HIS26 Introduction To Immunohaematology And Hospital Blood Banking Flashcards
Blood groups
Expression of Ag on RBC membrane
- proteins / carbohydrates
- many blood groups (ABO, Rh, Duffy, Kidd etc.)
- ABO, Rh most important (∵ most immunogenic)
Significance of different blood groups depends on:
- Frequency of Ag in population
- Effect of corresponding Ab on RBC
Significance of Blood group Ab
- Class/subclass of Ig
- Titre of Ab
- Thermal amplitude
—> Warm Ab: IgG (active at body temp)
—> Cold Ab: IgM (1-4oC) - Potency of Ab:
- Ability to fix complement
- Ability to cross placenta (haemolytic disease of fetus)
- Some Ab are naturally occurring e.g. Anti-A, Anti-B (***Isohaemagglutinins)
- Most Ab are acquired (after exposure to corresponding Ag)
***ABO blood group
Most important of all blood group systems
Naturally occurring Ab:
- Anti-A, Anti-B
- **IgM (Cold Ab but active at body temp)
- can fix complement —> **Haemolytic
- ***cannot cross placenta
AB Antigen on RBC:
- ***Oligosaccharides
some Anti-A / Anti-B can be immune in origin (i.e. IgG) —> can cross placenta (but haemolytic disease caused usually mild / self-limiting)
***ABO gene
- located on Chromosome 9
- encode enzyme with **Glycosyltransferase activity which **modify oligosaccharides on surface membrane glycoprotein
- default oligosaccharide structure - ***H-antigen (H gene on chromosome 19) —> precursor of A, B antigen
“A” allele: encode an enzyme adding ***N-acetyl galactosamine to H-antigen —> convert it to A antigen
“B” allele: encode an enzyme adding ***galactose to H-antigen —> convert it to B antigen
“O” allele: encode protein with ***loss of transferase activity (modify nothing on RBC) —> contain no Ag on RBC
ABO genotypes and phenotypes
Inheritance: ***Autosomal dominant
A, B: Co-dominant
O: Recessive
AA, AO —> A blood group
BB, BO —> B blood group
AB —> AB blood group
OO —> O blood group
Groupings of ABO blood group
- Cell grouping: presence/absence of A/B antigens on RBC
- Plasma / serum grouping: presence/absence of Anti-A/Anti-B in serum
If a particular Ag is present, there is no corresponding Ab
***Significance of ABO system
- Acute haemolytic transfusion reaction:
- Hyperacute ejection
- due to ABO incompatibility (ONLY reason)
- incompatible RBC lysed by ***Complements activated by naturally occurring IgM ABO Ab reactive at body temp
- fatal if unrecognised / untreated - ABO haemolytic disease of newborn (HDN):
- due to immune ABO Ab (IgG) by mother crossing placenta to destroy RBC of fetus
- more common in group O mothers (have Anti-A + Anti-B) with non-O fetus
- mild and self-limiting
***Acute vs Chronic Intravascular Haemolysis
Acute Intravascular Haemolysis:
—> free Haemoglobin released into blood
—> Haemoglobinaemia / Haemoglobinuria (vs haematuria: RBC in urine) / ↓ Haptoglobin level / ↑ Bilirubin level
Chronic Intravascular Haemolysis:
—> Fe deposition in urine / urothelial cells (i.e. renal tubules)
—> Haemosiderinuria (not detected until later phase)
***Rhesus system
- More than 50 antigenic specificities
- Transmembrane ***polypeptide
- Rh antigens high immunogenic
- Main antigenic specificities: D, C, c, E, e
- 2 closely linked genes: D and CcEe (CE, Ce, cE, ce)
- ***RhD: most immunogenic besides ABO system
Rh Ab:
- **Acquired after exposure to Ag that patient lacks (i.e. not pre-existing like in ABO)
- **Warm IgG
- can ***cross placenta
Example:
RhD -ve: no RhD Ag on RBC (of mother)
—> if given RhD +ve RBC transfusion (from 1st Rh+ve baby)
—> high chance to develop Anti-RhD Ab (in mother)
—> problems with subsequent transfusion (esp. for women at reproductive age) (2nd Rh+ve baby)
—> IgG: can cross placenta and cause HDN
***Significance of Rhesus system
- Transfusion reaction:
- due to immune Rh Ab
- usually **extravascular (∵ IgG) destruction of unlike RBCs
- **extravascular haemolysis: Anti-Rh attach to RBC —> when pass through splenic/liver vessels —> **macrophage activated —> RBC destroyed by **reticuloendothelial system
