Comprehensive Study Guide for Transfusion Medicine and Blood Group Systems

Donor Types and Selection Criteria

• Allogeneic Donor: General donor population providing blood for others.

• Autologous Donor: Individual donating blood for their own future use.

  • Hemoglobin ($Hb$) must be $\ge 11.0\,g/dL$ ($\ge 110\,g/L$).

  • Hematocrit ($HCT$) must be $\ge 33\%$.

  • Blood collection must occur more than $72$ hours before the scheduled surgery.

  • Bacteremia is a definitive cause for deferral.

  • Units are labeled specifically and used only for autologous transfusion.

• Directed or Designated Donor: A specific donor chosen by or for a patient, often when rare Antigen-negative ($Ag$-negative) blood is required.

  • Must meet the exact same eligibility requirements as an allogeneic donor.

  • If not needed by the intended recipient, these units may be used for other patients.

Collection of Donor Blood

• Venipuncture Site Preparation: Employs aseptic technique. The site must be scrubbed for a minimum of $30$ seconds using providine-iodine scrub or isopropyl alcohol to minimize bacterial contamination risk.

• Standard Collection Volume: $450\,mL \pm 10\%$ or $500\,mL \pm 10\%$, depending on the collection bag used.

• Maximum Collection Limit: $10.5\,mL$ of whole blood per kilogram ($kg$) of donor weight, which includes samples taken via a diversion pouch or a second phlebotomy.

• Low-Volume Collection: Defined as $300\text{--}404\,mL$ in a $450\text{--}mL$ bag or $333\text{--}449\,mL$ in a $500\text{--}mL$ bag.

  • Must be specifically labeled as low-volume.

  • Red Blood Cells ($RBCs$) may be transfused from these units, but the plasma and platelets should be discarded due to improper anticoagulant-to-blood ratios.

• Anticoagulant Volumes:

  • $63\,mL$ of anticoagulant for a range of $400\text{--}500\,mL$ (average $450\,mL$).

  • $70\,mL$ of anticoagulant for a range of $450\text{--}550\,mL$ (average $500\,mL$).

• Collection Duration: Typically takes less than $10$ minutes. If the collection lasts longer than $15\text{--}20$ minutes, the unit may be unsuitable for the preparation of platelets or plasma.

• Materials: Collection bags are made of polyvinyl chloride.

Blood Anticoagulants, Additives, and Storage Lesions

• Expiration Limits for Anticoagulants and Additives:

  • $ACD$, $CPD$, and $CPD2$: $21$ days.

  • $CPDA-1$: $35$ days.

  • Additives (e.g., Saline adenine glucose mannitol or $SAGM$): Extend shelf life to $42$ days.

• Additive Processing: Plasma is expressed from Whole Blood ($WB$), and the additive is transferred from an attached satellite bag to the $RBCs$ within $72$ hours of collection.

• Hematocrit Standards: $55\%\text{--}65\%$ with additives; $65\%\text{--}80\%$ without additives.

• Rejuvenating Solutions: Used to restore $2,3\text{-}DPG$ and $ATP$ levels in stored units.

• Storage Lesion (Changes during storage):

  • Increased: Plasma $Hb$, Plasma $K^{+}$, Lactic acid, and Microaggregates.

  • Decreased: Viable $RBCs$, Plasma $Na^{+}$, Plasma $pH$, $RBC$ $ATP$, $RBC$ $2,3\text{-}DPG$, Glucose, and Coagulation factors.

Apheresis and Specialized Collection

• General Definition: An automated blood collection system that removes one or more specific components and returns the remainder to the donor. It reduces the number of donors a patient is exposed to.

• RBC Apheresis: Allows for the collection of $2$ units at the same time from donors with higher $HCT$.

• Plateletpheresis:

  • Used to collect $HLA$ matched platelets for patients refractory to random platelets.

  • Units can be leukoreduced during collection.

  • Must contain $\ge 3 \times 10^{11}$ platelets.

  • Frequency for single plateletpheresis: Every $2$ days (maximum $2$ times in $7$ days; maximum $24$ times in $12$ months).

  • Frequency for double and triple plateletpheresis: Every $7$ days.

  • Donor platelet count must be $\ge 150,000/\mu L$ (measured before or after the procedure).

• Plasmapheresis:

  • Frequent donation: Every $2$ days (maximum $2$ times in $7$ days).

  • Infrequent donation: Every $4$ weeks (maximum $13$ times per year).

• Granulocytes (Leukapheresis): For neutropenic patients. Collected every $2$ days (maximum twice a week, $24$ times per year).

• Stem Cells: Used for bone marrow reconstitution in patients with cancer, leukemia, or lymphoma. Collected from bone marrow, umbilical cord blood, or peripheral blood ($ABO$ compatibility not required).

