blood bank rotation exam B, EXAM C bloodbank, exam D blood bank

definition of a titer

the minimum volume of a solution needed to reach the endpoint in a titration

The titer of a immunologic reaction is:

The highest —-->

Serial dilutions are made of the antibody containing plasma and tested against selected red blood cells to determine the highest dilution causing a positive reaction.

The result or titer is expressed as the reciprocal of the highest plasma dilution reacting 1+.

Enzymes

1. Enzymes are proteins that increase the rate of a chemical reaction.

2. In transfusion medicine, enzymes are used to alter red blood cell antigens.

3. The most commonly used are ficin and papain.

4. Less commonly used enzymes are bromelin, pronase, trypsin, and chymotrypsin.

5. Some red blood cells are destroyed by enzymes, while the reactivity of other antigens is enhanced.

Enzyme treatment can be performed in two ways:

The one-step technique: Enzyme is added as an enhancement to the tube containing test plasma and red blood cells and carried through the IAT phase of testing.

The two-step technique: Requires treatment of the red blood cells with enzymes and washing prior to the addition of patient plasma. This method is often preferred because the enzymes in the one-step technique are added directly to the patient's plasma and any antibodies present may be damaged.

Neutralization

The inactivation of an antibody by combining it with a soluble form of the corresponding antigen.

P1 antigen

neutralization: Hydatid cyst fluid, Pigeon eggs

Rare hemolytic IgG component

Associated with PCH, parasitic infection

I antigen

Neutralization: Breast milk

Tested with cord blood

Stimulated by M. pneumoniae (I-like Ag)

Fy a antigen %

66%

Fy b antigen %

83%

K antigen %

9%

k antigen %

99.8%

S antigen %

55%

s antigen %

89%

Jka antigen %

77%

Jkb antigen %

72%

Kpa is a low-frequency antigen

only 2%

Kpb is a high-frequency antigen

(99.9%)

Js a antigen %

0.1% in white

20% in black

Jsb antigen %

both white and black 99%

Cold Hemagglutinin Disease

→ anti-I, anti-i, anti-Pr

Acute: secondary to lymphoproliferative diseases or M. pneumoniae

Chronic: elderly patients, may result in Raynaud's and hemoglobinuria

Lab findings: reticulocytosis, positive DAT due to complement (IgM), negative eluate

Agglutinated RBCs, polychromasia, anisocytosis, poikilocytosis

Rabbit Erythrocyte Stroma Test (REST)

Used for patients that were recently transfused

Anti-I prefers the I antigens on rabbit cells → Antibody is absorbed out

CAUTION: may absorb clinically significant antibodies (D, E, Vel) and IgM

Can be used to adsorb certain antibodies.

Removes cold autoantibodies, anti-B, and antibodies in the P blood group system (anti-P1).

Cold autoadsorption

Cannot be performed on recently transfused patients

Procedure: keep EDTA sample warm, separate serum, wash RBCs with warm saline, add aliquot of serum to RBCs, incubate for 1 hour at 4°C, harvest serum and test screen cells

All negative → no alloantibody

Positive and negative → alloantibody present, run panel

All positive → unsuccessful, repeat with absorbed sample

Paroxysmal Cold Hemoglobinuria

→ anti-P, Donath-Landsteiner Antibody

historically associated with syphilis infections

Usually an acute transient condition secondary to viral infection in children

Biphasic hemolysin causes hemolysis after exposure to cold

Lab testing: DAT + IgG autoantibody causes complement → C3d is +

Eluate = since only complement is coating cells

Warm autoimmune hemolytic anemia

Most common type of AIHA and most difficult problem encountered in transfusion service

Intravascular immune RBC destruction occurs

DAT Results:

IgG and C3d - 67%, IgG alone - 20%, C3d alone - 13%

In rare cases, = DAT due to RBCs coated with IgA or IgM

Serum Testing

May have little free autoantibody, all on RBCs instead

50% of WAIHA have autoantibodies reactive with all cells tested (nonspecific)

CAUTION: alloantibody may also be present and hidden by autoantibodies

Eluate + (except if complement only or drug-induced)

