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Completing the CIBMTR Completing the CIBMTR Confirmation of HLA Typing Form Confirmation of HLA Typing Form
(Form 2005)(Form 2005)
Stephen SpellmanResearch ManagerNMDP Scientific Services
Maria BrownScientific Services Specialist
Data Management Conference 2007
2
Introduction to Form 2005
Basic Biology of HLA
Overview of HLA nomenclature
Laboratory report interpretation and data transcription
3
IntroductionIntroduction
The CIBMTR product information is collected on three separate forms:
• The IDM form (Form 2004)
• The Confirmation of HLA Typing form (Form 2005)
• Infusion form (Form 2006)
HLA form modeled on NMDP Form 22/117
4
Confirmation of HLA Typing FormConfirmation of HLA Typing Form
The Confirmation of HLA Typing form is completed for:
• Non-NMDP allogeneic or syngeneic donors
• Non-NMDP recipients
• Non-NMDP cord blood units
Complete a separate copy of this form for each donor, recipient, or cord blood unit
Not required on NMDP-facilitated transplants!
5
FormsNetFormsNet 2.02.0
All data fields identical to the paper form
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Basic Biology of HLABasic Biology of HLA
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What is HLA?What is HLA?
Human Leukocyte Antigen (HLA) First discovered through antigenic differences on white blood cells
Highest expressed antigens are the class I and class II HLA molecules (0.5 - 1 million per cell)
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HLA and the MHCHLA and the MHC
HLA is part of a larger system know as the Major Histocompatibility Complex (MHC)
MHC is a set of closely linked genes inherited as a unit (complex)
MHC was initially recognized by its major influence on transplant rejection
9
Histocompatibility MoleculesHistocompatibility Molecules
MHC is divided into two main regions:
•HLA class I
•HLA class II
Encoded by genes on chromosome 6
10
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HLA InheritanceHLA Inheritance
HLA genes on the same haplotype are inherited together, along with the rest of the MHC ‘Complex’
Maternal Copy
Paternal Copy
HLA Class II HLA Class IC
C
CC
C
C DRB1 DQB1 B C A
DRB1 DQB1 B C A
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HLA InheritanceHLA Inheritance
d
O F
F S
P R
I N
G
c
db
b
b/a
25%
25%
Recombinant Sibling (1%)
c
a25%
25%a
dA B DR
a
b
c
d
FATHER
MOTHER
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HLA DiversityHLA Diversity
Highest degree of polymorphism in human genome
The diversity makes possible thousands of different combinations
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HLA Typing MethodsHLA Typing Methods
SerologyIdentifies HLA molecules on the cell surface using antigen specific anti-sera
DNAIdentifies HLA molecules by defining the DNA code in the cell nucleus
AntigenProteinDNA
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HLA NomenclatureHLA Nomenclature
DNA DNA SerologySerology
DNA SerologyA*0201 A2A*0202 A2A*0203 A2A*0204 A2A*0205 A2…..A*0273 A2
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Number of Antigens/Alleles
17
Why So Diverse?Why So Diverse?
Present many different types of peptides
Protection from a myriad of pathogens
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The Function of HLA MoleculesThe Function of HLA Molecules
Present antigen (peptides) to T-lymphocytes
Facilitate immune surveillance
Detection of:
• Viral or bacterial infected cells
• Mutated genes or proteins
19
Class I HLA MoleculesClass I HLA Molecules
Co-dominantly expressed on the surface of almost all nucleated cells (except RBCsand CNS)
Present intracellular peptides to CD8+ T cells
Individuals express up to 6 different class I molecules; two HLA-A, two HLA-B and two HLA-C
21
Class II HLA MoleculesClass II HLA Molecules
Expressed on the surface of Antigen Presenting Cells (APCs)• B lymphocytes, dendritic cells and thymic
epithelial cells• Class II HLA-DRB1/3/4/5, DQA1, DQB1, DPA1,
DPB1
Present extracellular (and intracellular) peptides to CD4+ T cellsConsist of a and b chains
23
HLA Class I and IIHLA Class I and II
Antigen Presenting Cell
Groove-Domain CytoplasmicTM
Class I
Class II
1 5 6 87Exon 4Exon 3Exon 2
1 4 65Exon 3Exon 2
24
HLA Class I and IIHLA Class I and II
Antigen Presenting Cell
Peptide
Groove-Domain CytoplasmicTM
Class I
Class II
1 5 6 87Exon 4Exon 3Exon 2
1 4 65Exon 3Exon 2
25
Groove-Domain CytoplasmicTM
Class I
Class II
1 5 6 87Exon 4Exon 3Exon 2
1 4 65Exon 3Exon 2
HLA Class I and IIHLA Class I and II
Antigen Presenting Cell
T Cell