• Therapeutic Plasmapheresis (Plasma Exchange): Removes abnormal plasma proteins, immune complexes (e.g., in $SLE$), alloantibodies, autoantibodies, Immunoglobulins (e.g., in $WM$), and lipoproteins. Replaced with crystalloid, albumin, or $FFP$.

• Therapeutic Cytapheresis: Removes cellular elements (e.g., exchange transfusion for sickle cell patients, leukemic $WBCs$, or abnormal platelet/lymphocyte counts).

Blood Component Storage and Expiration

• Thawed $FFP$: Stored at $1\text{--}6^{\circ}C$. Expires after $24$ hours.

• Thawed Plasma: Stored at $1\text{--}6^{\circ}C$. Expires after $5$ days.

• Cryoprecipitate $AHF$: Stored at $\le -18^{\circ}C$. Expires after $12$ months.

• Pooled Cryoprecipitate (after thawing):

  • Closed system: Expires in $6$ hours.

  • Open system: Expires in $4$ hours.

  • Stored at $20\text{--}24^{\circ}C$.

• Platelets (Single or Apheresis): Stored at $20\text{--}24^{\circ}C$ with continuous agitation. Expires in $5$ days.

• Pooled Platelets (Open system): Only one $ABO$ type per pool. Expires in $4$ hours. Stored at $20\text{--}24^{\circ}C$ with continuous agitation.

• Apheresis Granulocytes: Stored at $20\text{--}24^{\circ}C$ without agitation. Expires in $24$ hours.

• 
  

  Quality Control ($QC$) Requirements:

  • Cryoprecipitate: Factor $VIII$ $\ge 80\,IU$ and Fibrinogen $\ge 150\,mg$.

  • Platelets: $pH$ $\ge 6.2$ at the end of storage in $40\text{--}70\,mL$ plasma.

Transfusion Practice and Administration

• Platelet Count Elevation:

  • $1$ unit of whole blood-derived platelets raises count by $5,000\text{--}10,000/\mu L$.

  • $1$ apheresis unit (equivalent to $4\text{--}6$ units) raises count by $20,000\text{--}60,000/\mu L$.

• Administration Rules:

  • Informed consent from recipient is mandatory.

  • Must start before component expiration and complete within $4$ hours (ideally $1\text{--}2$ hours) to prevent bacterial growth.

  • Direct observation for the first $15$ minutes of infusion; periodic observation thereafter.

• Reissue of Blood: $RBC$ units must return to the lab within $30$ minutes if not kept at $1\text{--}6^{\circ}C$. Container closure must be undisturbed, and at least one sealed segment must remain attached.

• Universal Selection:

  • $RBC$ Transfusion: Group $O$ $Rh\text{-}$ is the universal donor; Group $AB$ $Rh+$ is the universal recipient.

  • Plasma Transfusion: Group $AB$ is the universal donor; Group $O$ is the universal recipient.

Adverse Effects of Transfusion

• Classification by Timing:

  • Acute: Occurs immediately or within $24$ hours.

  • Delayed: Occurs after $24$ hours.

• Top Causes of Transfusion-Associated Mortality:

  1. $TRALI$ (Transfusion-Related Acute Lung Injury).

  2. Hemolytic Transfusion Reactions.

  3. $TACO$ (Transfusion-Associated Circulatory Overload).

Genetics of Blood Groups

• International Society of Blood Transfusion ($ISBT$): Standardizes nomenclature.

• Definitions:

  • Genotype: Actual genetic makeup.

  • Phenotype: Physical expression of inherited traits.

• Antigen Types:

  • Oligosaccharide/Carbohydrate: $ABO$, $H$, $Le$, $P1PK$, $I$, $GLOB$.

  • Protein: $Rh$, $FY$, $JK$, $KEL$, $MNS$.

• Inheritance Patterns:

  • Most are autosomal dominant or autosomal codominant.

  • Inheritance of $A$ and $B$ genes results in expression of both ($Codominant$).

  • Group $O$ is autosomal recessive; the $O$ gene is an amorph, producing no detectable antigen.

The ABO and H Blood Group Systems

• Significance: Most important system; incompatible transfusions cause immediate hemolysis, renal failure, and death.

• Landsteiner’s Rule (1900-1901): Individuals naturally possess antibodies to the $ABO$ antigens they lack.

• Antibody Characteristics: Naturally occurring; detectable in infants at $3\text{--}6$ months, reaching adult levels by age $5$.

• Genetics:

  • $ABO$ locus on Chromosome $9$.

  • $H$ gene ($FUT1$) and $h$ allele on Chromosome $19$.

  • The $H$ gene must be inherited to produce the $H$ antigen, which is the precursor substrate for $A$ and $B$ antigens.