- Hyperbilirubinaemia, Haptoglobin ↓
- ***NOT see Haemosiderinuria / Haemoglobinemia / Haemoglobinuria - HDN due to Anti-D:
- most common HDN in Caucasians (10-13% Rh(D)-ve)
- Rh(D)-ve mother + Rh(D)+ve father —> Rh(D)+ve fetus
- 1st pregnancy not affected but after that mother acquire Anti-RhD Ab
- sensitisation usually occur during labour where fetal RBC enter maternal circulation
—> **Allo-immunisation
- subsequent pregnancies are affected (if fetus is Rh(D)+ve), usually severe
- mother should receive **Rh(D) Ig (i.e. Anti-RhD) as prophylaxis
—> ***destroy any Rh(D)+ve fetal RBC
—> prevent formation of Anti-RhD Ab
All Chinese can practically be considered Rh(D)+ve (only 0.3% Rh(D)-ve)
—> 唔怕阿媽produce Anti-RhD, 因為本身阿媽都係Rh(D)+ve —> 唔會develop Ab
Other blood group systems
Only significant if corresponding Ab is warm reacting
- Duffy, Kidd, Kell, Ss
- Transfusion reaction, HDN
Not significant if it is cold Ab
- MN, Lewis etc.
***Type and Screen (Pre-transfusion compatibility test)
Type: ABO and RhD blood group (睇RBC有咩Antigen)
- by commercially available Ab (i.e. Anti-serum)
- Anti-sera have known blood group specificity Ab e.g. Anti-A, Anti-D etc.
- DAT: RBC + Anti-serum —> Agglutination —> positive result
—> add Anti-human Globulin to augment agglutination reaction
Screen: Antibody screen (睇有咩Antibody)
- Positive (~7%) / Negative
- screen patient’s plasma for Ab against non-ABO but clinically significant blood group Ag
- formation / acquisition of Ab: through Sensitisation / Transfusion
- primary exposure: short duration, moderate response
- secondary exposure: long duration, larger response —> Anamnestic response (Ab titre ↑ greatly —> haemolysis)
- IAT: Recipient Serum + Donor RBC —> Agglutination —> positive result
—> add Anti-human Globulin to augment agglutination reaction
Principles of Blood bank testing
Separate blood into cells + serum
Plasma/Serum:
- Plasma/Serum grouping
- Antibody screen
- Crossmatch
Cells:
- Cell grouping
- Red cell phenotype
- ***Direct antiglobulin test (DAT)
Direct Antiglobulin test (DAT)
Detection of Antibody ***on RBC (see whether RBC are sensitised by Ab) (可以睇RBC有咩Antigen)
- test for presence of Ag-Ab reaction
- If RBC coated by human Ig
—> Anti-human-Ig Ab (AHG reagent) can be added
—> used to agglutinate RBC
Indirect Antiglobulin test (IAT)
Detection of Antibody **in serum (睇有咩Antibody)
- basis of **Ab screening and identification
- Ag-Ab reaction
- Incubate recipient serum (i.e. Ab) with ***donor RBC (i.e. Ag) (usually a panel of RBC is employed)
—> Ab will only coat RBC with corresponding Ag
—> 再add AHG agent to augment reaction
—> agglutination
—> Since Ag of RBC is known —> the specificity of Ab can be deduced
Crossmatch
Screen Recipient Serum —> Pair with Donor RBC
Electronic Crossmatch (用電腦配對 rather than 用人手溝血睇下是否compatible)
- After fulfilling certain criteria e.g. Ab screen negative (not have any active RBC atypical Ab) with confirmed ABO blood group
—> computer will select donor RBC with compatible ABO / Rh blood group
—> much more efficient, safe
—> For most patients with Ab-negative status, random but ABO RhD compatible blood can be issued through electronic crossmatch
Unmatched blood = “uncrossmatched” blood (not undergone screening yet)
- may be used if there is urgent need of transfusion when Ab screening has not been completed (~ 30-45 mins)
- there is chance in patient having Ab in serum, but only 7% chance
—> either unmatched group O blood (if patient’s blood type is not known) / unmatched ABO group specific blood
Unmatched group O (esp Rh -ve) blood: can just give right away (since no Ag on RBC)
Unmatched ABO group specific blood: if you can wait for ABO, RhD results ~25 mins
***Immune haemolytic anaemia
- Autoimmune haemolytic anaemia (AIHA)
Warm Ab type
- idiopathic
- associated with autoimmune diseases, lymphoproliferative disorders, drugs etc.
