is it time to re-evaluate the gold standards in blood banking ?

Jerry A. Holmberg, MT(ASCP)SBB, Ph.D.

June 6, 2013

201 Florida Association of Blood Banks

Is it time to re-evaluate the gold standardsin blood banking?

| Re-evaluating the Gold Standards| Holmberg | 24April2013 | CBBS 20132

Agenda

Current status of transfusions in the U.S.

Traditional serological typing methods

Red cell genotyping as a supplemental method

Regulatory considerations

Recent Blood Product Advisory Committee discussion


Disclaimer

All blood group genotyping tests commercially available in the US and Canada are for research use only. Not for use in diagnostic procedures.

Transfusions/1,000 Population Comparison in Selected Countries

US England Australia Denmark Sweden

Units of RBC per 1000 Population

48.7 (2001)48.9 (2006)48.8 (2008)

44.9 28 54.0858.6 (2000-2002)

45.3 (1996-20020

Recipient Age

<41 18.8%41-65 27.8%>65 53.3%

<40 14.4%40-70 38.4%> 70 47.2%

<40 15.4%40-70 36.7%> 70 47.9%

<39 9.4%40-59 18.2%> 60 72.4%

<39 9.8%40-59 15.1%> 60 75.2%

Gender M 48.5%F 51.5%

M 50.4%F 49.6

M 52.5%F 47.5%

M 53.2%F 46.8%

M 52.9%F 47.1%

2009 National Blood Collection and Utilization Survey

Kamper-Jorgensen Transfusion 2009; 49:888-894

Cobain Transfusion Medicine 2007; 17, 10-15


5

2009 HHS Blood Collection and Utilization Survey of RBC Use by Hospital Service

General Surgery

11%

Orthopedic Surgery

6%

Cardiac Surgery

7%

Trauma/ER9%

Hem/Onc15%

Transplant1%OB/GYN

2%Ped/Neonates

2%

Neph/Dialysis2%

ICU11%

General Medicine

28%

Other6%


2009 Health and Human Services’ Blood Collection and Utilization Survey

6

Risk Management

Risk Communication

Transfusion Risk

Risk Assessment


Challenges in Transfusion Medicine

Source: FDA 2013, http://www.fda.gov/BiologicsBloodVaccines/SafetyAvailability/ReportaProblem/TransfusionDonationFatalities

Notes: TRALI is transfusion related acute lung injury, which occurs within 6 hours of a transfusion. HTR is hemolytic transfusion reaction, which results in destruction of red cells and is usually caused by incompatibility. TACO is transfusion associated circulatory overload and is characterized by a sharp increase in blood pressure.

31 22

7

http://www.fda.gov/BiologicsBloodVaccines/SafetyAvailability/ReportaProblem/TransfusionDonationFatalities


Challenges in Transfusion Science

RBC Alloimmunization: lessons from sickle cell disease• Alloimmunization in general public is 0.5-1.5%

• Alloimmunization in individuals receiving 3 or more units is 8.4%

• A review of 12 publications found a mean rate of 25% in SCD

Miller ST et al. TRANSFUSION 2013;53:704-709.

RBC Alloimmunization in SCD prevalence in 2010• Editorial (TRANSFUSION 2013;53:692-695.)

• Drs Treml and King advocates for genotype matching and leads to the conclusion that alloimmunization rates remain high partially to transfusions at institutions not providing extended matching


Challenges in Transfusion Science

RBC Alloimmunization in transfused patients with myelodysplastic syndrome or chronic myelomonocytic leukemia• Alloimmunization occurs in 15% of MDS and CMML

• Alloimmunization mostly involves Rh system and Kell

• Antigen matching should include Kell and CcEe

Sanz C et al. TRANSFUSION 2013;53:710-715.

Immunohematologic and patient safety benefit of a centralized transfusion database• Puget Sound advocates for universal transfusion records since

many patients bounce from hospital to hospital

Delaney M TRANSFUSION 2013;53:771-776.