  • Secretor gene ($Se$ or $FUT2$) on Chromosome $19$ regulates $ABO$ antigens in secretions (saliva, tears, etc.).

• Antigen Precursors:

  • Type $1$ oligosaccharide: Present in secretions.

  • Type $2$ oligosaccharide: Present on $RBCs$.

• H Antigen Quantity by Blood Group: $O > A2 > B > A2B > A1 > A1B > \text{para-Bombay} > \text{Bombay}$.

• Subgroups: $80\%$ of group $A$ are $A1$; $20\%$ are $A2$. They are differentiated using anti-$A1$ lectin (Dolichos biflorus).

• Bombay ($O_{h}$) Phenotype: $hh$ genotype. Lacks all $ABH$ antigens and possesses potent anti-$A$, anti-$B$, anti-$A,B$, and anti-$H$. They can only receive blood from another Bombay individual.

ABO Discrepancies

• Definition: Red cell (forward) grouping does not match serum (reverse) grouping. Reactions are typically strong ($3+$ to $4+$).

• Forward Grouping Discrepancies (Red Cell Related):

  • Subgroups of $A$ (e.g., $A2$ with anti-$A1$).

  • Rouleaux formation (pseudoagglutination) due to high globulin (Multiple Myeloma, Waldenström's Macroglobulinemia) or fibrinogen.

  • $O_{h}$ Bombay phenotype.

  • Leukemia or Hodgkin’s disease (weakened antigens).

  • Chimerism or transfusion history (mixture of cell types).

• Reverse Grouping Discrepancies (Serum Related):

  • Age extremes (newborns or elderly).

  • Hypogammaglobulinemia/Immunodeficiency.

  • Cold reacting antibodies (anti-$H, I, M, N, P1, Lewis$).

  • Prozone/Postzone effects or plasma expanders.

The Rh Blood Group System

• Immunogenicity: Second most important system. Antigen immunogenicity order: $D > c > E > C > e$.

• Genetics: Two closely linked codominant genes ($RHD$ and $RHCE$) on Chromosome $1$ control the expression of $D, C, E, c, e$.

• Weak D Antigen:

  • Weak D (Quantitative): Reduced quantity of $D$ antigen; no anti-$D$ produced.

  • Partial D (Qualitative/Variant): Missed epitopes on the $D$ protein; can produce anti-$D$ against the missing parts. Must receive $D\text{-}$negative blood.

  • Testing: Requires Indirect Antiglobulin Test ($IAT$) phase at $37^{\circ}C$. Donor blood must be tested for weak $D$.

• Nomenclatures:

  • Fisher-Race: Uses $D, C, E, c, e$ (e.g., $DCe/dce$).

  • Wiener: Uses shorthand like $R_{1}, R_{2}, R_{0}, r$ (e.g., $R_{1} = DCe$, $r = dce$, $R_{2} = DcE$).

  • Rosenfield: Alphanumeric ($Rh:1$ is $D$, $Rh:2$ is $C$, etc.).

• Rh Rh Antibodies: Immune origin (pregnancy/transfusion); primarily $IgG$; react at $AHG$ phase. Capable of causing severe Hemolytic Disease of the Fetus and Newborn ($HDFN$) and Hemolytic Transfusion Reactions ($HTR$).

Minor Blood Group Systems

• Lewis ($LE$): Alleles $Le, le$. Antigens ($Le^{a}, Le^{b}$) are adsorbed from plasma onto cells, not synthesized by $RBCs$. Not on cord cells. Phenotype $Le(a\text{-}b-)$ is common in pregnancy.

• I System:

  • Cord blood: $i$+ , $I$ -.

  • Adult blood: $I$+ , $i$ -.

  • Anti-$I$ is an $IgM$ autoantibody associated with Mycoplasma pneumoniae.

  • Anti-$i$ is associated with Infectious Mononucleosis ($IM$).

• P1PK and GLOB:

  • Anti-$P1$: IgM neutralized by hydatid cyst fluid.

  • Anti-$P$: IgG autoantibody (Donath-Landsteiner) found in Paroxysmal Cold Hemoglobinuria ($PCH$).

• MNS System: Antigens $M, N, S, s, U$. Anti-$M, N$ are usually $IgM$; Anti-$S, s, U$ are $IgG$ and clinically significant. The $S\text{-}s\text{-}U\text{-}$ phenotype is found in individuals of African descent.

• Kell ($KEL$): Antigen $K$ is highly immunogenic. Antigens are destroyed by $DTT$ and $AET$. McLeod phenotype is $X\text{-}$linked and associated with Chronic Granulomatous Disease ($CGD$).

• Duffy ($FY$): Antigens $Fy^{a}, Fy^{b}$. $Fy(a\text{-}b-)$ phenotype provides resistance to Plasmodium vivax.

