate methods
ConfirmationConfirmation
Rule of 3 and 3 Antigen type patient’s RBCs.
Landsteiner’s Law!!!
The PanelThe Panel
Series of 8 to 20 Group O RBCs Various distribution of the most common RBC
antigens Suspended in a preservative to protect antigen
integrity for 2 -4 weeks Packaged with a lot-specific antigram
AntigramAntigram
Donor
Cell number
D C c E eCw K k
Kpa
Kpb
Jsa
Jsb
Fya
Fyb
Jka
Jkb
Lea
Leb
P
1M N S s
Lua
Lub
Xga
RZR1 1 + + 0 + + 0 0 + 0 + 0 + 0 0 0 + 0 + 0 0 + 0 + 0 + +
R1wR1 2 + + 0 0 + + + + 0 + 0 + 0 + + + 0 + + 0 + 0 + 0 + +
R2R2 3 + 0 + + 0 0 0 + 0 + 0 + + + 0 + + 0 + + + 0 + 0 + 0
r’r 4 0 + + 0 + 0 0 + 0 + 0 + 0 + 0 + + 0 + + + + + 0 + +
r’’r 5 0 0 + + + 0 0 + 0 + 0 + + 0 + 0 0 + 0 + + 0 + 0 + +
rrK 6 0 0 + 0 + 0 + + 0 + 0 + 0 + + 0 0 + + 0 + 0 + 0 + +
rrFya 7 0 0 + 0 + 0 0 + 0 + 0 + + 0 + + 0 0 + 0 + + + 0 + +
Ror 8 + 0 + 0 + 0 0 + 0 + + 0 0 0 + 0 0 + + 0 + 0 0 0 + +
rr 9 0 0 + 0 + 0 + + 0 + 0 + + + + 0 0 + + + 0 + 0 + + +
R2r 10 + 0 + + + 0 + 0 0 + 0 + + + + 0 0 + + + + + + + + +
R1R1 11 + + 0 0 + 0 0 + 0 + 0 + + 0 + + + 0 0 + + 0 + 0 + +
PatientCells
Panel AntigramPanel Antigram
Auto ControlAuto Control
Patient’s serum/plasma tested against a suspension of patient’s RBCs Optional Evaluate results in conjunction with patient history
Autoantibody Newly forming alloantibody
If patient has a positive DAT, auto control will be positive
Usually the same as was used for the antibody screen
Must include incubation at 37oC Must include an AHG phase with reagent
containing anti-IgG
Test MethodTest Method
Interpreting Interpreting Panel ResultsPanel Results
Cell D C c E e Cw K k Kpa Kpb J
sa
Jsb
Fya
Fyb
Jka
Jkb
Lea
Leb
P
1M N S s
Lua Lub X
ga
AHG
CC
1 + + 0 0 + 0 + + 0 + 0 + + + + 0 0 + + + + 0 + 0 + + 3+
2 + + 0 0 + + 0 + 0 + 0 + 0 + 0 + 0 + + + + 0 + 0 + + 0 2+
3 + 0 + + 0 0 0 + 0 + 0 + + 0 + + 0 + + + 0 0 + 0 + + 0 2+
4 + 0 + 0 + 0 0 + 0 + + 0 0 0 + 0 0 + + 0 + 0 0 0 + + 0 2+
5 0 + + 0 + 0 0 + 0 + 0 + 0 + + + 0 + + + + 0 + 0 + + 0 2+
6 0 0 + + + 0 0 + 0 + 0 + + + + + + 0 + 0 + 0 + 0 + + 0 2+
7 0 0 + 0 + 0 + + 0 + 0 + 0 + + 0 0 + + 0 + 0 + 0 + 0 3+
8 0 0 + 0 + 0 0 + 0 + 0 + + 0 + + + 0 0 + 0 0 + + + 0 0 2+
9 0 0 + 0 + 0 0 + 0 + 0 + 0 + 0 + 0 + + + 0 + 0 0 + + 0 2+
10 + + + 0 + 0 + + 0 + 0 + + + 0 + 0 0 + + 0 + + 0 + + 3+
11 + w + + w 0 0 + 0 + 0 + 0 0 + 0 0 + + 0 + + 0 0 + + 0 2+
Auto 0 2+
ExclusionExclusion
Begin with the RBCs that failed to react The antibody in the serum is not directed
against the antigens on these RBCs, so we can eliminated these antibody specificities
