Blood Bank - Clinical Rotation Study Guide

Component Preparation - Whole Blood

CPD, C2PD - 21 days

CDPA - 35 days

AS - 42 days

Storage temp - 1-6 C

Volume - 450-500 mL

Dosage - H&H increase by 1 g/dL + 3%

Component Preparation - Irradiated Whole Blood

Original expiration or 28 days after IRR

Storage temp - 1-6 C

QC - IRR label color change

Volume - 450-500 mL

Dosage - H&H increase by 1 g/dL + 3%

Component Preparation - pRBCs

CPD, C2PD - 21 days

CDPA - 35 days

AS - 42 days

Storage temp - 1-6 C

Volume - 450-500 mL

Dosage - H&H increase by 1 g/dL + 3%

Component Preparation - RBC aliquots

Closed - no change to exp

Open - 24 hour exp

Storage temp - 1-6 C

Volume - varies based on weight of Pt

Dosage - per 10 mL/kg increase HGB 2 g/dL

Component Preparation - RBC IRR

Original expiration or 28 days after IRR

Storage temp - 1-6 C

QC - IRR label must change color

Volume - 300-500 mL

Dosage - H&H increase by 1 g/dL + 3%

Component Preparation - RBC LR

Closed = no change to exp

Open = 24 hour exp

Storage temp - 1-6 C

QC - >85% RBC mass, <5×10^6 WBCs

Volume - 300-500 mL

Dosage - H&H increase by 1 g/dL + 3%

Component Preparation - Washed RBC

24 hour expiration

Storage temp - 1-6 C

QC - HCT 70-80%

Volume - 200 mL

Dosage - H&H increase by 1 g/dL + 3%

Component Preparation - Frozen RBC

10 year exp

Storage temp - < -65 C

Component Preparation - Deglycerolized RBC

24 hour exp

Storage temp - 1-6 C

QC - 80% RBC recovery + <1% detectable glycerol

Volume - 200 mL

Dosage - H&H increase by 1 g/dL + 3%

Component Preparation - Random Donor PLT

5-7 day exp

Storage temp - 20-24 C

QC - >5.5×10^10 PLTs, pH >6.2

Volume - 50-100 mL

Dosage - PLT count 5-10,000

Component Preparation - Single Donor PLT

5-7 day exp

Storage temp - 20-24 C

QC - >3×10^11 PLTs, pH >6.2

Volume - 200-400 mL

Dosage - PLT count 30-60,000

Component Preparation - PLTs IRR

5 day exp

Storage temp - 20-24 C

QC - IRR label color change

Volume - varies by RDP vs SDP

Dosage - varies by RDP vs SDP

Component Preparation - PLTs Pooled

4 hour exp

Storage temp - 20-24 C

QC - pH >6.2

Volume - varies

Dosage - varies

Component Preparation - FFP - Frozen, 1 year, 7 years

1 year exp, 7 year exp

Storage temp - < -18 C, < -65 C

Volume - 200-400 mL, 200-1,000 mL

Dosage - Coag factors by 20-30% per 10-20 mL

Component Preparation - FFP - Thawed

24 hour exp

Storage temp - 1-6 C

Volume - 200-400 mL

Dosage - Coag factors by 20-30% per 10-20 mL

Component Preparation - PF24 - Frozen, 1 year, 7 years

1 year exp, 7 year exp

Storage temp - < -18 C, < -65 C

Volume - 200-400 mL

Dosage - Coag factors by 20-30% per 10-20 mL

Component Preparation - PF24 - Thawed

5 day exp

Storage temp - 1-6 C

Volume - 200-400 mL

Dosage - Coag factors by 20-30% per 10-20 mL

Component Preparation - Cryoprecipitate

Frozen - 1 year exp

Thawed - 6 hour exp

Pooled / Open - 4 hours exp

Storage temp - frozen @ < -18 C, thawed @ 20-24 C

Volume - 10-25 mL

Dosage - FBG + coag factors per 5-10 mL

Component Preparation - Granulocytes + IRR Granulocytes

24 hour exp

Storage temp - 20-24 C

QC - > 1×10^10 IRR

Volume - 200-600 mL

Antibody Identification - Impacts of Dosage

Different strength reactions

If panel cells are homozygous, a strong reaction may be seen

If panel cells are heterozygous, reaction may be weak or even non-reactive

Duffy, Rh, Kidd, MNSs all exhibit dosage

Antibody Identification - Impacts of Multiple Antibodies

Different strength reactions

Matching pattern is difficult

Number of selected cells needed depends on how many antibodies are identified

Emergency Issue of Blood Products - General Policy

Emergencies or life-threatening situations (no time for T/S results)