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HLA and TransplantationHLA and Transplantation
Stem cell transplantation is transplanting of the immune system
Graft rejection occurs when the residual host immune system attacks the transplanted immune system
Graft vs. Host Disease (GvHD) occurs when the new immune system begins to attack the body of the recipient
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HLA Basic Biology HLA Basic Biology -- SummarySummary
HLA genes part of MHC complex
Inherited as maternal and paternal haplotypes
HLA genes are the most polymorphic in the human genome
Facilitate immune surveillancePresent self and non-self peptides (viral, bacterial, mutated proteins, etc…) to circulating T cells
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Overview of HLA NomenclatureOverview of HLA Nomenclature
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Examples of Serological TypingExamples of Serological TypingNomenclatureNomenclature
order ofantigen
discovery
gene
A10 B15 DR4
A2 B35 DR15
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Examples of DNA Typing Examples of DNA Typing NomenclatureNomenclature
gene
order ofallele discovery
A*2501 B*1503 DRB1*0402
A*0210 B*3502 DRB1*1504
“ * ” DNAmethodology allele group
•structure•serologic
31
DNA Nomenclature:DNA Nomenclature:Silent SubstitutionsSilent Substitutions
DRB1*150201DRB1*150202DRB1*150203DRB1*150204
The fifth & sixth digits indicate a difference in DNA structure, but the HLA antigens specified do not differ
Example: DRB1*1502
32
DNA Nomenclature OdditiesDNA Nomenclature Oddities
A*020101 = Silent mutationA*2409N = (Null) No antigen expressedA*2402L = Low antigen expressionB*4402S = HLA protein is secretedB*3565Q = Antigen level is questionable
Antigen
33
NMDP Allele CodesNMDP Allele Codes
11AAF
01AG13ADG
07XX01PP
01AD
15AAA02GS
04FH
01TY
01FG
13ST
01DD
11AKA
02EF 04FH
13EG
01TY01DD01BAB
03HJ
34
NMDP Allele Codes used for Multiple NMDP Allele Codes used for Multiple Allele DesignationsAllele Designations
Used to report multiple possible alleles (intermediate/high resolution)
Provides maximum information on the person’s typing
NMDP Bioinformatics Web site contains look-up Web tool
www.bioinformatics.nmdp.org
Used to report multiple possible alleles (intermediate/high resolution)
Provides maximum information on the person’s typing
NMDP Bioinformatics Web site contains look-up Web tool
www.bioinformatics.nmdp.org
35
NMDP Codes for Multiple Allele NMDP Codes for Multiple Allele DesignationDesignation
AB = 01/02
A*0201 or *0202 = A*02AB
B*4001 or *4002 = B*40AB
DRB1*1501 or *1502 = DRB1*15AB
A*0201 or *0202 = A*02AB
B*4001 or *4002 = B*40AB
DRB1*1501 or *1502 = DRB1*15AB
36
CIBMTR Confirmation of HLA CIBMTR Confirmation of HLA Typing Form (Form 2005)Typing Form (Form 2005)
37
Key Fields BoxKey Fields Box
•The Key Fields Box contains information about the transplant –recipient, donor, cord, transplant date and type of transplant.
•The Key Fields link the donor forms to the patient forms.
38
Key Fields BoxKey Fields Box
The section on the bottom of Page 5 is used if multiple donors or cords were infused into one recipient.
39
Whose HLA typing data are you Whose HLA typing data are you reporting? reporting?
40
Are you submitting a laboratory Are you submitting a laboratory report?report?
If you have any questions about how to interpret a lab report, please contact either:
Maria Brown 612/627-8186 [email protected] Spellman 612/617-8334 [email protected]
41
Transcribing HLA Typing Data Transcribing HLA Typing Data from Laboratory Reportsfrom Laboratory Reports
42
Is it Serology or DNA?Is it Serology or DNA?
Serology
LymphotoxicityMicrolymphotoxicitySerology
DNALuminexMolecularPCRPCR-SSPSBTSSOSSOPDNA
43
General Instructions General Instructions –– DNA TypingDNA Typing
One possible allele in each set of boxes
Use as many boxes as you have alleles to report
If more than six possible alleles, fill the boxes provided then write the remainder of the alleles in the available space on the page, clearly indicating the appropriate locus
44
How to Read a Lab ReportHow to Read a Lab Report
“/” is used to separate alleles (example DRB1*0701/03 equals 0701 or 0703)
“-” is used to indicate a range of alleles (example B*0801-0803 equals 0801/0802/0803)
45
DNADNA--Based Typing in Based Typing in FormsNetFormsNet
0201/09/43N/66/75/83N/89●
●
●
●
0301/01N/20/21N
●
0702/44
5701
0702
0602
04070701
The HLA field in FormsNetsupports up to 250 characters
HLA Laboratory Report
46
DNADNA--Based TypingBased Typing
HLA Laboratory Report
Notice that the 02 is in front of the A*0289.