Look at alleles to avoid problems with dosage! Exclusion should be done using RBCs having
homozygous antigen expression. Exceptions are low prevalence antigens
Cell D C c E e Cw K k Kpa Kpb J
sa
Jsb
Fya
Fyb
Jka
Jkb
Lea
Leb
P
1M N S s
Lua Lub X
ga
AHG
CC
1 + + 0 0 + 0 + + 0 + 0 + + + + 0 0 + + + + 0 + 0 + + 3+
2 + + 0 0 + + 0 + 0 + 0 + 0 + 0 + 0 + + + + 0 + 0 + + 0 2+
3 + 0 + + 0 0 0 + 0 + 0 + + 0 + + 0 + + + 0 0 + 0 + + 0 2+
4 + 0 + 0 + 0 0 + 0 + + 0 0 0 + 0 0 + + 0 + 0 0 0 + + 0 2+
5 0 + + 0 + 0 0 + 0 + 0 + 0 + + + 0 + + + + 0 + 0 + + 0 2+
6 0 0 + + + 0 0 + 0 + 0 + + + + + + 0 + 0 + 0 + 0 + + 0 2+
7 0 0 + 0 + 0 + + 0 + 0 + 0 + + 0 0 + + 0 + 0 + 0 + 0 3+
8 0 0 + 0 + 0 0 + 0 + 0 + + 0 + + + 0 0 + 0 0 + + + 0 0 2+
9 0 0 + 0 + 0 0 + 0 + 0 + 0 + 0 + 0 + + + 0 + 0 0 + + 0 2+
10 + + + 0 + 0 + + 0 + 0 + + + 0 + 0 0 + + 0 + + 0 + + 3+
11 + w + + w 0 0 + 0 + 0 + 0 0 + 0 0 + + 0 + + 0 0 + + 0 2+
Auto 0 2+
ExclusionExclusion
InclusionInclusion
Of the antibody specificities that have not been excluded, match the pattern of positive and negative reactions with the pattern of antigen positive and antigen negative cells.
There must be an explanation for each positive reaction seen.
Cell D C c E e Cw K k Kpa Kpb J
sa
Jsb
Fya
Fyb
Jka
Jkb
Lea
Leb
P
1M N S s
Lua Lub X
ga
AHG
CC
1 + + 0 0 + 0 + + 0 + 0 + + + + 0 0 + + + + 0 + 0 + + 3+
2 + + 0 0 + + 0 + 0 + 0 + 0 + 0 + 0 + + + + 0 + 0 + + 0 2+
3 + 0 + + 0 0 0 + 0 + 0 + + 0 + + 0 + + + 0 0 + 0 + + 0 2+
4 + 0 + 0 + 0 0 + 0 + + 0 0 0 + 0 0 + + 0 + 0 0 0 + + 0 2+
5 0 + + 0 + 0 0 + 0 + 0 + 0 + + + 0 + + + + 0 + 0 + + 0 2+
6 0 0 + + + 0 0 + 0 + 0 + + + + + + 0 + 0 + 0 + 0 + + 0 2+
7 0 0 + 0 + 0 + + 0 + 0 + 0 + + 0 0 + + 0 + 0 + 0 + 0 3+
8 0 0 + 0 + 0 0 + 0 + 0 + + 0 + + + 0 0 + 0 0 + + + 0 0 2+
9 0 0 + 0 + 0 0 + 0 + 0 + 0 + 0 + 0 + + + 0 + 0 0 + + 0 2+
10 + + + 0 + 0 + + 0 + 0 + + + 0 + 0 0 + + 0 + + 0 + + 3+
11 + w + + w 0 0 + 0 + 0 + 0 0 + 0 0 + + 0 + + 0 0 + + 0 2+
Auto 0 2+
InclusionInclusion
Other Points to ConsiderOther Points to Consider
In what phase(s) of testing is the antibody reactive? May give clue as to antibody identity and clinical
significance. Is the strength of reaction the same for each cell that
reacts, or is there variation in strength? Dosage, antigen variability, or multiple antibodies.
Is the antibody reacting only with “homozygous” cells of a certain specificity? Weak antibody showing dosage.
Did the autologous control react? Autoantibody or newly forming alloantibody.
