ImmunohematologyHemovigilance

Slaviková M., Zemanová M., Galuszková D.Transfusion Department, Faculty Hospital Olomouc

Introduction

• First part:• Blood Group Systems

– AB0 system – Rh system

• Other blood group systems• Techniques used in blood grouping

• Second part:• Hemovigilantion

What is blood group?

• Definition– An inherited character of the red cell

surface, detected by a specific alloantibody.

• Blood group antigens may be:– Proteins

– Glycoproteins

– Glycolipids

Blood group systemsThere are over 300 known red blood group antigens

30 blood group systems

Terminology and classification

• ISBT classification

• Based on blood group systems

• Each antigen within that system has a three digit number plus a three to five upper case letter symbol

http://blood.co.uk/ibgrl/

Other blood group systems

• All authenticated antigens fall into one of four classifications:

• A blood group system consists of one or more antigens controlled at a single gene locus, or by two or more very closely linked homologous genes with little or no observable recombination between them.

• Collections consist of serologically, biochemically, or genetically related antigens, which do not fit the criteria required for system status.

• The 700 Series contains antigens with an incidence of less than 1% and which cannot be included in a system or collection.

• The 901 Series contains antigens with an incidence of greater than 90% and which cannot be included in a system or collection.

Blood group systems

• AB0• MNS• P• Rh• LUTHERAN• KELL• LEWIS• DUFFY• KIDD • DIEGO• Yt• Xg• SCIANNA • DOMBROCK • COLTON• LANDSTEINER-WIENER

• CHIDO/RODGERS• H• Kx• GERBICH• CROMER• KNOPS • INDIAN• Ok• RAPH• JOHN MILTON HAGEN• I• GLOBOSIDE• GILL • Rh-ASSOCIATED

GLYCOPROTEIN

Blood group collections (200)

ACTUALLY:

• 205: COST

• 207: Ii

• 208: Er

• 209: GLOB

• 210: Lec; Led

• 212: VEL

OBSOLETE COLLECTIONS:

201 Gerbich

202 Cromer

203 Indian

204 Auberger

206 Gregory

211 Wright

Low incidence antigens (700 series)

• BATTY• CHRISTIANSEN • BILES• BOX• TORKILDSEN • PETERS• REID• JENSEN• LIVESAY• MILNE

High incidence antigens (901 series)

• LANGERIES• AUGUST• Jra

• EMM• ANTON

The AB0 blood groups

• In 1900 discovery by Landsteiner

• There are four phenotypes: A, B, AB and 0

• Individuals lacking A or B antigen form their red cells have the coresponding antibody in their plasma (Landsteiner’s rule)

AB0 antigens

Rh blood group system

• Discovered in New York in 1939 with an antibody in serum of women who had given birth to a stillborn baby and then suffered a hemolytic reaction as the result of transfusion with blood from her husband

• Rh phenotypes are controlled by two genes :

– RHD which encodes D antigen

– RHCE which encodes Cc and Ee antigens

DDDDDD

E/eE/e

C/cC/c

E/eE/eE/eE/e

C/cC/cC/cC/cC/cC/c

RhD neg.RhD neg.RhD RhD pospos.. RhD neg.RhD neg.RhD RhD pospos..

Techniques used in blood grouping

• Routine blood grouping still relies primarily on haemagglutination reactions between antigens and antibodies that are read and interpreted either manually or by automated means

Ab Rbc Haemagglutination

Factors affecting antigen-antibody reactions

• Temperature• Antibodies that promote acclerated

red cell destruction in vivo (apart from antibodies of the AB0 system) react optimally at 37 °C!!!

• Time and ionic strenght• Low Ionic Strenght Saline (LISS)• By reducing the numbers of free ions

in solutions, this neutralising effect is diminished thus enhancing the speed at which the antigen-antibody complexes are formed

• pH• Optimal pH is 7 – 7,5

• Antigen density• The number of antigen site per cell

and the accesibilityof those antigens affects the optimal uptake of apropriate antibody

Routine laboratory testing

• Saline test• Fyziologic solution• Detection of IgM antiglobulin class

antibodies

• Direct agglutination test• Indirect agglutination test• Enzyme test

Enzyme techniques• Enzymes often used in blood group serology

are the proteases:– bromelin, ficin, and papain which have broad

specifity for peptide bonds– Trypsin, chymotripsin, which have more precise

specifities– Neuraminidase, which cleaves sialic acid residues

Enzyme treated cells (ETC):