Donath-Landsteiner Test

Two tubes of blood are drawn

One tube placed at 37°C for 1 hr, the other placed at 4°C for 30 min then 37°C for 30 min

Both tubes centrifuged and examined for hemolysis

Hemolysis in cold incubated tube, none in warm tube

= Donath-Landsteiner Ab

Effects on the Infant on HDFN

Severe cases: heart failure, profound anemia, hepatosplenomegaly

Hydrops fetalis - high output cardiac failure with generalized edema

Kernicterus - indirect bilirubin levels > 20 mg/dL results in mental retardation or death

-unbound unconjugated bilirubin crosses blood-brain barrier → neurotoxicity

-occurs at lower levels in presence of acidosis, prematurity, hypoalbuminemia, and some drugs

Less severe: accelerated RBC destruction, generation of large quantities of bilirubin

deficiency in uridine diphosphoglucuronic acid in immature fetal liver

Hemolytic Disease of the Newborn

Background

Shortened lifespan of fetal or neonatal RBCs by action of maternal IgG antibodies attaching to the corresponding antigens of paternal origin on baby's RBCs.

Necessary conditions for HDN to develop:

1. Mother must lack antigen present on fetal RBCs

2. Fetal Ag must be well developed in utero

3. Mother is exposed to fetal Ag → , previous pregnancy, exposure to RBCs

-Transfusion, amniocentesis, miscarriage, abortion, rupture to ectopic pregnancy, etc.

-Biggest risk of exposure during birth; HDN can occur with <0.1mL

4. Mother produces IgG Ab capable of crossing the placenta

Detection of Fetomaternal Hemorrhage (FMH)

Weak D test

Fetal D+ coated with reagent anti-D, agglutination evaluated microscopically

Very insensitive and requires skill; not used much anymore

Rosette Screening Test → qualitative

Maternal RBCs incubated with anti-D which coats D Ag of any fetal RBCs present

Anti-D is washed off, D+ indicator RBCs bind to second Fab of anti-D coating cells

Rosettes form around the cells

NOTE: if baby types weak D+, perform Kleihauer Betke instead of rosette

Kleihauer Betke Acid Elution

Hemoglobin F is resistant to acid elution whereas adult hemoglobin is not

Procedure: smear stained with acid buffer → adult Hgb eluted out → stained with hematoxylin and erythrosin B → count the number of fetal cells in 2000 adult cells

Adult RBCs appear as ghost cells while fetal RBCs are bright pink and refractile

% of fetal RBCs (50) / 30 → round to whole number and add 1 vial for safety

If test is =, give 1 vial RgIg as a precaution

CAUTION: false positives may occur if patient has HbgF due to certain diseases:

Sickle cell anemia, thalassemia, aplastic anemia, and other hemoglobinopathies

Flow Cytometry

Uses anti-hemoglobin F to bind fetal RBCs followed by incubation with labeled anti-Ab

What is elution?

free antibody that is bound to a red blood cell → the term refers to removing (or "dissociating") an antibody that is attached to the surface of a red blood cell.

Uses of elution include:

Identify one or more antibodies attached to the red blood cell membrane.

Can be combined with adsorption techniques to remove specific antibodies.

Adsorption and elution techniques can be used to detect a weakly reactive antigen, such as a weak ABO subgroup.

Methods that are best at releasing cold-reactive antibodies or antibodies that have a broad temperature range of reactivity include:

Lansteiner and Miller heat elution

Weiner's freeze-thaw method

Lui's freeze-thaw method and sonication

Methods better suited for recovering warm-reactive alloantibodies or auto antibodies include

use of organic solvents such as ether, xylene, methylene chloride, and chloroform or acids such as digitonin, citric, or glycine.

Preparation of a successful eluate can be influenced by several factors:

Improper technique

Inadequate stroma removal

Insufficient washing of the sensitized red blood cells before eluting the antibody

Incorrect method of transferring washed red blood cells

Incorrect procedure used during the wash phase

Use of saline to prepare eluates

what is Adsorption

A technique used to remove an antibody or antibodies from plasma → adsorption refers primarily to the adherence of an antibody onto the surface of a red blood cell, described briefly below and in more detail in the linked definitions.

