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Transfusion 231 > Week 1 > Flashcards

Week 1 Flashcards

1
Q

What is immunohematology

A

study of blood as it relates to AG and AB

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2
Q

Why are screen performed on patients

A

to detect allo AB in patients plasma sample so you know what crossmatching procedure needs to be done to cross match units for the pt

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3
Q

What are the blood products that transfusion department issues

A

packed red cells, fresh frozen plasma, platelets, fibrinogen
concentrates, and any factor concentrates main goal is to give ABO compatible blood but sometimes you cant

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4
Q

What is the primary immune response

A

Humoral with B lymphs
-after a transfusion B lymphs recognize AG on red cells through B receptors and AB production may not start

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5
Q

What is humoral immunity

A

-fluid parts of the immune system like AB and complement components
-mediated by B cells
-initiate AB production
-AB produced by B cells

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6
Q

What is cellular immunity

A

Mediated by T Cells and Lymphokines (activates/deactivates cells)

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7
Q

What is the function of the AB

A

to bind foreign molecules -AG
-AG are found on the surface of the foreign cells
-one AB binds with one AG (epitope)
-study of AB -AG reactions = serology

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8
Q

What are AG

A

-what the body thinks is foreign and causes the body to produce AB
-large molecular weight proteins and polysaccs
-on surface of cell membrane or integral part of cell membrane
-determined by inheritance

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9
Q

What is Antigenicity

A

Ability of antigen to be recognized by antibodies

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10
Q

What does antigenicity depend on

A

Chemical Composition and Complexity – Proteins are the best immunogens (produces immune response). Can be protein, glycolipid, glycoprotein, lipoprotein, nucleic acid
Degree of Foreignness – Greater the difference has a greater chance of producing a reaction
Molecular Weight – Larger molecular weight are better immunogens
Dosage and Antigen Density – Number of cells and amount of antigen introduced
Route of Administration – Intramuscular or intravenous

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11
Q

What are AG composed of

A

Protein
glycolipid - (carb and lipids)
glycoprotein (carb and protein)
lipoprotein
nucleic acid

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12
Q

What are AG composed of

A

Non- ABO like Rh/Kell =proteins
ABO = carbs

Lewis AG are soluble substances in plasma so they are absorbed into the membrane

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13
Q

Difference between protein and carb AG

A

Protein Antigen – Integrated into the RBC membrane, developed at birth
Gene codes directly for protein (Rh, Kell, Kx, Deigo)

2.Carbohydrate Antigen – found on Surface of the RBC membrane
Gene codes for an enzyme like glycosyltransferase which adds sugar to a precursor on RBC membrane to form AG
-enzyme production starts early but AG not developed at birth
-ABC AG similar structure to pollen or bacterial membrane

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14
Q

Why Testing Cord/Baby Samples May Have Weak Reactions

A

Antigens are NOT well developed – Despite enzyme production beginning early

Wharton’s Jelly (substance surrounding umbilical cord) can interfere with testing

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15
Q

What are AB

A

proteins produced by B lymphs because there was stimulation by AG
-Immunoglobulins
-Gamma globlin region in SPE
-isotypes/classes depend on Heavy chain
-specificity based on variable regions
-IgG, IgM, IgA, IgD, and IgE
Allo antibodies are antibodies to a foreign antigen of the same species
* Autoantibodies are antibodies to an antigen one possesses

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16
Q

What sites does the AG bind

A

AB binds the specific antigen
* Fc Portion determines the antibody function and contains the complement
binding region and the cell activation region

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16
Q

What is the structure of the AB

A

2 Heavy chains + 2 Light chains – Can either be in constant or variable region

Fab region + Fc region – Fab (Binds antigen), Fc (determines antibody function, contains complement binding region and cell activation region)

-IgG – 2 binding sites; temperature: most optimal at 37 deg C- can cross placenta HDFN may need enhancement techniques
-IgM – 10 binding sites; most optimal at 4 deg; pentamer
Can cause immediate transfusion reaction

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17
Q

Types Of Exposure To Antigens

A

Naturally Occurring (Non Red Cell Immune) – After birth or throughout life; Exposure to antigen similar to RBC antigen
Red Cell Immune – Through transfusion or pregnancy; Exposure to antigen on foreign RBC

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18
Q

how does a B cell respond to an AG

A

-AG stimulates lymphs with best fit receptor . Lymphs signalled to produce B cell clone >plasma cells > ONE ab with ONE specificity

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19
Q

Types of Antibody Production Response

A

Primary Antibody Response – Formation of IgM, followed by IgG and memory B cells
-peak response is smaller with low affinity 5-10 days
–more memory cells produced and response sustained for longer