The Future

Are we moving towards more personalized transfusion medicine?• Impact on collections?

• Impact on testing?

• Impact on data management?

• Impact on outcomes?


Agenda






12

Contents

•Preface •About the Author •Acknowledgments •1. Blood and the cells it contains

•Blood contains cells, proteins, and sugars

•Red blood cells transport oxygen

•White blood cells are part of the immune response

•Platelets help blood to clot

•Your complete blood count

•Hemoglobin binds oxygen

•Resources

•2. Blood group antigens are surface markers on the red blood cell membrane

•Antigens stimulate an immune response •Red blood cell antigens can be sugars or proteins •Red blood cell antigens determine your blood group •Blood groups differ around the world •The classification of blood cell antigens •References

•3. Blood transfusions and the immune system

•How to launch an immune response against transfused red blood cells

•"Blood type and cross match"

•Transfusion reactions: Immune-mediated

•Transfusion reactions: Non-immune

•4. Hemolytic disease of the newborn

•Maternal antibodies cross the placenta and attack fetal red blood cells

•Sensitization occurs during the first pregnancy

•HDN occurs in subsequent pregnancies

•The Coombs test detects Rh incompatibility between mother and fetus

•Preventing HDN

•References

•5. The ABO blood group

•At a glance

•Background information

•Basic biochemistry

•Clinical significance of ABO antibodies

•Molecular information

•References

•6. The Hh blood group

•At a glance



•Clinical significance of H antibodies


•References

•7. The Rh blood group

•At a glance



•Clinical significance of Rh antibodies


•References

•8. The Kell blood group

•At a glance



•Clinical significance of Kell antibodies


•References

•9. The Duffy blood group

•At a glance



•Clinical significance of Duffy antibodies


•References

•10. The Kidd blood group

•At a glance



•Clinical significance of Kidd antibodies


•References

•11. The Diego blood group

•At a glance



•Clinical significance of Diego antibodies


•References

•12. The MNS blood group

•At a glance



•Clinical significance of MNS antibodies


•References

< PrevNext >Print View

Bookshelf ID: NBK2264Chapter 2Blood group antigens are surface markers on the red blood cell membrane

Before the 1900s, it was thought that all blood was the same, a misunderstanding that led to frequently fatal transfusions of animal blood into humans and hazardous transfusions of blood between people. Human blood is not the same—people belong to different blood groups, depending upon the surface markers found on the red blood cell.The cells that make up the body's tissues and organs are covered with surface markers, or antigens. Red blood cells are no different. This chapter will describe the types of red blood cell antigen and explain why they are so important in medicine today.Antigens stimulate an immune response

Go to:An antigen is any substance to which the immune system can respond. For example, components of the bacterial cell wall can trigger severe and immediate attacks by neutrophils. If the immune system encounters an antigen that is not found on the body's own cells, it will launch an attack against that antigen. Conversely, antigens that are found on the body's own cells are known as "self-antigens", and the immune system does not normally attack these.The membrane of each red blood cell contains millions of antigens that are ignored by the immune system. However, when patients receive blood transfusions, their immune systems will attack any donor red blood cells that contain antigens that differ from their self-antigens. Therefore, ensuring that the antigens of transfused red blood cells match those of the patient's red blood cells is essential for a safe blood transfusion.Red blood cell antigens can be sugars or proteins

Go to:Blood group antigens are either sugars or proteins, and they are attached to various components in the red blood cell membrane. For example, the antigens of the ABO blood group are sugars. They are produced by a series of reactions in which enzymes catalyze the transfer of sugar units. A person's DNA determines the type of enzymes they have, and, therefore, the type of sugar antigens that end up on their red blood cells.In contrast, the antigens of the Rh blood group are proteins. A person's DNA holds the information for producing the protein antigens. The RhD gene encodes the D antigen, which is a large protein on the red blood cell membrane. Some people have a version of the gene that does not produce D antigen, and therefore the RhD protein is absent from their red blood cells.The figure below shows the red blood cell membrane and some of the blood group antigens attached to it. Aside from the sugar (glycan or carbohydrate) antigens, the red blood cell membrane contains three types of protein that carry blood group antigens: single-pass proteins, multi-pass proteins, and glycosylphosphatidylinositol (GPI)-linked proteins. Click on the blood groups to find out more about the antigens that define it.