Done most often to remove autoantibodies from a patient's specimen in order to see if other antibodies are present in the plasma.

Autologous adsorption

Preferred method to remove autoantibodies.

Cannot be used if the patient has been transfused within the previous 3 months because the patient's specimen will also contain donor red blood cells.

Allogeneic adsorption

used when a patient with an autoantibody has been transfused.

When choosing red blood cells for adsorption, the antigens that stimulate the most clinically significant antibodies are considered.

Titration is

A technique used to measure the strength (concentration) of an antibody.

The sum of these scores reflects the total binding strength of the antigen and antibody molecules (known as titration score or avidity).

Two antibodies can have the same titer but different titration scores, demonstrating the same strength but differing avidity.

Titration is most commonly used for determining antibody activity and the potential severity of HDFN in alloimmunized pregnant women.

A titration can distinguish between a characteristic high-titer, low-avidity antibody (HTLA) and a weakly reactive antibody with a low titer.

High titer = need barely any to cause a reaction

If the autocontrol or DAT is negative and the patient has been transfused or pregnant--> mean

reactions are likely due to alloantibodies

because Both combinations of multiple antibodies and a single antibody to a high-incidence antigen may cause this pattern of reactivity.

If the autocontrol or DAT is positive and the patient has not been transfused recently---> mean

results are likely due to an autoantibody, either warm reactive, cold reactive, or both.

if the autocontrol or DAT is positive and the patient has been transfused recently,---> mean

results may indicate either an autoantibody or a delayed serologic transfusion reaction.

False-positive panreactivity may occur with

rouleaux or antibodies to reagents.

The typical reaction pattern when a patient has developed an antibody to a high-incidence antigen is

negative autocontrol or DAT, but almost all screen, panel, and donor red blood cells reacting positive.

how to confirm the specificity of the antibody.

Testing with rare selected cells that lack the antigen is the best way

Warm Autoantibodies

known as autoimmune hemolytic anemia.

Warm autoantibodies react with the patient's own red blood cells at body temperature. The typical pattern of reactivity is a positive autocontrol and DAT; and all screen, panel, and donor red blood cells reacting positive.

An antibody that causes all cells tested to agglutinate is called a panagglutination.

what is panagglutinin aka

warm autoantibody

, in Warm Autoantibodies, If the patient has not been transfused in the last 3 months

an autoadsorption can be performed and the adsorbed plasma tested for alloantibodies.

Warm Autoantibodies, If the patient has been transfused

allogenic adsorptions may be used to identify any underlying alloantibodies.

Cold agglutinins

Antibodies that optimally react at temperatures between 39.2° F and 77° F (4° C and 25° C) and can be autoantibodies or alloantibodies.

They are differentiated by running an autocontrol or DAT.

Cold Autoantibodies

Cold autoantibodies include the specificities to I, i, IH, and Pr antigens; autoanti-I is the most common.

Although the autoantibodies can be distinguished using cord and adult red blood cells, identification is generally not necessary.

If cold autoantibodies may be masking alloantibodies, the autoantibodies can be removed from plasma by autoadsorption using a low temperature or by adsorption with rabbit erythrocyte stroma.

Cold Alloantibodies

Include specificities such as anti-M, anti-N, anti-P1, anti-P, anti-Pk, anti-Lea, anti- Leb, and anti-Lua.

These antibodies rarely cause in vivo hemolysis.

3 simple techniques for avoiding detection of cold alloantibody when using a tube method are (1) eliminating the immediate spin reading; (2) eliminating microscopic readings; and (3) testing with anti-IgG instead of polyspecific antihuman globulin.

Prewarm Technique

used to detect the presence of clinically significant antibodies in the presence of cold-reactive antibodies.

Plasma and cells are warmed separately at 98.6° F (37° C) prior to testing in an attempt to avoid binding of cold antibodies.

If performing a tube technique, warm saline may be used for the wash step in the IAT

Prewarm is also used to resolve ABO discrepancies due to cold antibodies.

Delayed Hemolytic or Serologic Transfusion Reaction

caused by multiple alloantibodies may show the same pattern of reactivity as a warm autoantibody.