Emergency issue blood products require physician signature to accept risks associated w/ uncrossmatched blood

Emergency issue blood may be sent through the tube system or in a cooler, but must be specially labeled from regular blood products

BB must obtain: ordering provider, patient’s name, patient’s MRN, patient’s approximate age, patient’s sex, + location of patient

WellSky: from patient dropdown menu, use Patient Registry tab to search for patient’s MRN

If purple click on Registry button + save; if not purple continue on to Emergency Issue

Proceed to Product Selection + Emergency Issue

Emergency Issue:

Select appropriate number of requested products

If RBC/WB requested: label tube w/ unit number + remove a segment from each unit for compatibility testing, apply bright orange “Uncrossmatched Blood” label front of RBC unit or on transfusion tag, apply a Hemo Temp II temperature indicator on the back of the unit

Click on “Emergency Issue” tab

Enter patient’s MRN into the field + tab. Information will populate or register the patient if not

Enter pickup information: ID, location, inspection, physician. Select Products button will become active

Click on Select Products button

The Product Code Selection form appears. The product group, number of units, + attending physician all must be entered in order to select units. Click OK when fields are complete

The Product Selection window appears. Scan the unit number + product code of each unit. Once all units are entered, click OK

Verify all information on screen. Click Save

Select Emergency Issue + then say yes

Print Emergency Release tag. Confirm information on tag. Write time + date of issue. Attach tag to unit

What pRBC products are issued

O pos → men + women >50 years old

O neg → women <50 years old

What plasma products are issued

A FFP

Fisher Race + Weiner Conversions

dce → r

dCe → r’

dcE → r”

dCE → ry

Dce → R0

DCe → R1

DcE → R2

DCE → Rz

DAT

Detects in vivo sensitization of RBCs w/ IgG or complement that is associated w/ the following conditions: Transfusion reactions - alloantibodies coating donor cells; HDFN - maternal antibodies coating fetal cells; Autoimmune disorders hemolytic anemia - autoantibodies coating self cells; Drug-induced hemolytic anemia - drug reactions in plasma destroy RBCs

DAT is not required in pre transfusion testing for blood products

DAT is positive - something (IgG or C3b) is coating the surface of the patient’s RBCs in-vivo; IgG AHG+ implies antibody is coating RBC cell surface → investigate; C3b AHG+ implies complement is coating RBC cells surface → no further workup

DAT Procedure - Tube Method

Blood collection in EDTA tube

Blood made into 3-5% cell suspension + 2 drops are added to a new tube

New tube’s blood is washed x3 w/ saline

Reagent AHG is added →

If no agglutination occurs → indicator cells are added; new reaction should be positive = negative interpretation

If agglutination occurs → follow up w/ an eluate if patient has been transfused in last 2-3 weeks = positive interpretation

IAT

In-vitro test to evaluate the presence of allo or auto antibodies in patient plasma via 3 commonly performed tests: Antibody screening/Antibody ID (panels)/Antibody titers, Antigen typing, Crossmatching

A positive IAT test typically means that in-vitro sensitization of RBCs w/ IgG or complement has occurred due to: Alloimmunization from previous transfusions; Alloimmunization from previous or current pregnancy; Autoimmune disorders; Drugs or medications that may cause hemolysis

IAT Procedure - Tube Method

Blood collected in EDTA tube + centrifuged

Patient plasma added to tube w/ reagent cells

Centrifuged + read at immediate spin

Add enhancement media (LISS, PEG, 22% Albumin)

Incubate at 37 C for 15 min

Centrifuge + read at 37 C (LISS + Albumin)

Wash x3 w/ saline

Add AHG →

If negative → indicator cells added to validate reaction; should be positive = negative interpretation

If positive → follow up w/ appropriate reflex test = positive interpretation

DAT vs IAT

Cord Blood Testing On

Infants born to Rh negative mothers

Infants born from Group O mothers

Infants in the NICU

Infants born from mothers w/ clinically significant antibodies

Infants born from mothers w/ no prenatal history

Candidates for RhIG

Rh negative women

Weak D negative women

Non-immunized Rh-negative women

Rh negative mothers w/ Rh positive infants

After invasive procedures (miscarriages, abortions, ectopic, + amniocentesis)