• are used in antibody screening and, importantly, in antibody identification procedures

• They are particularly useful for determing specifities within mixtures of antibodies, where one or more are directed against enzyme-sensitive antigens

Pretransfusion compatibility testing

• Pretransfusion testing involves these tests:

• Determining the ABO and RhD types of both patient and donor blood

• Screening patient sera (or plasma) for RBC alloantibodies

• And performing a major crossmatch(testing the patient's serum against the donor's RBCs)

Sample and requirement

6 ml K3EDTA

AB0 blood grouping in laboratory

• Blood presure, temperature

• Bed-side test (EBR - ABO compatibility)

• observation

• complications: blood sample from patient + TU, repeat testing

Pretransfusion testing - patients

Prenatal and PerinatalImmunohematologic Testing

• AB0 RhD typing: all women should be tested for ABO and D as early as possible in pregnancy, preferably during their first trimester visit. ABO typing is done primarily for patient identification.

• Antibody screening• Positiv antibody screening

– ID antibodies• Antibody titer: Rh or

other clinically signficant antibody

Prenatal and PerinatalImmunohematologic Testing

• Not all clinically significant antibodies detected during pregnancy will cause HDN.

• The mother's antibody may have been induced during a previous pregnancy by a different consort, in which case the current fetus may be antigen negative and not at risk for HDN.

• It is worthwhile to phenotype RBCs from the putative father whenever the potential for HDN exists. The probability that a fetus carries the antigen to which the maternal serum contains antibody can be estimated from maternal and paternal phenotypes. If there is a high likelihood that the father is heterozygous for the gene encoding the offending antigen, molecular genotyping of amniotic fluid cells or chorionic villi can performed. PCR assays are available for for Rh, Kell, Duffy or Kidd genotypes.

•standard practise in hospitals is to transfuse group-specific red cellsto all recipients, but group O red cells may be transfused for all patients, especially neonates or patients requiring urgent transfusion before their blood group is known

• some group O donors may have high titre of anti-A and anti-B in their plasma, which could cause lysis of A and or B cells, so, blood group O is tested for high titre anti-A and anti-B

•many blood centres also undertake a more comprehensive red cell antigen phenotyping (RhD, Kell, MNS, Duffy, Kidd, Lutheran, Lewis) in order to identify donors whose red cells could be used for transfusion to recipient whose plasma is known to contain clinically significant blood group antibodies or patients at high risk of forming multiple alloantibodies

Testing of blood donations

•laboratory screening test for HBV (HBsAg), HCV (anti-HCV), HIV (anti HIV 1+2, p24) and syphylis remains mandatory in many countries including Czech republic

•the application of polymerase chain reaction (PCR) methodology is available in some countries, an advantage is identifying viral infections at the earliest possible stages of infection and high sensitivity for detection

Microbiological testing

Hemovigilance definition

• TRIP (Transfusion Reactions in Patients)definition– Hemovigilance is the systematic monitoring of side

effects and adverse incidents throughout the chain from blood donor to recipient, and all other activities which can lead to safer and more effective use of blood components.

• EU Blood Directive 2002/98/EC Definition– Haemovigilance shall mean a set of organised

surveillance procedures relating to serious adverse or unexpected events or reactions in donors or recipients, and the epidemiological follow-up of donors.

Reporting procedure

• Reports of transfusion reactions are sent to to SÚKL on a voluntary basis and are treated confidentially.

• Who?– The hospital submits a report if a reaction

has been noted in the hospital.

•the detection and correction of errors

•strict protocol should be drawn up showing action to be taken in such an eventuality

• all staff in a blood transfusion service is trained to accept quality control as a necessary part of everyday work

•quality system is based on the principles of Good Practice and Quality management, as described in ISO 9001 standards

•the quality system should assure the quality and safety of blood and blood components and to fulfil customer satisfaction

Quality system

• the area for blood donors and laboratory areas should be separated from all processing areas

• storage conditions should be controlled, monitored and checked, appropriate alarms should be present and regulatory checked, the checks should be recorded

• all equipment should be designed, validated and maintained to suit its intended purpose, maintenance, cleaning and calibration should be performed regularly and recorded, instruction for use, maintenance, service cleaning and sanitation should be available

• only reagents and materials from approved suppliers should be used

Quality system

• each activity which may affect the quality of the blood and blood components should be documented and recorded

• documentation to ensure that work performed is standardised and that there is traceability of all steps in the process

• all changes to documents should be acted upon promptly and should be dated and signed by an authorised person

Documentation

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