With a delayed hemolytic reaction the patient exhibits symptoms of hemolysis,

whereas in a delayed serologic reaction the DAT becomes positive but the patient does not exhibit symptoms of hemolysis.

Rouleaux

Caused by unusual properties in a patient's plasma that can aggregate red blood cells and mimic agglutination.

Rouleaux can occur as a result of various intravenous injections or an abnormal concentration of serum proteins

performing a saline replacement procedure.

Resolutions for eliminating the reaction include:

Antibiotics: Wash reagent red blood cells prior to testing

Sugars: Inhibit by adding glucose to test medium

Dyes: Wash patient red blood cells to remove patient plasma prior to testing

Bacteriostatic/antifungal reagents: Wash patient red blood cells to remove patient plasma prior to testing

Miscellaneous: Wash reagent red blood cells prior to testing

where is HLA molecules are expressed

on immature red blood cells that contain nuclei but often disappear as the erythrocytes mature.

Compatibility Testing aka

crossmatch.

Performed prior to release of red blood cells for transfusion

.Uses in vitro testing results to try to predict if patient/recipient plasma will react in vivo with donor RBCs if transfused.

Immediate spin testing is conducted to detect any ABO incompatibilities.

The incubation/enhancement and IAT test phases are designed to detect unexpected patient antibodies against corresponding antigens on donor cells.

Agglutination in any phase of testing is considered incompatible and is not released for transfusion.

Pretesting of Specimens from Neonates

The ABO group of the neonate is determined only by typing red blood cells with anti-A and anti-B antisera (forward blood grouping).

The major concern with antibody screen testing is detecting and identifying possible passive maternal red blood cell antibodies.

transfuse only group O red blood cells to neonates.

Specimen Retention how many days

7 days after transfusion.

Immediate Spin Crossmatch

Testing that occurs when donor red blood cells are physically tested against recipient/patient plasma is called a serologic crossmatch.

This type of testing occurs at room temperature and is designed to detect ABO incompatibilities.

As there is no incubation or IAT phase in the immediate spin crossmatch, most non-ABO antibodies will not be detected.

The IS crossmatch should only be used if the recipient appears not to have a clinically significant antibody.

Cold-reactive antibodies other than ABO may cause a positive reaction in the IS crossmatch. Repeating the test using either a prewarm technique or incubation phase may be done.

Drug-Induced Immune Hemolytic Anemias (DIIHA)

Detects antibodies that recognize drug bound to RBCs.

dRUG-TREATED CELL are prepared by incubating group O RBCs with a solution of the drug believed to be causing the anemia.

The most common causes are the cephalosporins.

DAT +, ELUTION =

Rh-Immune Globulin (RhIg)

Used to protect D= individuals exposed to D+ RBCs. IgG anti-D coats D+ RBCs causing destruction before they can be recognized, thereby preventing B cell activation and formation of memory cells.

Immunization occurs if fetal RBCs entering maternal circulation exceeds 30 mL of whole blood

Amniotic Fluid Analysis

Level of bilirubin in amniotic fluid correlates with hemolysis of fetal RBCs in utero

Amniocentesis

Perform at 28 weeks if previous child with HDN

Perform at 22 weeks if previous child was severely affected

Perform if maternal antibody increases before 34th week

Direct Coombs Test (DAT)

Detects in-vivo sensitization of RBCs with IgG or complement (Cd3)

Used in investigating AIHA, drug-induced hemolysis, HDFN, and alloimmune reactions to recent transfusion

Washed red cells from patient or donor are tested directly with AHG reagents

Does not require the incubation phase because the antigen-antibody complexes formed in vivo

Does not use LISS

Indirect Coombs Test

Detect in-vitro sensitization of RBCs

Used for detection of incomplete antibodies to potential donor RBCs (compatibility testing), antibody screening, determination of RBC phenotype, and for titration of incomplete antibodies

Serum or plasma is incubated with red cells, which are then washed to remove unbound globulins

RBCs are incubated at 37C to sensitized the cells with the IgG or complement (Cd3)

The use of LISS

what is complete antibody

· antibody that is able to inherently agglutinate red blood cells in physiological saline solution without the need of an AHG reagent. It is an IgM antibody with a large pentamer structure that allows the complete antibody to bridge the zeta potential gap between the RBCs

what is incomplete antibody

· antibody binds to its respective antigen on a RBC but will not cause an agglutination reaction without the addition of AHG reagent. The smaller, monomeric structure of the IgG antibody cannot bridge the ionic gap between the RBCs. Incomplete antibodies are IgG antibodies that usually react at 37C and are considered clinically significant antibodies.