Red blood cell antigens determine your blood group

Scientific Overview Blood Group Antigens

Dean L. Blood Groups and Red Cell Antigens [Internet]. Bethesda (MD): National Center for Biotechnology Information (US); 2005. Chapter 2, Blood group antigens are surface markers on the red blood cell membrane. Available from: http://www.ncbi.nlm.nih.gov/books/NBK2264/ | Re-evaluating the Gold Standards| Holmberg | 24April2013 | CBBS 2013

http://www.ncbi.nlm.nih.gov/books/n/rbcantigen/A2/

http://www.ncbi.nlm.nih.gov/books/n/rbcantigen/rbcantigen_book_info/

http://www.ncbi.nlm.nih.gov/books/n/rbcantigen/A201/

http://www.ncbi.nlm.nih.gov/books/n/rbcantigen/ch1/






































































































































































http://www.ncbi.nlm.nih.gov/books/n/rbcantigen/ch05ABO/


































http://www.ncbi.nlm.nih.gov/books/n/rbcantigen/ch06Hh/

































http://www.ncbi.nlm.nih.gov/books/n/rbcantigen/ch07Rh/

































http://www.ncbi.nlm.nih.gov/books/n/rbcantigen/ch08Kell/

































http://www.ncbi.nlm.nih.gov/books/n/rbcantigen/ch09Duffy/


































http://www.ncbi.nlm.nih.gov/books/n/rbcantigen/ch10Kidd/

































http://www.ncbi.nlm.nih.gov/books/n/rbcantigen/ch11Diego/


































http://www.ncbi.nlm.nih.gov/books/n/rbcantigen/ch12MNS/






































http://www.ncbi.nlm.nih.gov/books/NBK2264/?report=printable

http://www.ncbi.nlm.nih.gov/books/NBK2264/?report=printable

http://www.ncbi.nlm.nih.gov/bookshelf/br.fcgi?book=rbcantigen&part=ch12MNS

http://www.ncbi.nlm.nih.gov/bookshelf/br.fcgi?book=rbcantigen&part=ch11Diego

http://www.ncbi.nlm.nih.gov/bookshelf/br.fcgi?book=rbcantigen&part=ch10Kidd

http://www.ncbi.nlm.nih.gov/bookshelf/br.fcgi?book=rbcantigen&part=ch09Duffy

http://www.ncbi.nlm.nih.gov/bookshelf/br.fcgi?book=rbcantigen&part=ch08Kell

http://www.ncbi.nlm.nih.gov/bookshelf/br.fcgi?book=rbcantigen&part=ch07Rh

http://www.ncbi.nlm.nih.gov/bookshelf/br.fcgi?book=rbcantigen&part=ch06Hh

http://www.ncbi.nlm.nih.gov/bookshelf/br.fcgi?book=rbcantigen&part=ch05ABO


Antigen Function

CARBOHYDRATESABOP1PKLEGLOB

HIFORS

ADHESION MOLECULES

LWXGFYLUOK

INSCMER2JMH

TRANSPORTERS AND CHANNELS

RHRHAGJKDILAN

COGILXRJR

COMPLEMENT REGULATION

CH/RGCROMKN

HIFORS

ENZYMES

KELYT

STRUCTURAL OR UNKNOWN

MNSGEDO


Traditional Serological Testing Methods

Limitations of serology to detect antigens in some cases• Recent transfusion (mixed field)

• Warm auto-antibody

• Antisera - Limited availability of some rare antisera

- Quality

Weak expression of some RBC antigens Impractical to screen large numbers of donors for all

potential antigens• Typing for rare antigens is often not performed

• Potential allo-immunization or reactions

Can be time consuming and costly


European PerspectiveBritish Committee for Standards in Haematology

4.8.4. Where there is a discrepancy in reaction strength between different anti-D reagents, or where the reagent fails to give a clear-cut strong positive reaction,1 a decision to investigate further needs to be made based on whether the development of anti-D is likely to cause clinical problems.