Secondary Antibody Response – more production of IgG and less IgM; Faster and increased potential for RBC destruction > Leads to plateau
-in 1-3days
-higher average affinity

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20
Q

What does the likelihood of an immune response depend on

A

-immunogenicity of the AG - able to stimulate an immune response
-pt own immune systems sensitivity- some people develop multiple AB from ONE transfusion
-iron supplements are given to pt to raise hgb instead of transfusing

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21
Q

Types of AG exposure

A

-AB formation can occur without apparent AG exposure- naturally occurring (ABO groups)
- can occur after birth “Naturally occuring”
-AG on foreign RBC
-through transfusion or pregnancy = Red Cell Immune

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22
Q

What happens after AB production

A

AB binds to specific AG
-AG (donor RBC) are targeted for destruction
-IgM INTRAvas clearance
-IgG EXTRAvas clearance

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23
Q

What is the complement pathway

A

-activated proteins in immunological reactions
-activation occurs as cascade
-proteins circulate in inactive state but activate and convert into active enzymes

24
Q

Types of complement pathways

A

Classical Pathway – Activated by presence of antibody bound to an antigen

Alternative Pathway – Activated by bacteria, viruses, foreign constitutes

25
Q

Main Components of Complement Cascade

A

-C3 convertase (C4b2a)-
-C5 convertase (C4b2a3b)
-C3a: anaphylatoxin, eosinophil chemotaxin
-C3b: mediates phagocytosis, opsonin
-C5a: anaphylatoxin & chemotaxin for leukocytes and monocytes
-C5b-9: Membrane Attack Complex (MAC - C4b2a3b)

26
Q

What is the classical pathway composed of

A

C1 complex
C2b and C4b
C3 convertase

27
Q

What is the alternative pathway composed of

A

C3 hydrolysis
C3b and C3a
-C3b cleaves C5 into C5a/b and then C5B- C9 come together from MAC attack

28
Q

Detection of Complement –

A

Anti-C3d tests for C3d (breakdown of C3b)

because C3b and 4b are left behind on the RBC membrane and degraded into C3D/4D
-AB can dissociate while complement portions stay on the red cell
-Pos reactions are helpful in transfusion

29
Q

Where does INTRAvas occur , what does it involve, lead to

A

Occurs in the veins, arteries, capillaries
-Involves IgM to detect ABO
-Complement cascade leads to MAC formation, then hemolysis

30
Q

Where does EXTRAvas occur , what does it involve, lead to

A

-Occurs in the spleen and liver
-Involves IgG to detect Rh
-Complement cascade stops at C3b (no MAC forms). Splenic macrophages detect Fc of IgG

31
Q

Serology – Antigen and Antibody Reactions

SOURCE of AG
source of AB
detection

A

Source of antigen – Patient

Source of antibody – Donor’s plasma or commercially prepped antisera

Detection of reaction – Hemolysis (these are rejected) or Agglutination

32
Q

What tubes are preferred for blood bank

A

lav or pink top -(Lavender Tubes) EDTA – Chelates Ca2+ and prevents the vitro activation of the complement pathway; Preferred for RBC and Plasma

33
Q

Types of Antigen and Antibody Reactions
Sensitization

A

Antibody binds to Antigen epitope&raquo_space; Invisible
Dependent on temperature, incubation time, pH, ionic strength

34
Q

Types of Antigen and Antibody Reactions
Precipitation –

A

Precipitation – Soluble antigens&raquo_space; Visible

35
Q

Types of Antigen and Antibody Reactions
Agglutination –

A

Agglutination – Lattice formation occurs (involves IgM pentamer)&raquo_space; Visible
Dependent on zeta potential which affects crosslinking, zone of equivalence, centrifugation
-to decrease zeta potential isotonic saline, albumin, LISS, PEG, AHG and enzymes in testing.

36
Q

Types of Antigen and Antibody Reactions
Hemolysis

A

Hemolysis – Complement is activated&raquo_space; Visible

37
Q

What are potentiators

A

Used to decrease zeta potential aka enhances antibody reactions (such as albumin, LISS, PEG, AHG, enzymes in testing)

37
Q

Factor Affecting Antigen and Antibody Reactions

Type of immunoglobin

A

-immunoglobin IgG or IgM
-RBC Solution – Suspension material, pH
-Concentration of antigen and antibody, Cell to serum ratio
Prozone – Antibody excess
Equivalence Zone – Proper proportion of antibody and antigen
Post Zone – Antigen excess
-temps
-time = increased incubation can cause AB to dissociate
-size, shape or charge

38
Q

Affinity

Factor Affecting Antigen and Antibody Reactions

A

strength of the binding between a single antibody and an epitope of an
antigen.

