IHBB FINALS

HIGH-YIELD EXAM REVIEWER: IMMUNOHEMATOLOGY (MLS-416) Lessons 10–13

LESSON 10: COMPATIBILITY TESTING

KEY DEFINITIONS

  • Compatibility testing = entire pretransfusion process (request → release of blood bag)

  • Crossmatching = only ONE part of compatibility testing

  • No Minor crossmatch performed anymore (redundant with antibody screening)

SPECIMEN OF CHOICE: Pink/Purple top (K2EDTA) — preferred over red top

  • Plasma preferred over serum (no fibrin clot, no false positives)

  • Samples must be tested within 72 hours (complement becomes unstable)

  • Hemolyzed samples = automatically rejected

PATIENT ID: 2 unique identifiers required (name + DOB or MRN)

CROSSMATCH TYPES:

Major

Minor

Sample

Patient serum + Donor RBCs

Donor serum + Patient RBCs

Detects

Patient antibodies vs donor RBCs

Donor antibodies vs patient RBCs

Required?

YES (routinely)

NO (since 1976)

3 PHASES OF CROSSMATCH:

  • IS (Immediate Spin) → detects IgM cold-reacting antibodies (Anti-Le, Anti-M, Anti-N, Anti-P)

  • 37°C (LISS/Thermophase) → detects IgM and IgG

  • AHG (Antihuman Globulin/IAT) → detects warm-reacting IgG (Rh, Kell, Duffy, Kidd)

COMPLETE vs INCOMPLETE CROSSMATCH:

  • Incomplete (IS only) → antibody screen NEGATIVE, no history of antibodies

  • Complete (IS + 37°C + AHG) → antibody screen POSITIVE or history of antibodies

ANTIBODY SCREEN vs ID:

  • Screen = qualitative (present or absent)

  • ID = identifies specific antibody (done only if screen is POSITIVE)

  • Uses 2–3 Screening Cells; ID uses 10+ Panel Cells

CROSSMATCH INTERPRETATION TABLE:

Ab Screen

Crossmatch

Action

Positive

Negative

Compatible; antibody vs screening cell, not donor

Negative

Positive

Incompatible; perform DAT, antibody ID

Positive

Positive

Incompatible; give antigen-NEGATIVE units

COMPUTER (ELECTRONIC) CROSSMATCH:

  • No actual mixing of samples

  • Requires TWO validated ABO results on file

  • Use only when antibody screen is NEGATIVE

LIMITATIONS OF COMPATIBILITY TESTING:

  • Cannot detect antibodies too weak to detect (e.g., Kidd — titer drops easily)

  • Cannot prevent clerical errors

  • Cannot prevent nonhemolytic reactions (cytokines not tested)

  • Cannot prevent TTIs

  • Only detects RED CELL antibodies

SPECIAL CIRCUMSTANCES:

Situation

Key Points

Emergency

O-negative pRBCs; physician must sign; crossmatch still done AFTER

Massive Transfusion

>5L or 10 units/24hrs; give ABO-identical units

Neonatal (<4 months)

No antibodies; ABO/Rh compatible blood; may use maternal serum for crossmatch

Intrauterine

Group O, D-negative; irradiated; CMV-safe; HbS-negative; <5–7 days old; Hct 70–85%

Autologous

Donor = patient; no crossmatch needed

BACTERIAL CONTAMINATION:

  • Normal plasma

  • Abnormal: RED or BLACK → bacterial contamination

  • Most common in platelets (stored at RT)

  • Common bacteria: Yersinia enterocolitica (cold-tolerant, RBCs), Pseudomonas aeruginosa

BLOOD BAG RETURN CRITERIA (memorize for MTLE):

  1. Unopened and intact (no leaks/punctures)

  2. Temperature maintained at 1–10°C

  3. Returned within 30 minutes if unrefrigerated

  • ANY LEAKAGE = DISCARD regardless of time

LESSON 11: DONOR SELECTION & BLOOD COLLECTION

GOVERNING AGENCIES:

  • USA: FDA (primary regulator), CMS (CLIA), AABB (standards), CAP, Joint Commission

  • Philippines: DOH-NVBSP; RA 7719 (voluntary donation); RITM TTI-NRL (TTI testing); NKTI (immunohematology)

DONOR CATEGORIES:

  • Allogeneic/Homologous/Random = general use

  • Autologous = for own use

  • Directed/Recipient-specific = for a specific patient

DONOR SCREENING (4 parts): Registration → Health History Interview → Physical Exam → Informed Consent