7.9. Females of child-bearing potential- 7.9.1. Females of child-bearing potential should receive K negative red

cells unless they are unavailable in an emergency (Lee & de Silva, 2004; BCSH, 2006a).

7.16. Foetal transfusions- 7.16.2. D negative, K negative (and further antigen negative where

appropriate) units should be crossmatched against the maternal plasma by IAT if the maternal plasma contains red cell antibodies of likely clinical significance.

1. Guidelines for the Blood Transfusion Services in the UK 20122. Guidelines for pre-transfusion compatibility procedures in blood transfusion laboratories, British Committee for Standards in Haematology Transfusion Medicine doi: 10.1111/j.1365-3148.2012.01199.x



4.1.4. Interpretation of D grouping has become more complex, with the increase in variety of monoclonal reagents, and molecular testing. The historical distinction between weak and partial D, based on whether the individual is able to make anti-D, has become blurred and a new algorithm is included in Fig. 1.

Guidelines for pre-transfusion compatibility procedures in blood transfusion laboratories, British Committee for Standards in Haematology Transfusion Medicine doi: 10.1111/j.1365-3148.2012.01199.xGuidelines for the Blood Transfusion Services in the UK 2012)



Guidelines for pre-transfusion compatibility procedures in blood transfusion laboratories, British Committee for Standards in Haematology Transfusion Medicine doi: 10.1111/j.1365-3148.2012.01199.xGuidelines for the Blood Transfusion Services in the UK 2012)


Agenda







Bloodgen Consortium

Lund, Sweden

Ulm, GermanyBristol, UK(UWEand BITS)

Rotterdam,Netherlands

Amsterdam,Netherlands

Prague, Czech RepublicDerio, Spain

Courtesy of Dr. Nuria Nogues, Barcelona


Genotyping for RBC Antigens

Case for genotyping

• Basis for predicting phenotypes from genotyping results

• 324 blood group antigens recognized

• 33 blood group systems / 40 unassigned antigens

• Most blood group polymorphisms are now understood

Potential clinical application

• Determination of blood type in transfused patients

• Patients with “allo” or “auto” antibodies

• warm autoantibodies

• positive direct antiglobulin test

• Resolution of phenotype discrepancies


Potential Clinical Application of RBC Molecular

Detection of rare but significant antigens

Typing for antigens when antisera are not available

Identification of a fetus at risk for HDFN

• Determination of parental zygosity

• Typing of fetal blood

Mass screening of donors for antigen-negative blood

Identification of null and novel alleles



German Consensus on Blood Group Genotyping

In 2000 a German consensus statement was developed on the use of blood group genotyping and its potential application in clinical situations (Legler et al. Infusioinsther Transfusionmed 2000; 27: 215-16)

In fetus from amniotic fluid or trophoblastic cells (chorionic villi)

In multiply transfused patients, if standard serology fails

In case of auto and allo-immunohemolytic anemia, if standard serology fails

For weak D types and other variant RH alleles, if serology is inconclusive


Prior to 2001 the usual source of fetal DNA was a sample

of amniotic fluid or chorionic villi.

Cell-free fetal DNA is detectable in the blood of pregnant

women amount increasing throughout gestation.

23

Fetal Blood Group Typing in Europe

Fetal RhD type can be predicted reliably from the fetal

DNA in the plasma of D neg pregnant women from

beginning of 2nd trimester.



Non-invasive fetal blood group typing from maternal

plasma cell-free fetal DNA is now a clinical reality.

Offered by many laboratories in Europe, to identify the

fetus not at risk of HDFN.