Process for Receiving Units from Licensed Blood Supplies

Unpack carefully + inspect products for proper storage temp + condition

Mark on supplier’s packing list in appropriate area whether or not the units were received in good condition + the temp was satisfactory

Use the Inventory Module, Receive Blood Products pane: Enter batch details at top of the window: for receipt number enter your initials, enter supplier, enter received date/time, enter temp/inspection/quantity of products to receive

Tab to Unit Number field + scan the product’s unit number → tab to ABO/Rh field + scan the product’s ABO/Rh barcode → tab to Product code field + scan the product code → tab to Exp D/T field + scan product’s expiration date barcode → tab to volume field, if necessary, enter the volume of unit (Ex: FFP + Plateletpheresis) → if unit is Autologous/Directed/Reserved, enter the corresponding letter into A/D/R field → if unit has any special attributes, such as CMV negative, HGB S negative, or antigen typings, enter “Y” in AAA field (to select attributes) or scan corresponding barcode → tab through remaining fields, pane will expand to next line for any additional units to be received → after all units have been entered, click the Save button → file all checked forms in the file “Miller Shipping Forms” + return originals to supplier

Process for Confirming Units from Licensed Blood Suppliers

WB/RBC/AGRs (storage = refrigeration 1-6 C): detach 3 segments from unit → attach unit number to each segment + place 2 labeled segments into biohazard bag labeled w/ date received. Place bar-coded unit number on test tube for UNIT ABORH confirmation testing + place a segment in this test tube → place bag of segments into current week’s storage box → segments are retained for minimum of 7 weeks → using remaining segment, confirm ABO/Rh typing by performing unit ABO/Rh confirmation testing (only need forward type)

Results reporting: results of unit confirmation entered in LIS Testing Module using Active Tests pane: highlight all units to be tested, right click Enter Test Results → batch worksheet without Observations for Ortho Vision results, batch worksheet w/ Observations for manual tube/gel results; for Batch worksheets without Observations (Ortho Vision results) → reactions from Ortho Vision will appear in corresponding fields, click Verify + Save; for Batch worksheet w/ Observations (manual results) → enter reagent rack, enter reactions + interpretations for each unit to be tested, click Verify + Save

ABO Antigen Frequencies

O = 45%

A = 35%

B = 15%

AB - 5%

Rh Antigen Frequencies

D = 85%

C = 70%

E = 30%

c = 80%

e = 98%

K Antigen Frequencies

K = 9%

k = 98%

Kpa = 2%

Kpb = 99%

Jsa = <1%

Jsb = 99%

Duffy Antigen Frequencies

Fya = 65%

Fyb = 83%

Kidd Antigen Frequencies

Jka = 77%

Jkb = 73%

Lewis Antigen Frequencies

Lea = 22%

Leb = 72%

P Antigen Frequencies

P = 100%

P1 = 79%

MNSs (U) Antigen Frequencies

M = 78%

N = 72%

S = 55%

s = 89%

U = 99%

Lutheran Antigen Frequencies

Lua = 8%

Lub = 99%

Probability of Finding Compatible Donor Units

# of units needed / antigen frequency (decimal)

If more than one antigen frequencies multiply them together (decimal)

ABID Panels

All panel cells are Group O

Cells have been fully phenotyped: + refers to presence of antigen in reagent cell, 0 refers to absence of antigen in reagent cell

Autocontrol run to determine if antibody is an allo or auto-antibody

1 drop of panel cell + 2 drops of patient’s serum

Add 2 drops of AHG

Centrifuge + examine for agglutination

Add 1 drop of Coombs control cells (should agglutinate)

“Ruling out” means crossing out antigens that did not react w/ the patient’s plasma (which contains antibody): antibody will only react w/ cells that have the corresponding antigen

Consider antibody’s usual phase of reactivity - IgM like it cold (RT) while IgG like it hot (37 C)

Look for matching pattern

Electronic Crossmatch

Detect ABO incompatibilities between donor red cells + recipient serum using computer LIS system

Requires:

Patient has 2 separate ABO/Rh typings on record

Patient has no clinically significant antibodies or history of a clinically significant antibodies

Patient has a current specimen

Immediate Spin Crossmatch

Detect ABO incompatibilities between donor red cells + recipient serum

Requires:

Patient has no present or previous clinically significant antibodies

In-date T&S

Extended Crossmatch

Recipient plasma + donor red blood cells are mixed, incubated, + taken through to Coombs

Prevents transfusion of incompatible red cells + to select blood products that when transfused to the recipient will have acceptable survival

Requires:

Patient has positive antibody screen

History of an antibody (including non-specific, non-clinically significant antibody, WAA, + cold reacting antibodies

Selection of Appropriate Blood Products Based on Patient + Donor Blood Types

A - WB = A, RBC = A or O, Plasma = A or AB, PLTs = A or O

B - WB = B, RBC = B or O, Plasma = B or AB, PLTs = B or O

AB - WB = AB, RBC = AB or A or B or O, Plasma = AB, PLTs = AB or A or B or O

O - WB = O, RBC = O, Plasma = O or A or B or AB, PLTs = O

Acute Hemolytic Transfusion Reaction

Chills accompanied by 1 or more of the following signs or symptoms:

Shock, hypotension, back pain, dyspnea, chest pain, nausea, hemoglobinuria, oliguria or anuria, generalized bleeding, flushing

Febrile Non-Hemolytic Transfusion Reaction

Temperature rise of 1 C or more occurring in association w/ transfusion + without any other explanation

Temp rise may occur early in the transfusion or may not occur until an hour or 2 after the transfusion has finished

Anaphylactic Transfusion Reaction

Occur after an infusion of only a few mLs of blood or plasma

No fever or temp rise

Following symptoms may occur: coughing, bronchospasm, respiratory distress, vascular instability, nausea, abdominal cramps, vomiting, diarrhea, shock, or loss of conscientiousness

Urticaria/Allergic Transfusion Reactions

Characterized by local erythema, hives, + itching usually without fever or other adverse effects

Hypotensive Transfusion Reaction

Characterized by hypotension occurring during or within 1 hour after cessation of transfusion (all other adverse reactions presenting w/ hypotension are excluded)

Adults experience a drop in systolic BP of >30 mmHg + systolic BP <80 mmHg

Infants, children, adolescents experience >25% drop in systolic BP from baseline

TACO

Characterized by hypervolemia due to excess volume or speed of infusion

Symptoms may include: congestive heart failure, dyspnea, severe headache, peripheral edema during or soon after transfusion

TRALI

Characterized by acute respiratory distress, bilateral pulmonary edema, + hypoxemia in setting of transfusion of plasma-containing blood components

Hypotension + fever are frequent symptoms

Onset of symptoms is 1-2 hours following beginning of transfusion (can be up to 6 hours after transfusion)

TAD

Characterized by acute respiratory distress occurring within 24 hours of cessation of transfusion + without any other explanation

Delayed Serological Transfusion Reaction - Primary Immunization

Mild, occurs several weeks after transfusion + result of patient’s initial development of antibody

Antibodies are detectable no earlier than 7-10 days after transfusion + usually several weeks or months later

Unexplained fall in HGB concentration, positive DAT, detection of new red cell alloantibody

Delayed Serological Transfusion Reaction - Anamnestic Responses

Secondary response to transfused red cell antigens in a previously immunized recipient

Some alloantibodies formed after primary immunization may diminish to levels undetectable in serum

Pre-transfusion testing reveals no unexpected antibody + no serologic incompatibility, but within 3-7 days after transfusion, an anamnestic response leads to high levels of IgG antibodies that react w/ the transfused cells

Fever, unexplained fall in patient’s HGB, mild jaundice, hemoglobinuria may occur; acute renal failure is uncommon

Use of Ficin / Enzyme Treated Panels + How That Impacts ABID Workup

Can be used to modify the RBC surface of reagent cells of an ABID panel to increase or decrease expression of non-ABO blood group antigen systems

Used when there is reason to suspect underlying alloantibodies in the presence of an antibody to a high prevalence antigen on the reagent cell panel

Cannot rule out antigens that are destroyed by enzymes even if the reagent cell is unreactive