PHYSICAL EXAM REQUIREMENTS:

Parameter

Requirement

Weight

>50 kg / 110 lbs

Hgb (allogeneic female)

>12.5 g/dL (Hct >38%)

Hgb (allogeneic male)

>13.0 g/dL (Hct >39%)

Hgb (autologous)

>11.0 g/dL (Hct >33%)

Hgb (double RBC apheresis)

Hct >40%

Temperature

<37.5°C / 99.5°F

BP systolic

<180 mmHg

BP diastolic

<100 mmHg

Pulse

50–100 bpm (<50 acceptable for athletes)

Copper Sulfate Test:

  • Blood SINKS (SG >1.053 or 1.055) = ACCEPT

  • Blood FLOATS = REJECT (low Hgb)

DEFERRALS:

PERMANENT:

  • HIV/AIDS, HTLV I&II, Hepatitis B & C (chronic)

  • Prion disease (CJD) — cannot be destroyed; silent 4–20 years; no test available

  • History of parenteral drug use, prostitution

  • Tegison/Etretinate use (teratogenic, stored in fat)

  • Received clotting factor concentrate

  • Xenotransplantation

  • Autoimmune disease

INDEFINITE:

  • MSM (men-to-men sex) since 1977

  • Born/lived in African countries on FDA HIV-1 Group O list

  • CJD/vCJD related: spent 3 months UK (1980–1996), 5 years Europe/France (1980–present)

  • Leukemia, lymphoma, malignant melanoma

  • Babesiosis, Chagas disease

TEMPORARY DEFERRALS (key ones):

Condition

Deferral Period

Whole blood donation (AABB)

8 weeks (56 days)

Whole blood donation (Philippines, 450 mL)

12 weeks (3 months)

Double RBC apheresis

16 weeks

Infrequent plasmapheresis

4 weeks

Double/triple plateletpheresis

1 week

Post-partum (AABB)

6 weeks

Post-partum (Philippines)

1 year after delivery OR 3 months after weaning

Malaria (confirmed)

3 years after recovery, asymptomatic

Travel to endemic area (>24hr, <5yrs ago)

1 year

Travel to Iraq (leishmaniasis)

1 year

Syphilis/gonorrhea, high-risk sexual contact

12 months

Tattoo (unregulated)

12 months

Major surgery/transfusion/transplant

12 months

Incarceration >72 hours

12 months

VACCINE DEFERRAL (Sir's tip):

  • Single L vaccines (Measles/Rubeola, Mumps, BCG) = 2 weeks

  • Double L vaccines (Rubella, Varicella/Chickenpox) = 4 weeks

  • Smallpox = 8 weeks

  • Hepatitis B immune globulin = 12 months

  • Immunoglobulins = 1 year

  • Inactivated/killed/synthetic vaccines = NO DEFERRAL (Flu, Hepatitis A, DPT, Anthrax, Cholera, Lyme, etc.)

MEDICATION DEFERRAL:

Drug

Deferral

Tegison/Etretinate

Permanent

Growth hormone (pituitary) / Bovine insulin

Permanent

Soriatane/Acitretin (psoriasis)

3 years

Experimental/unlicensed vaccine

1 year

Accutane/Isotretinoin, Finasteride

1 month

Avodart/Dutasteride (BPH)

6 months

BLOOD COLLECTION:

  • Needle gauge: 16–17

  • Donation time: 8–12 minutes (>15 minutes = unsuitable for platelets, FFP, cryo)

  • Disinfectants: Iodophor and 10% povidone-iodine

  • Most common contaminant during collection: Staphylococcus aureus / S. saprophyticus (skin flora)

  • Post-donation rest: 15–20 minutes

APHERESIS TYPES:

  • Plasmapheresis = plasma collected

  • Plateletpheresis = platelets collected

  • Erythrocytapheresis = RBCs collected

  • Leukapheresis = leukocytes/granulocytes collected (only effective method for WBC collection)

  • Sedimenting agent for leukapheresis: Hydroxyethyl starch (HES) + corticosteroids

INFECTIOUS DISEASE TESTING:

Disease

Screening

Confirmatory

HIV

EIA, ChLIA, NAT

Western Blot; Philippines: rHIVda

Hepatitis B

HBsAg, Anti-HBc, HBV DNA

—

Hepatitis C

Anti-HCV, NAT

—

HTLV I/II

EIA, ChLIA

Western Blot, IFA

Syphilis

RPR, VDRL

FTA-ABS

LESSON 12: BLOOD COMPONENT PREPARATION & THERAPY

RBC STORAGE LESION (what DECREASES during storage):