Different assays are currently used for reliable

genotyping of D, C, c, E and K by quantitative real-time

PCR techniques.

Fetal Blood Group Typing in sensitized pregnant womenEuropean Experience



Fetal RHD typing: Application to all D neg pregnant womenEuropean Experience

Trials of high-throughput methods have demonstrated that

accurate fetal D testing in all D- pregnant women is feasible.

Fetal RHD typing to target antenatal anti-D prophylaxis already

introduced in Denmark (2010) and the Netherlands (2011).

TRANSFUSION Volume 52, April 2012


Blood Donors Molecular Typing Barcelona Blood Center Experience

Focused on specific groups of donors:

Extensive genotyping of blood donors from immigrant

populations Aim: Identify blood donors expressing low-incidence antigens

(Dia, Jsa, Mia, Cob, Kpa) or donors with rare phenotypes:

Di(a+b), Co(a-b+), Fy(a-b-), HPA-1(a-b+)

Utility:

- For transfusion (rare phenotype blood units)- RBC panel units (Antibody identification)- Diagnostic (reagent cells)



Utility:

- Quality control of D negative units

- Estimate the real incidence of Del units

RHD Genotyping of D negative blood donors with RhC

and/or RhE positive

Used the BLOODchip Reference platform

Total of 2.200 blood donors extensively genotyped

1.6%

Blood Donors Molecular Typing Barcelona Blood Center Experience



Finnish Red Cross Blood Service experience

Routine genotyping of blood donors with ID-Core+ since

September 2012

Criteria– blood group A or O, K neg

– previous donation within a year

– rare donors, ethnic minorities

Blood Donors Molecular Typing Finnish Red Cross Blood Service Experience



Agenda








Regulatory Process for Serology• Biologics License Application (BLA)

• Applicable regulations• 21 CFR 600-660• 21 CFR Part 809 10 (Labeling)• 21 CFR Part 820 (Quality System Regulations)

Definitions:• Biological Product – 21 CFR Part 600.3 (h)

• Blood Grouping Reagent - 21 CFR Part 660, Subpart C

• Reagent Red Blood Cells – 21 CFR Part 660, Subpart D

• Anti-Human Globulin – 21 CFR Part 660, Subpart F



Hemagglutination assays (HA) defined immunohematology

• Over 300 blood group antigens recognized

Reagent Approvals - FDA

• Polyclonal antisera

• Monoclonal antisera

• Anti-Human Globulin (AHG)

• Panel or screening cells with known antigens


Regulatory Consideration

High degree of accuracy has been reported

FDA established molecular testing laboratory to created DNA reference panels

No licensed or approved molecular device for RBC genotyping

Pathway• Investigational device exemption (IDE)

• Premarket approval (PMA)- Assurance of safety and effectiveness

- Scientific evidence

- 180-day review clock


Regulatory ConsiderationRBC Genotyping Workshop, NIH

RBC genotyping promises significant technical and clinical benefits for transfusion medicine

Assay systems for RBC genotyping will be reviewed under the PMA pathway

FDA currently considers that:• In-house validation of RBC genotyping systems will require panels

of specimens pedigreed by gene sequencing of the donor

• Candidate technology should be defined within review process to permit appropriate labeling, limitations, and comparison to serology

Guidance and policy will be needed

Dr. Epstein, Director of CBRR, Sept 14, 2012


Agenda







Blood Product Advisory CommitteeSupports integral labeling with historic antigen type results

December 2012, BPAC asked to comment on • Reporting based on historical RBC typing results from two donations

• Validated process to confirm donor identification and accurate linkage to historical data

• Confirmation of relevant negative results on the current unit prior to transfusion, when feasible to mitigate risk form historical data

• Would BPAC’s response to the first questions vary if serology or molecular testing, or both were performed.

Committee supported historical data as part of the label and their responses would not vary whether the test was serologic or molecular


Thank you

The secret to complex serology is in the genes

is it time to re-evaluate the gold standards in blood banking ?

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