39
Q

Avidity:
Factor Affecting Antigen and Antibody Reactions

A

overall strength of the reaction
depends on the affinity of the antibody, valency and noncovalent
attractive forces

40
Q

What is zeta potential

A

Net negative charge around the RBC repelling each other

Too much zeta potential = No agglutination

IgM is a pentamer and can cause agglutination because it can bridge the gap; IgG is too small

41
Q

How to decrease zeta potential?

A

Increase protein to reduce the positive and negative ions that prevent antibodies from binding with antigens

42
Q

how can enzymes enhance AB reactions

A

Can either enhance antibody reactions OR depress reactivity of other antibodies (such as Ficin, Papain, Trypsin, Bromelin)

43
Q

how can we make AB-AG reactions more visible

A

goal is to enhanceAB uptake and make reaction visible
-ALB reduces repulsion between cells making the agglutination more visible to the eye
-PEG increases detection sensitivity
LISS – Used for screening cells; Speeds up agglutination, economical, provides good sensitivity but can enhance cold autoantibodies

44
Q

During agglutination how is testing prepped

A

Packed RBCs are DILUTED
-3-5% suspension is needed
-Plasma and Antisera are NOT dilutedW

45
Q

What is the Immediate Spin
(Direct Antiglobulin Test - DAT)
technique entail

A

Detects in vivo antibody or complement that’s attached to RBC membrane
-Detects IgM (ABO/Rh)
-add plasma w/3-5 % RBC
-reagent is monoclonal
-AB from Hybridoma

46
Q

What is the Antiglobulin Test
(Indirect Antiglobulin Test - IAT)
technique entail

A

Detects in vitro antibody or complement that’s attached to RBC membrane
-Detects IgG
-add plasma w/ 3-5% red cells
-incubate, wash and add AHG, centrifuge and read
-needs to be incubated at for sensitization to occur

47
Q

What is the Antihuman Globulin (AHG) Reagent

A

helps with the detection of AG and AB reaction; contains AB to target human AB

Polyspecific AHG Reagents – Anti-IgG and Anti-C3d
Monospecific AHG Reagents – ONLY Anti-IgG or ONLY Anti-C3d

48
Q

How to Perform DAT (IS)

A

Mix plasma (antisera; antibody) with 3-5% RBCs (antigen). Centrifuge.

49
Q

How to Perform IAT

A

IS Phase – Mix 2 drops plasma (antisera; antibody) with 1 drop 3-5% RBCs (antigen). Centrifuge. Record at IS and RT. Sensitization may have occurred here.

Incubate at 37C – Centrifuge and record at 37C.

Wash with saline 3 times – Removes free antibody from the mixture

Add AHG – Agglutination will occur with sensitization occurred. Centrifuge. Read at AHG

***If negative – Add Coombs Control Cells (CCC)/Check Cells (RBCs coated with sensitized IgG antibodies; will react with AHG > produce Positive Result)

50
Q

What is the Antiglobulin Reagent

A

-human IgG is injected into rabbit to produce human AB
-uses an AB against a human to help with AB-AG detection
-the human AB is the AG
-sometimes the AG-AB dont have bridging to produce agglutination so the AHG gives the complex a bridge and helps to form a lattice making the reaction visible

51
Q

When will we see a positive IAT test

A

-if AB is in the plasma and sensitization occurs - AB attaches to RBC+AG
-Add AHG = agglutination IF there is sensitization = POSITIVE

52
Q

When will you see a true negative IAT

A

NO AB in the plasma
-add AB
-no agglutination
-if reaction is negative add CCC which binds with AHG
-CCC needs to be added to validate the test

53
Q

What happens if there was AB in the plasma but washing did not occur properly

A

-unbound AB binds with AHG and neutralizes = no agglutination and free AB not bound to RBC are in the test mixture (prozone phase)
-false neg

54
Q

Coombs Control Cells CCC

A

-controls with AB on RBC
-helps tell the difference between true neg and false neg
-addtion of AHG if the reaction is negative under the mic, CCC is added to make sure

55
Q

No Antibody is detected and
negative results under the
microscope

A

-Add CCC
-agglutination occurs
-Reaction with CCC and AHG
Confirms a TRUE Negative IAT

56
Q

No Reaction after CCC is added
Indicates a false Negative IAT.

A

AHG was Neutralized
Must Repeat Testing

Transfusion 231 (10 decks)

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