  • Glucose, ATP, pH, 2,3-DPG, viable cells

  • Left shift of O2 dissociation curve → less O2 delivery

What INCREASES: Lactic acid, plasma K+, plasma hemoglobin

ANTICOAGULANT-PRESERVATIVES:

Solution

Shelf Life

ACD-A

21 days

CPD

21 days

CP2D

21 days

CPDA-1

35 days

With AS (AS-1, AS-3, AS-5, AS-7)

42 days

COMPONENT SHELF LIVES:

Component

Storage Temp

Shelf Life

Whole Blood

1–6°C

21–35 days

Packed RBCs

1–6°C

21–42 days (depends on anticoagulant)

Irradiated RBCs

1–6°C

28 days from irradiation (or original expiry, whichever is EARLIER)

Washed RBCs

1–6°C

24 hours (open); original expiry (closed)

Frozen RBCs

≤-65°C

10 years

Deglycerolized RBCs

1–6°C

24 hours (open); 14 days (closed)

Platelets (whole blood/apheresis)

20–24°C, continuous agitation

5–7 days

Pooled Platelets

20–24°C

4 hours

Granulocytes

20–24°C, NO agitation

24 hours

FFP

<-18°C

1 year; <-65°C = 7 years

FFP (thawed)

1–6°C

24 hours

Cryoprecipitate (frozen)

≤-18°C

1 year

Cryoprecipitate (thawed)

20–24°C

6 hours (closed); 4 hours (open)

Liquid Plasma

1–6°C

5 days after WB expiry

SPECIAL RBC COMPONENTS:

Component

Purpose

Key Fact

Irradiated RBCs

Prevent TA-GVHD

25 Gy; RED indicator = insufficient irradiation

Leukoreduced RBCs

Prevent FNHTR, CMV, HLA alloimmunization

<5×10⁶ WBCs; ≥85% RBC recovery

Washed RBCs

Severe allergic reactions, IgA deficiency

1–2L saline; removes plasma proteins

Frozen RBCs

Rare blood phenotypes

Glycerol 40%; 10-year storage

Deglycerolized

Rare blood types, IgA deficiency

Washed with decreasing osmolarity NaCl

PLATELET KEY FACTS:

  • Visual swirling = VIABLE platelets (discoid form)

  • No swirling = pH dropped below 6.2 = DISCARD

  • pH must be ≥6.2

  • Most prone to bacterial contamination (stored at RT)

  • Platelet refractoriness = CCI ≤5,000–7,500 (caused by HLA alloimmunization)

CCI FORMULA:
CCI = (Platelet Increment/µL × BSA m²) ÷ Platelets transfused (×10¹¹)

  • ≤5,000 = Refractory; ≥5,000 = Effective

CRYOPRECIPITATE CONTAINS: Fibrinogen, Factor VIII, vWF, Factor XIII

  • Minimum: 150 mg fibrinogen; 80 IU Factor VIII

  • Indicated for: Hemophilia A (2nd line), von Willebrand disease, hypofibrinogenemia, Factor XIII deficiency

FFP CONTAINS: ALL clotting factors including labile Factors V and VIII

COMPONENT PREPARATION SPINS:

  • pRBC = 1 light spin

  • PRP (Platelet Rich Plasma) = 1 light spin

  • Platelet Concentrate = 1 light spin + 1 hard spin

  • Cryoprecipitate = 1 light spin + 2 hard spins

BACTERIAL CONTAMINANTS:

Component

Bacteria

Platelets

S. aureus, S. saprophyticus (skin flora)

RBCs

Yersinia enterocolitica, Pseudomonas aeruginosa

IRRADIATION INDICATOR:

  • RED = insufficient (re-irradiate)

  • BLACK = adequate

LESSON 13: ADVERSE EFFECTS OF BLOOD TRANSFUSION

FIRST ACTION IN TRANSFUSION REACTION: STOP THE TRANSFUSION

POST-TRANSFUSION WORKUP:

  1. Clerical check

  2. Hemolysis check

  3. Direct Antiglobulin Test (DAT)

  4. ABO testing

ACUTE vs DELAYED HEMOLYTIC REACTIONS:

Feature

AHTR (Acute)

DHTR (Delayed)

Onset

<24 hours

>24 hours (days to weeks)