• Kidd ($JK$): Antigens $Jk^{a}, Jk^{b}$. Antibodies show dosage and are notorious for causing Delayed Hemolytic Transfusion Reactions ($DHTR$) because titers drop below detectable levels quickly.

• Lutheran ($LU$): $Lu^{b}$ is high prevalence; $Lu^{a}$ is low prevalence. Anti-$Lu^{a}$ shows mixed-field reactivity.

HLA and Platelet Antigens

• Human Leukocyte Antigens ($HLA$):

  • Class I: Platelets, leukocytes, nucleated cells.

  • Class II: Antigen Presenting Cells ($APCs$) like B-lymphocytes and macrophages.

  • Associations: $HLA\text{-}B51$ with Behçet disease.

  • Complications: Febrile Non-Hemolytic Transfusion Reaction ($FNHTR$), $TRALI$, and Graft-versus-Host Disease ($GVHD$).

• Platelet Specific Antigens: Human Platelet Antigen ($HPA\text{-}1a$) is associated with Neonatal Alloimmune Thrombocytopenia ($NAIT$) and Posttransfusion Purpura ($PTP$).

Pretransfusion and Compatibility Testing

• Coombs’ Test (Antiglobulin Test):

  • Direct Antiglobulin Test ($DAT$): Detects in vivo sensitization ($IgG$ or complement on patient cells). Used for $HDFN$, $AIHA$, and transfusion reactions.

  • Indirect Antiglobulin Test ($IAT$): Detects in vitro sensitization (antibodies in serum). Used for antibody screening, crossmatching, and weak $D$ testing.

• Enhancement Media (Potentiators):

  • LISS (Low-Ionic-Strength Saline): Speeds up antibody uptake.

  • PEG (Polyethylene Glycol): Removes water to increase antibody concentration.

  • Proteolytic Enzymes (Ficin, Papain): Enhance $Kidd$, $Lewis$, $Rh$, $P$, $I$; destroy $M, N, S, s, Duffy$.

  • $22\%$ BSA: Reduces red cell charge.

• Crossmatch:

  • Major: Patient plasma vs. Donor cells.

  • Minor: Donor plasma vs. Patient cells.

  • Check Cells: IgG-sensitized cells added to negative results to confirm $AHG$ was added and not neutralized.

Hemolytic Disease of the Fetus and Newborn (HDFN)

• Pathophysiology: Maternal $IgG$ crosses the placenta and attacks fetal $RBCs$ containing paternal antigens.

• Rh HDFN: Anti-$D$ formed by $Rh\text{-}$ mother; usually affects subsequent pregnancies; severe anemia/bilirubinemia.

• ABO HDFN: Usually occurs in Group $O$ mothers with Group $A$ or $B$ infants; can affect the first pregnancy; typically mild.

• Management of IUT (Intrauterine Transfusion):

  • Requires Group $O$ $Rh\text{-}$ negative RBCs.

  • Must be fresh (less than $7$ days old) to avoid hyperkalemia.

  • Must be irradiated ($prevent GVHD$), Leukoreduced, $CMV$ seronegative, and negative for $Hb\text{-}S$.

• Prevention: Rh Immune Globulin ($RhIG$/RhoGAM). Administered at $26\text{--}28$ weeks gestation and within $72$ hours of delivery to $Rh\text{-}$ mothers.

• Fetal Maternal Hemorrhage ($FMH$) Evaluation:

  • Rosette Test: Qualitative screen for $Rh+$ fetal cells.

  • Kleihauer-Betke ($KB$) Acid Elution: Quantitative test where fetal $HbF$ resists acid elution (stays pink) while maternal cells become ghosts.

Questions & Discussion

• Q: A nurse from Nigeria living in Saudi Arabia for 2 years wants to donate blood.

  • Considerations: Malaria endemicity/travel history.

• Q: A woman lived in London (1993-1994), Italy (1999-2000), and Australia (2009-2015). Why defer?

  • Deferral related to potential exposure to Variant Creutzfeldt-Jakob Disease ($vCJD$) in the UK/Europe.

• Q: Screening shows $HBsAg$ reactive, Anti-$HBc$ reactive, Anti-$HBs$ non-reactive. Cause for deferral?

  • Indicates active/chronic Hepatitis B infection.

• Q: Can a sickle cell trait individual donate?

  • Yes, though the blood may not be suitable for specific procedures like IUT.

• Q: Donation for money?

  • In many jurisdictions, this is prohibited; volunteer donation is the standard.

• Q: Heterozygous C genotype?

  • Result: $dcE/dce$.

• Q: Fisher-Race antigens for individual with anti-c?

  • Result: $DCe/DCe$ (Must lack the 'c' antigen).

• Q: Interpretation of Anti-D titer 256 for IUT?

  • Blood used should be Group $O$, $D\text{-}$negative $RBCs$.