· Phenotype Duffy: Fy (a-b-) disease

o Resistant to infections from Plasmodium knowlesi and Plasmodium vivax (malaria)

· Anti-i diseases

Infectious mononucleosis, Epstein-Barr virus, and Cytomegalovirus often have transient but potent anti-i.

o Associated with *Congenital dyserythropoietic anemia Type II*

Anti-P disease

o Echinococcus granulosis (parasitic infection) , facioliasis. (parasitic infections caused by tapeworms/flukes)

o Autoanti-P is associated with paroxysmal cold hemoglobinuria and is a cold-reactive IgG autoantibody that is described as a biphasic hemolysin

anti-kell diseases

o HTR, McLeod Syndrome (anti-K, -k)

§ Chronic granulomatous disease (CGD)

What is an antibody screen

o Method to detect clinically significant antibodies in both the blood donor and recipient as part of pretransfusion compatibility testing

Describe reagent red cells used in the antibody screen.

o RBC reagents are from group O that have been typed for most common & significant antigens

§ Group O cells used so anti-A & anti-B won't interfere with detection of unexpected Abs

o Packaged in sets of 2-3 cell suspensions with unique combination of antigens

§ Usually R1R1, R2R2, and rr

Should have at least one cell positive for each significant antibodies

What is zeta potential

o Net negative charge surrounding red blood cells

§ Ionic cloud of cations (positively charged ions) that are attracted to the zone of negative charges on the RBC membrane

§ Ionic cloud forces the cells apart

What does the use of albumin and enzymes do to the zeta potential

★ Albumin

o Increases of the dielectric constant → reduces the zeta potential

★ Enzymes

o Releases sialic acid from the RBC membrane → reduces the zeta potential

When are elutions of value? What is the acid in an acid eluate? How low is the pH? How is the pH returned to normal prior to testing?

Elution methods are used to free antibody from the cell surface to allow for identification

o Bound antibody may be released by changing the thermodynamics of an antigen-antibody reaction, neutralizing or reversing forces of attraction that hold antigen-antibody complexes together, or disturbing the structures of the antigen-antibody binding site

§ Eluate may be tested against an RBC panel to identify the antibody

§ Usually used after a positive DAT

A total elution

o Antibody is released and RBC antigens are destroyed

o Necessary when performing antibody identification

Partial elution

o Antibody is removed but RBC antigens remain intact

o Useful to prepare RBCs for phenotyping and to use in autoadsorption

Acid in acid elution

o HCl or glycine acid

o pH of 3

To return pH to normal

o Add a buffer

Why are cord cells used in antibody identification?

· Cord cells are used in antibody identification to detect cold agglutinins. Anti-I is a common agglutinin, most adults should have the I antigen present while cord cells have little to no I antigen at birth.

· Alloanti-D may be produced in D+ individuals with partial D phenotype, although not all persons who are D+ and produce what appears to be anti-D should be assumed to have epitope-deficient red cells. A D+ person who have a weakly reactive anti-LW may be indistinguishable on initial serologic testing from an individual with a partial D antigen who has made anti-D to missing epitopes. Cord cells on the other hand expresses anti-LW with equal reactivity regardless of the D-type and will differentiate anti-LW from an alloanti-D

13) Describe the reactivity of the Lewis antibodies.

· Lewis antibodies are naturally occurring antibodies and are not clinically significant. Lewis antibodies are IgM antibodies and will usually react at room temperature or colder.