Antibody type

IgM (ABO)

IgG (Rh, Kidd, Kell, Duffy)

Hemolysis type

Intravascular

Extravascular (spleen)

Hemoglobinemia/uria

YES

NO

Bilirubin elevated

NO

YES (jaundice)

DAT

Positive

Positive

LDH

Elevated (High)

Mildly elevated

Spherocytes

NO

YES

Major complication

DIC, acute renal failure, shock

Mild anemia

ACUTE IMMUNE-MEDIATED REACTIONS:

Reaction

Cause

Key Distinguishing Feature

AHTR

ABO incompatibility (IgM)

Hemoglobinemia, hemoglobinuria, back pain, DIC

FNHTR

Cytokines from WBCs

Fever ≥1°C, NEGATIVE DAT, no hemolysis

Allergic

IgE vs plasma allergens

Urticaria, pruritus (mild); PAUSE then resume

Anaphylactic

Anti-IgA (IgA deficiency)

Severe systemic; STOP; give epinephrine

TRALI

Donor anti-HLA/anti-HNA (2-hit model)

Noncardiogenic pulmonary edema, hypotension, fever

FNHTR: Temperature rise >1°C; most common reaction type; treat with acetaminophen; PREVENT with leukoreduction

ALLERGIC vs ANAPHYLACTIC:

  • Allergic = pause transfusion, antihistamine (diphenhydramine), may resume

  • Anaphylactic = STOP immediately, epinephrine; IgA deficiency patients

TRALI - TWO-HIT MODEL:

  • Hit 1 (patient): Sepsis, trauma, surgery, massive transfusion → primes neutrophils

  • Hit 2 (donor): Anti-HLA or Anti-HNA antibodies (especially multiparous women) → activates primed neutrophils

  • Result: Noncardiogenic pulmonary edema, hypotension, fever

  • Prevention: Use male donors; avoid multiparous female donors; wash blood products

TACO vs TRALI:

Feature

TACO

TRALI

Edema type

Cardiogenic

Noncardiogenic

Blood pressure

HYPERTENSIVE

HYPOTENSIVE

Fever

Absent

Present

BNP

Elevated

Normal

Mechanism

Volume overload

Neutrophil activation

TA-GVHD:

  • Donor T-lymphocytes attack immunocompromised recipient

  • Requires: HLA differences + viable T-lymphocytes + immunocompromised recipient

  • Almost universally fatal within 1–3 weeks

  • Prevention: IRRADIATION (25 Gy)

POST-TRANSFUSION PURPURA (PTP):

  • Platelet count drops to <10,000/µL, 5–12 days after transfusion

  • Cause: Anti-HPA-1a antibodies destroy BOTH donor AND patient platelets

  • Treatment: Leukoreduced components, washed RBCs, HPA-matched platelets

NON-IMMUNE MEDIATED REACTIONS:

Reaction

Cause

Key Feature

Non-immune hemolysis

Thermal (>50°C or <0°C), osmotic, mechanical

No antibodies; physical damage

Bacterial sepsis (TTBI)

Contaminated blood bag

High fever, shock, DIC; Gram stain + culture to confirm

TACO

Volume/rate overload

Cardiogenic pulmonary edema; hypertension

Hemosiderosis

Chronic transfusions

Iron overload; treat with deferiprone/deferoxamine

Citrate toxicity

Excess citrate chelating Ca²⁺/Mg²⁺

Hypocalcemia, hypomagnesemia; massive/rapid transfusion

ONLY 0.9% NORMAL SALINE approved for mixing with blood components

OSMOTIC DAMAGE: Never mix blood with D5W, 0.45% saline, or Ringer's lactate

HEMOSIDEROSIS treatment: Iron chelation therapy (Deferiprone or Deferoxamine)

CITRATE TOXICITY treatment: IV calcium chloride/gluconate; slow infusion rate

QUICK MNEMONICS

  • MOMOYS = 2-week vaccine deferral (Measles, Others, Mumps, Oral polio, Yellow fever, Smallpox [BCG also included])

  • Double L = double weeks (Rubellla = 4 weeks, Varicella = 4 weeks)

  • "all Ergic = IgE; anAphylactic = IgA"

  • TACO = Hypertension; TRALI = Hypotension

  • AHTR = Intravascular = IgM = ABO; DHTR = Extravascular = IgG = Rh/Kidd/Kell/Duffy

  • Kidd antibody = most notorious (titer drops easily; still give antigen-negative units if history of antibody)