Cold Ab type
- cold agglutinin syndrome: idiopathic / associated with infections (infectious mononucleosis and mycoplasma) / LPD
- paroxysmal cold haemoglobinuria (rare, biphasic cold Ab)
- Alloimmune haemolytic anaemia
- Haemolytic transfusion reaction
- Haemolytic disease of newborn (HDN) - Drug-induced immune haemolytic anaemia
- Methyldopa
Serological diagnosis of Immune haemolytic anaemia
Autoimmune HA:
- Positive DAT —> RBC sensitised by Ab
- Positive IAT —> **AutoAb with **broad specificity (react with all RBC in panel)
- Important to exclude primary disorders, drugs
Alloimmune HA:
- Usually Positive IAT and/or DAT where **AlloAb with **defined specificity (patient RBC lack corresponding Ag)
- Usually history of transfusion / pregnancy
Clinical transfusion practice: Preparations for transfusion
- Whole blood collected in closed system of multiple plastic bags
—> separated by centrifugation into different components - Harvest of specific blood component by ***apheresis (platelets, plasma, leukocytes)
—> return other components back to donor (collect what you want)
—> collected dose (single donor unit) already equivalent to 5 donor units
—> better because purely from one donor (whereas normally one unit of specific blood components formed from several donors)
—> limited by availability of donor, takes more time (~2 hours) - Synthetic substances for transfusion (future direction)
- Other pharmacological agents (e.g. growth factors, fibrinogen concentrate etc.)
Blood components
- RBC: in additive
- Platelets: random donor platelets / apheresis platelets
- Plasma
- Leukocytes
- Cryoprecipitate: provide clotting factors
- Cryoreduced plasma: seldom used, used to treat TTP
Transfusion principle
***Only give what the patient needs
- RBC to proven anaemic patients
- Platelets to proven thrombocytopenic patients
- Plasma to selected patients with coagulation problem / bleeding
Special blood components
For special clinical situations
- Irradiated blood components
- for severely immunocompromised patients e.g. post-BM transplantation
—> to prevent transfusion associated ***graft vs host disease - Leukocyte-depletion blood components
- remove Buffy coat / filtered by bedside filter or pre-storage filtration
Indications:
- prevention of ***FNHTR (Febrile non-haemolytic transfusion reaction) (Ab to transfused WBC —> febrile reaction)
- prevention of ***HLA alloimmunisation (development of Ab against WBC)
—> otherwise problem with future transplantation / platelet transfusion (for patients on long-term transfusion support / potential BMT recipient) - prevention of ***CMV transmission for CMV negative recipients (CMV hide inside WBC)
Acute and Delayed complications of transfusion (less important)
Acute:
1. Acute haemolytic transfusion reaction (ABO incompatibility)
2. Non-haemolytic febrile transfusion reaction (Ab to transfused WBC)
3. Urticaria (Ab to proteins in transfused blood components)
4. Pulmonary oedema (fluid overload)
5. Infective shock (bacterial contamination of platelet concentrates)
6. Anaphylaxis, transfusion related acute lung injury
Delayed:
1. Delayed haemolytic transfusion reaction (patient already immunised against specific RBC Ag —> extravascular haemolysis 5-10 days later)
2. Development of Ab to transfused RBC (Alloimmunisation)
3. Development of Ab to transfused WBC / platelets (Alloimmunisation to HLA antigens —> NHFTR, platelet refractoriness)
4. Iron overload
5. Transfusion-transmitted infections (usually viral infections: HIV, HBV, HCV, CMV, HTLV1)
Transfusion safety
Regional Blood Transfusion Service i.e. Hong Kong Red Cross Blood Transfusion Service
Hospital Transfusion service:
- Hospital Transfusion Committee
- Hospital Blood Bank
- Front-line medical / nursing staff
Corporate:
- Central Committee (Transfusion Service)
Role of Regional BTS in transfusion safety
- Stringent donor selection criteria
- Sensitive virological / microbiological assays
- Comprehensive serological workup
- Proper production protocols for preparation of blood components
- Proper storage conditions
- Strict quality assurance programme
Hospital transfusion service
Multi-disciplinary
- Medical: Prescription of transfusion (type, amount, and degree of urgency), sample collection, checking and monitoring
- Nursing: Checking and monitoring