0 2 0 16 6
4 3 N0 9A*0 2 8 9
7 5 8 3 N
0 7 0 2
5 7 0 1
0 6 0 2
4 4
0 7 0 2
0 4 0 7
0 7 0 1
0 1
2 1 N2 00 3 0 1 0 1 N
Which Typing do I Use?Which Typing do I Use?
Remember to check the comments or footnotes on your laboratory report.
There is often further clarification of the HLA typing.
0 2 7 70 2 7 10 2 0 1
0 2 7 4 0 1 0 2 7 5
4 4 0 2 4 4 4 1
0 2 6 8
A*0281-85/0287-90/0293-96
1 8 0 1
0 5 0 1 0 5 1 3
0 7 0 1
0 2 7 10 2 7 7
Patient
Donor
48
*0702*07@*1101*0301*0801*0702*3201*0201
Cw#2Cw#1DR#2DR#1B#2B#1A#2A#1
NT*0301*0201
DRB345#2DRB345#1DQ#2DQ#1
Laboratory Report
Enter the alpha/numeric data from the footnote on the forms and/or into FormsNet 2.0. Please note that without the key we cannot decipher your symbols.
@ = C*0701/0706/0718NT = Not Tested
What About Symbols @#$&%!What About Symbols @#$&%!
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Reporting NMDP Allele CodesReporting NMDP Allele Codes
02ANGA●
●
●
03XKS
1402
44AMUT
05AC0802
FormsNet
0 2 A N G A
0 3 X K S
1 4 0 2
4 4 A M U T
0 5 A C
0 8 0 2
Paper form
Directly transcribe any allele codes onto the form or into FormsNet.
50
What to do with Positives??What to do with Positives??
Example
Either write or type “positive”or “pos”
Please do not use “+” or “-”
P o s
0 5 R V
0 6 B V AH
51
Serologic Typing and DNA?Serologic Typing and DNA?ENTER BOTH!ENTER BOTH!
CUMULATIVE HLA TYPE RESULTS
MOLECULAR TYPING BY PCR-SSP, SBT AND/OR LUMINEX SSO
HLA Class I Antigens HLA Class II Antigens A B Cw DR DR51/52/53 DQ
A1 A24 B8 B35 Cw4 Cw7 DR1 DR17 DQ2 DQ5
HLA -A HLA -B HLA -Cw HLA-DPB1 A*0101 B*0801 Cw*0401 A*2402 B*3501 Cw*0701
HLA – DRB1 HLA – DRB3/4/5 HLA –DQB1
DRB1*0101 DQB1*0201 DRB1*0301 DQB1*0501
0 1 0 1
2 4 0 2
0 8 0 1
3 5 0 1
Serology
DNA
Serology in Serology in FormsNetFormsNet 2.02.0
●Drop down selection menu
Yes/No buttons
53
Example Laboratory Reports for Example Laboratory Reports for Completing Form: Completing Form:
Confirmation of HLA Typing Form Confirmation of HLA Typing Form (Form 2005)(Form 2005)
54
Example AExample A
55
Example A Example A continuedcontinued
2 9 X X
2 4 X X
0 7 X X
4 4 0 3
0 7 X X
1 6 0 1
1 5 0 1
1 3 0 1
Positive
56
Example BExample B
57
Example B continuedExample B continued
1 5
0 1
0 2
0 7
4 4
0 5
0 7 0 2 1 01 7
0 7 0 1 1
1 5 0 1 1
0 1 0 1
1 5
58
Example CExample C
59
Example C continuedExample C continued
3
1 1
2 7
1
1
4
60
Example DExample D
61
Example D continuedExample D continued
0 8 0 1― 0 3
1 3 0 1― 0 3
0 6 0 2
0 7 0 1 0 7 0 6
0 3 0 1
0 7 0 1 0 3
0 3
0 1 0 1 0 3
0 1 0 1 ―0 5
0 2 0 1― 2 2 / 2 5 ― 2 7 / 2 9
3 0 0 1― 0 4 / 0 6
Example EExample E
63
Example E continuedExample E continued
0 1 0 1
0 7 0 1
0 1
64
SummarySummary
Form 2005 required on non-NMDP donors and recipients
Multiple donors = multiple forms
Transcribe data directly as reported on laboratory report*
Include all possible alleles for DNA-based typing
Report both DNA and Serology data, if available
*Remember to watch for footnotes
65
Please remember:
If you have any questions about how to interpret a lab report, please contact either:Maria Brown 612/627-8186 [email protected] Spellman 612/617-8334 [email protected]