ALOPS hematology, coagulation and blood bank analyzers

What is the gold standard for blood bank testing?

Tube testing

What tests are included in immediate spin for Blood banking?

ABO, Rh, Donor confirmation

Indirect Antiglobulin test (IAT) involves what tests?

Antibody screen, Antibody ID, IgG cross match, phenotype, weak D

For screen cells testing, what reacts with the respective reagent cells?

Patient serum/plasma

Automated instruments can perform tests to perform blood bank tests such as

SPRCA assays, microtiter plates, gel technology

The TANGOTM Infonity (Bio Rad) system

Is a type of automated SPRCA system

Combines Erytype S and Solidscreen II

CaptureTM system (Immuncor)

Microtiter plates accommodate SPRCA or hemagglutination testing

Hemagglutination Assays

• Assays are performed on microplates using the same principles as in tube agglutination

• Bar-coded samples and reagents are used

• Plates are automatically read using a camera reader

BIO-RAD Erytype S

• Hemagglutination assay

• ABO, D, and other RBC antigen testing

• Monoclonal antisera are dried in microwells

Principle and diagram of Erytype S hemagglutination assay

SPRCA Assays Are used for

Indirect and direct antiglobulin tests

SPRCA Assays Components and result interpretations

Contain Microtiter wells that have RBC membranes bound to the surface

• Patient antibodies, if present, will attach to the corresponding antigen covering the well

• Negative results forma button at the bottom of the well

• Capture-R Select assay system (Immucor) immobilizes RBCs others than screen or panel cells

SPRCA test procedure diagram with steps

grading chart for SPRCA assay

BIO-RAD Solidscreen II In blood bank testing

• Solid-phase assay that detects RBC antibodies in serum or plasma

• Wells are precoated with protein A, which has a high affinity for the Fc portion of immunoglobulins

• Positive test: RBCs and antibody to immunoglobulin G (anti-IgG antibody) bind with protein A, forming a smooth monolayer

• Negative test: RBCs form a button at the bottom of the well

A negative test in Bio Rad solidscreen II testing for blood bank would be

RBCs forming a button on the bottom of the well

Solid screen II is suitable for what applications?

Solid phase: Immucor Lumena

• Holds

• 20 specimens • 16 reagents

• 32 strips

• 1 camera for analysis • 2 barcode readers

• 1 probe

• Strips transported to incubator, centrifuge, wash station, or pipette station

Gel: Ortho ProVue

• Carousel holds

• 48 specimens

• 16 reagents

• 2 diluents

• 3 cameras for analysis

• 24 position capacity card block • Probe performs dilutions

• Gel card gripper transports cards to centrifuge or service rack

Electrical Impedance Instruments

Blood Aspirated, 1st aliquot : RBC/Plt dilution chamber

• External electrode

• 3 apertures (ea w/ internal electrode)

3 RBC counts are obtained, compared, and evaluated. If agreement, the reported RBC count is an average of the 3 counts.

In Gaussian curve, RBC sizes are interpreted as

If RBCs larger than normal, right shift

If RBCs smaller than normal, left shift

1st Aliquot determinations:

Data determines:

◦ RBC count

◦ MCV

◦ RDW-CV and RDW-SD ◦ Plt count

◦ MPV

Calculated parameters:

◦ Hct

◦ MCH ◦ MCHC

2nd Aliquot

Delivered to WBC/hemoglobin dilution chamber

◦ Lytic agent lyses RBCs and converts released hgb to cyanmethomoglobin, and shrinks the leukocyte cell membrane and cytoplasm... allowing WBC count to represent cell volume rather than cell size.

◦ WBC count measured by electrical impedance from 3 apertures and reported count represents the average.

◦ Hgb determined by absorbance reading at 525 nm (Beer’s Law)

3rd Aliquot

Delivered to the orbital mixing chamber:

1. Blood mixed with heated lysing agent to remove RBCs

2. Stabilizing agent added to preserve WBCs

3. Cells sent through the volume-conductivity-scatter (VCS) flow cell by hydrodynamic focusing for 5-part differential

• Cell volume – by impedance

• Cell conductivity – by electromagnetic probe. Determines physical and chemical components

• Cell’s light scatter characteristics - determines internal contents and cell surface and size

Optical Light Scatter

Each cell flows in a single line through a flow cell

A laser device is focused

On striking cells, light is scattered in different directions

Sensor captures and multiplies scatter

• Forward angle light scatter (FALS)– cell size

• Side scatter (SS) – granularity

4th Aliquot

Delivered to a heated dilution chamber

◦ Mixed with new methylene blue reagent

◦ An acidic, hypotonic solution is added ◦ Eluteshemoglobin

◦ PreservesprecipitatedRNA

◦ SpheresRBCs(eliminatesinterferenceduetovariance

of shape)

◦ Sent to VCS for analysis to classify mature vs.

immature RBCs

1. Cell volume – by impedance

2. Cell conductivity – by electromagnetic probe.

Determines physical and chemical components

3. Cell’s light scatter characteristics - determines internal contents and cell surface and size

Platelet satellitosis would occur when

The sample is warm

When automated platelets counts encounter platelet clumping, what should you do

Recollect in heparin

What is a fully automated instrument used in Coagulation testing?

Sysmex C1000

Coagulation testing: Electromagnetic monitoring involves

monitoring of movement of a steel ball in plasma

Ball moves because of applied magnetic force

Clot = ↓ movement sensed by electronic sensor set to predefined limit

Photo-Optical Systems In coagulation testing involve

Detection of sample optical density due to formation of fibrin

•Records decreased light at the forward 180 ̊ angle

Nephelometric End Point Detection In coagulation testing

• Modification of photo-optical end point detection

• 90 degree or forward angle scatter contributes to measurement Ag-Ab form precipitates that scatters incident light

• Nephelometric instruments read loss of intensity of exiting beam as increasing amounts of impinging light scattered agglutinates form

Thromboelastography (TEG) - ROTEM

Whole blood clotting assay

Allows real-time comprehensive evaluation of hemostasis

Can analyze fibrinogen, factor activity, platelet function, and fibrinolysis

PFA 100 Platelet Function Analyzer (Dade-Behring)

assesses platelet dysfunction

Measures plt function in whole blood using: collagen/epinephrine, collagen/ADP

Detects platelet plug Formation

Platelet aggregation graph interpretation