- BB: Bank, maintaining stock level, pre-transfusion compatibility testing, issue of blood/blood components
Organisation of Hospital Trasnfusion service
Hospital Transfusion Committee (set up in all hospitals with transfusion service)
—> Hospital blood bank
—> Users of all disciplines
Responsibilities of the HTC:
Coordinate and oversee all matters related to blood transfusion in hospital
- formulate transfusion policy
- issue transfusion procedural guidelines
- monitor usage of blood / blood components
- collection and investigation of incidents related to transfusion
- education of hospital staff
Right sample from right patient for whom the transfusion is intended
Positive identification of patient:
- ask open-ended questions (What is your name)
- check identification information on bracelet
- verify prescription of transfusion on patient’s file
- Patient’s identification information: Name, HKID, Sex, DOB
Handle one patient at a time
Informed consent in transfusion
- Information provided by physician
- risk
- benefits
- alternative treatment e.g. autologous transfusion - Opportunity for questions and clarification
- patient competency
- patient (or surrogate decision maker) understands
- patient decides on basis of complete information - Patient agrees / refuses
- Documentation
Patient blood management targets
- Optimise patient’s Hb
- recognition, identification of causes and management of anaemia rather than just topping up Hb
- medical setting
- peri-operative setting - Minimise blood loss
- intra-operative cell salvage (recovering blood lost during surgery and re-infusing it into the patient)
- POCT-guided transfusion therapy
- better management of haemostasis (early coagulative therapy) - Rationalise transfusion
- judicious transfusion
- use alternatives of transfusion
- ***single unit transfusion followed by patient assessment
Alternatives to Red cell transfusion for patients with Fe deficiency anaemia
Fe replacement:
- Oral iron
- FeSO4
- Ferrum Hausman Solution
- Ferrum Hausman chewable tablets
- (Fortifer) - IV iron
- Venofer
- Monofer
ALL are effective. Choose according to patients’ tolerance and urgency of clinical conditions
Every transfusion is a clinical decision
- Transfusion risk is cumulative with the number of unit transfused
- Before considering transfusion, consider alternative of transfusion i.e. proper management of anaemia
- If transfusion is needed, transfuse minimal amount, ***adopt single unit transfusion for non-bleeding patient and reassess the patient for further need of transfusion
- Don’t just treat the number, always consider patient’s symptoms and clinical context
Implementation of patient Blood Management programme
- Strategic planning (Corporate —> Hospital-based)
- Multidisciplinary approach (Medical/Surgical/Obstetric/ICU/Paediatric/Laboratory and Blood bank)
- Education and training
- Evidence-based, data-driven
- Continuous evaluation of effectiveness
Summary
Serology:
1. Blood group
- Blood group: important if Ab is warm reacting IgG
- ABO most important (naturally-occurring IgM reactive at body temperature that can fix complement)
- Rh not so important in Chinese
- Serological test
- Ag-Ab reaction
- DAT and IAT - Serological problems in clinical practice:
- Autoimmune vs Alloimmune HA
- Warm vs Cold types
- Always rmb drugs as potential culprit
Transfusion practices:
1. Pre-transfusion compatibility testing
- Type (ABO, Rh) and Screen
- Electronic XM
- Full serological XM for Ab screening positive patients
- Clinical transfusion practice
- ***Component therapy: give only what patient needs
- Specialised preparations for special circumstances
- Adverse transfusion reaction - Transfusion safety
- right specimen from right patient
- right unit for right patient
- positive identification of patient - Medicolegal aspect of transfusion: Informed consent
***Summary: Comparison between ABO and Rhesus system
ABO system:
AB Antigen on RBC:
- ***Oligosaccharides
Naturally occurring Ab:
- Anti-A, Anti-B
- **IgM (Cold Ab but active at body temp)
- **can fix complement —> **Intravascular haemolysis
- **cannot cross placenta
Rhesus system:
Rh Antigen on RBC:
- Polypeptide
- 2 closely linked genes: D and CcEe (CE, Ce, cE, ce)
- RhD: most immunogenic
Rh Ab:
- **Acquired after exposure to Ag that patient lacks (i.e. not pre-existing like in ABO)
- **Warm IgG —> **Extravascular haemolysis
- can **cross placenta