Enhanced activity - Rh, Kidd, Lewis, I / i, ABO

Decreased / No activity - Duffy, MNSs, Xga

Unaffected - Kell

Weak D Testing for Rh-negative Babies

Prepare a 3-5% solution of patient’s cord red cells in isotonic saline - add 2 drops cord RBCs, add 2 mL saline + mix well

Add 1 drop of D antisera + 1 drop of 3-5% cord red cell suspension + mix well in new test tube

To another test tube add 1 drop of D control + 1 drop of 3-5% red cell suspension

Incubate at 37 C for 15-60 min

Wash cells (4x) w/ isotonic saline

Add 2 drops of IgG AHG

Mix well + centrifuge for 15 secs at 3400 rpm

Resuspend cells by gentle agitation + examine macroscopically for agglutination

Add Ortho Coombs Control to negative tests

Weak D - Explanations

Position effect - due to location of D + C genes, RhD is on the opposite chromosome (in trans) w/ the RhCE gene, DO NOT make an anti-D

Quantitative changes - D antigen is completely formed but not fully expressed, single nucleotide polymorphism (SNPs) that subtly change the D antigen structure, very very rarely make an anti-D

Del - extremely low number of D antigens

Partial D or D Mosaic - incomplete epitopes within the entire D protein, may make an anti-D antibody against the epitope(s) that they lack

PEG Adsorption Technique

PEG - enhances antibody uptake by removing water

Used to confirm the presence of the warm-reactive autoantibody + detect or identify any additional alloantibody(ies) that may be present

PEG treatment removes autoantibody → reducing the strength of the positive DAT + free antigen sites for adsorption

Used for patients that have not been transfused in the past 3 months

PEG Adsorption Procedure

Make 1:1 ratio of patient’s pRBCs, plasma + PEG

Mix well + incubate at 37 C for 15 min

Centrifuge at 3500 rpm for 5-10 min

Harvest the adsorbed plasma using a transfer pipette

Test absorbed plasma w/ non-treated screening cells I + II + the patient’s own cells (auto control) by the antiglobulin technique by: add 4 drops of adsorbed plasma + 1 drop of respective screening cells I + II + auto suspension, incubate screening cells + autocontrol at 37 C for 30 min, wash the contents of the tubes (4x), add 2 drops of Anti-IgG, mix well, + spin (centrifuge)

If 2 cell screen + autocontrol are negative - adorption is complete → crossmatch w/ adsorbed plasma, using the same procedure as for screening cells

If only one of the cells or both of the screening cells in the 2 cell screen are positive, the auto is negative, then a panel must be performed using the one-time adsorbed plasma to identify any underlying allo-antibody

If the 2 cell screen + autocontrol are positive - repeat adsorption using the 1x adsorbed plasma + new aliquot of patient’s untreated red cells (DO NOT add more PEG)

LISS (OAES) Enhancement Media + Why we Use it

Enhances antibody uptake + decreases incubation to 15 mins by reducing zeta potential (increases serum:cell ratio so increases ionic strength of Ab/Ag complexes)

FMH Testing

Massive Fetomaternal Hemorrhage (FMH) → 30 mL of whole blood, specimen must be post-delivery/trauma

Diagnostic test: Fetal Screen or rosette test:

Qualitative test

Purpose: detects Rh positive cells

Procedure: maternal red cells incubated w/ Anti-D serum → test washed to remove any unbound antibody → D positive indicator cells are then added → test is spun + read MICROSCOPICALLY

Interpretation: Positive = rosette formations, Negative = only loose red cells

Kleihauer-Betke Test

Quantitative test

Principle: Fetal HGB is resistant to acid elution, therefore fetal cells retain HGB + stain bright pink; Adult cells are not resistant, so they lose HGB + are only left w/ stroma (appear as ghost cells); Enables the calculation of additional doses of RhIG

Procedure: prepare slide + stain, count 2000 adult cells + note the number of fetal cells within that 2000 count, perform RhIG calculations

Negative FMS or KB testing of an Rh neg woman that gave birth to an Rh pos baby results in 1 vial of RhIg given

RhIG Calculations

% fetal cells = # of fetal cells / total cells counted

Volume of FMH (mL) = % fetal cells x 5000 (estimated total fetal blood in maternal circulation / volume of fetal blood)

Doses of RhIG = volume of FMH / 30 (1 vial of RhIG neutralizes 30 mL of whole blood)

>.5 = round up

<.5 = round down

Add safety vial