· Lewis antibodies occur almost exclusively in the sera of Le(a=b=) individuals, usually without known red cell stimulus. Individuals with the Le(a=b+) red cell phenotype do not make anti-Le(a) because small amounts of unconverted Le(a) are present in their saliva and plasma. It is unusual to fine anti-Le(b) in the sera of Le(a+b=) individuals but both anti-Le(a) and anti-Le(b) can be found in sera of Le(a=b=) individuals.

· Lewis antibodies are often found in the sera of pregnant women who transiently demonstrate Le(a=b=) phenotype.

o These Lewis antibodies however are almost always IgM and will not cross the placenta

o Lewis antigens are poorly developed at birth and are not associated with HDFN

· Lewis antibodies may bind complement, and fresh serum that contains anti-Le(a) may hemolyze incompatible red cells in vitro

What is the purpose of testing an auto control with the antibody screen? If the auto control is positive, what does this indicate and what should be done next

The autologous control is the patient's RBCs tested against the patient's serum or plasma in the same manner as the antibody screen. A positive autologous control may indicate the presence of autoantibodies or antibodies to medications. If the patient has been recently transfused (in the past 3 months), the positive autologous control may be caused by alloantibody coating circulating donor RBCs. An elution can be performed to isolate the autoantibody that is coating the cell and then an antibody panel can be used to identify the actual antibody in the eluate.

What is a crossmatch

The crossmatch test has been traditionally meant the testing of the patient's serum with the donor RBCs, including an antiglobulin phase or an immediate spin phase to confirm ABO compatibility.

1. What is the difference between a major and a minor crossmatch?

Crossmatch is a general term used for the testing of an individual's serum or plasma against another individual's red blood cells to check and see if there is ABO or Rh incompatibility. There are two main types of crossmatch: the major crossmatch and the minor crossmatch.

Major crossmatches test donor RBCs against recipient serum or plasma, and are required anytime a transfused blood product contains 2 mL of RBCs. Minor crossmatches which are not required in routine testing are but can be used to assess compatibility between donor serum or plasma against recipient RBCs. The major crossmatch is a core component of pretransfusion testing and serves as the final check of ABO compatibility between a donor and a recipient.

What is the purpose of the minor crossmatch? Why has it been eliminated from the crossmatch

In contrast to the major crossmatch (recipient serum vs donor red blood cells), the minor crossmatch is used test opposite compatibility. The purpose of the minor crossmatch is to test donor's serum or plasma with the recipient's red blood cells. Minor crossmatches have been eliminated and rarely performed anymore due to two main reasons:

· First, transfused blood is usually screened for unexpected non-ABO antibodies already, so performing a minor crossmatch to make sure non-ABO antibody won't be causing a problem doesn't really make sense

· Second, since the volume of transfused plasma is generally small compared to the patient's blood volume, minor compatibilities will not really be that great of a concern

1. Why is clotted blood preferred for crossmatching and antibody studies? How long may a clot which has been refrigerated be safely used for crossmatching?

Clotted blood is preferred for crossmatching and antibody studies because clotted blood is free from any excess serum or plasma that can lead to false positive. A clot which has been refrigerated can be safely used for crossmatching for 72 hours.

1. What is rouleaux formation? How is it differentiated from true agglutination? What is the relationship between high-molecular-weight plasma expanders (such as Dextran) and rouleaux?

Rouleaux are aggregates of red cells that characteristically, adhere to one another on their flat surface, give a "stack of coins" appearance when viewed microscopically.

Rouleaux can be differentiated from true agglutination by using the saline replacement technique. In the saline replacement technique, serum and cells are incubated to allow antibody-antigen association but serum is then removed and saline is added as the resuspending medium. If agglutination is caused by rouleaux formation, the saline replacement technique will disperse the rouleaux. If it is true agglutination, the agglutination will not go away with the saline replacement technique.

Abnormal concentrations of proteins, altered, serum protein ratios, or the presence of high-molecular-weight plasma expanders such as Dextran can cause non-specific red cell aggregation or rouleaux. It may produce hazy reactions or false-positive results.

1. Why would a patient with multiple myeloma be difficult to crossmatch?

Patient with liver disease, multiple myeloma, or another condition associated with abnormal globulin levels often have rouleaux formation that results from the abnormal serum protein concentrations. It may be difficult to detect antibody-associated agglutination in a test system containing rouleaux-promoting serum.