ProbabilityProbability
Rule of 3 and 3 For each antibody specificity, are there 3
antigen positive cells that reacted and 3 antigen negative cells that did not react? May use screen cells in addition to panel cells to
fill this rule Cells do not need to have homozygous antigen
expression to fill this rule
Cell D C c E e Cw K k Kpa Kpb J
sa
Jsb
Fya
Fyb
Jka
Jkb
Lea
Leb
P
1M N S s
Lua Lub X
ga
AHG
CC
1 + + 0 0 + 0 + + 0 + 0 + + + + 0 0 + + + + 0 + 0 + + 3+
2 + + 0 0 + + 0 + 0 + 0 + 0 + 0 + 0 + + + + 0 + 0 + + 0 2+
3 + 0 + + 0 0 0 + 0 + 0 + + 0 + + 0 + + + 0 0 + 0 + + 0 2+
4 + 0 + 0 + 0 0 + 0 + + 0 0 0 + 0 0 + + 0 + 0 0 0 + + 0 2+
5 0 + + 0 + 0 0 + 0 + 0 + 0 + + + 0 + + + + 0 + 0 + + 0 2+
6 0 0 + + + 0 0 + 0 + 0 + + + + + + 0 + 0 + 0 + 0 + + 0 2+
7 0 0 + 0 + 0 + + 0 + 0 + 0 + + 0 0 + + 0 + 0 + 0 + 0 3+
8 0 0 + 0 + 0 0 + 0 + 0 + + 0 + + + 0 0 + 0 0 + + + 0 0 2+
9 0 0 + 0 + 0 0 + 0 + 0 + 0 + 0 + 0 + + + 0 + 0 0 + + 0 2+
10 + + + 0 + 0 + + 0 + 0 + + + 0 + 0 0 + + 0 + + 0 + + 3+
11 + w + + w 0 0 + 0 + 0 + 0 0 + 0 0 + + 0 + + 0 0 + + 0 2+
Auto 0 2+
The Rule of 3 and 3The Rule of 3 and 3
Antigen TypingAntigen Typing
Confirms the antibody identification LANDSTEINER’S LAW
Test patient’s RBCs (unknown antigen) against appropriate anti-sera (known antibody)
Results should be negative
Run positive and negative controls for anti-sera A positive DAT or recent transfusion may invalidate
the typing results
Cell D C c E e Cw K k Kpa Kpb J
sa
Jsb
Fya
Fyb
Jka
Jkb
Lea
Leb
P
1M N S s
Lua Lub X
ga
AHG
CC
1 + + 0 0 + 0 + + 0 + 0 + + + + 0 0 + + + + 0 + 0 + + 3+
2 + + 0 0 + + 0 + 0 + 0 + 0 + 0 + 0 + + + + 0 + 0 + + 0 2+
3 + 0 + + 0 0 0 + 0 + 0 + + 0 + + 0 + + + 0 0 + 0 + + 0 2+
4 + 0 + 0 + 0 0 + 0 + + 0 0 0 + 0 0 + + 0 + 0 0 0 + + 0 2+
5 0 + + 0 + 0 0 + 0 + 0 + 0 + + + 0 + + + + 0 + 0 + + 0 2+
6 0 0 + + + 0 0 + 0 + 0 + + + + + + 0 + 0 + 0 + 0 + + 0 2+
7 0 0 + 0 + 0 + + 0 + 0 + 0 + + 0 0 + + 0 + 0 + 0 + 0 3+
8 0 0 + 0 + 0 0 + 0 + 0 + + 0 + + + 0 0 + 0 0 + + + 0 0 2+
9 0 0 + 0 + 0 0 + 0 + 0 + 0 + 0 + 0 + + + 0 + 0 0 + + 0 2+
10 + + + 0 + 0 + + 0 + 0 + + + 0 + 0 0 + + 0 + + 0 + + 3+
11 + w + + w 0 0 + 0 + 0 + 0 0 + 0 0 + + 0 + + 0 0 + + 0 2+
Auto 0 2+
Selecting Controls for Antigen TypingSelecting Controls for Antigen Typing
Value of Patient HistoryValue of Patient History
The following additional information may assist in determining the identity of the antibody: History of antibodies Transfusion, transplant, pregnancy (how many and
how long ago) Medications Diagnosis Ethnicity
ReportingReporting
Panel results are reported as “anti-” and then the specificity Anti-K
If specificity cannot be determined at this point, additional testing must be performed
The End