1. What effect will cold agglutinins have on the crossmatch?

Cold agglutinins will not have any effect on the crossmatch at 37C or AHG phases. At the IS phase, cold agglutinins may cause false positive results but the crossmatch will still be considered compatible if results are negative at 37C and AHG testing

1. Why is hemolysis important? What must be present for hemolysis to occur?

Hemolysis is a general term for the destruction of RBCs. In blood banking, if hemolysis is present, it must be interpreted as a positive result. Hemolysis in routine blood banking tests indicates that incompatible antibodies are interacting with the RBCs leading to the destruction of the RBCs.

What tests comprise full compatibility testing?

1. ABO grouping and Rh typing (including weak D test), and tests intended to prevent disease transmission must performed on sample of both donor and patients, which take at the time of collection

2. screening test for unexpected antibodies to RBC antigen om samp-le both donor and patients with history of pregnancy or transfusion

3. identification of abs if detect in screening test

4. confirm ABO cell group on all units and Rh typing on units labeled- Rh negative

5. crossmatch as final check for ABO compatibility and check for any presence of abs.

Major crossmatch is

recipient serum and donor RBC

MINOR CROSSMATCH

Donor serum and recipient RBC

why minor crossmatch has been eliminated?

1. transfused blood is screened for unexpected non-ABO abs already, so perform minor crossmatch to make sure non-ABO abs won't cause problem that doesn't make sense

2. volume of transfused plasma is small compared to patient's blood volume, minor crossmatch won't really great of concern.

how rouleaux affect crossmatch phase and what temp? how to confirm rouleaux in crossmatch

multiple myeloma and macroglobulinemia cause rouleaux because the imbalance of normal ratio of albumin and gamma globulin. so, rouleaux affect all tests, include autocontrol.

rouleaux usually strongest after 37C incubation, but not persist through washing before AHG test.

use saline replacement is solved, because rouleaux will disperse in saline if it is true agglutination.

how to eliminate cold abs

absorb serum in the cold with autologous red cell can remove autoabs, allow to detect alloabs.

- also can use patient's serum with enzyme treated autologous red cell--> will remove cold agglutinin.

- also can use ZZAP reagent (this is combination of proteolytic enzyme and powerful reducing agent). ZZAP will remove IgM and COMPLEMENT coating the autologous red cell, and uncover antigen sites

Describe the Donath-Landsteiner test

autoanti-P associate with PCH which is cold reactive IgG autoabs.

aka known as biphasic hemolysis. the antibody doesn't react with routine test system, it only demonstrate by Donath Landsteiner test.

IgG autoabs cause PCH as biphasic hemolysis in vitro. the IgG autoabs bind to red cell at cold temp. when test is warmed at 37C, complement is activated and lysis red cell occur.

Donath Landsteiner test should performed on patients positive DAT from C3d, and demonstrate hemoglobinemia, hemoglobinuria

what is material use in Autoadsorption to remove autoantibodies?

it is patient's own RBCs.

Procedure involves removing antibody from a sample of the patient's RBC and subsequently incubating the patient's plasma with their stripped RBCs to allow more autoantibody to attach. The plasma is then separated from the recoated RBCs.

should not be performed if the patient has been transfused in the previous 3 months.

if can't use autoadsorption to remove autoantibodies, what other we can use

an alloadsorption technique may be performed using donor or reagent RBCs with known phenotypes.

What are the symptoms of Hemolytic Intravascular Transfusion Reaction?

Fever, chills, shock, renal failure, DIC, pain in chest, back, or flank

which Reactions associated with leukocyte antigens

TRALI (donor leukocyte antibodies)

Febrile non-hemolytic (antibodies to transfused lymphs, grans, or platelets)

TA-GVHD

which Reactions associated with plasma proteins

Urticaria

Anaphylactic shock

which Reactions associated with platelet antigens

Post transfusion purpura (platelet antibodies)

What is the trigger of Hemolytic Intravascular Transfusion Reaction?

by antigen-antibody reaction which activate complement, coagulation system and endocrine response