ADVIA 2120i Technology2012-160208 - 127086901 - 6

ADVIA 2120i System:
Analytical Technology
Unrestricted © Siemens Healthcare Diagnostics Inc. 2013 All rights reserved. Answers for life.
ADVIA 2120i System — analytical technology overview
Cytochemistry, nuclear density, and light scatter for white cell identification
Dual-angle laser light scatter measures size and refractive index for
red cell differentiation
Nucleic acid staining for reticulocyte counting
Dual-angle laser light scatter for platelet differentiation by size and refractive index
Unrestricted © Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.

Page 2 A91DX-HHS-130985-GC1-4A00 HHS/Hematology
Interpreting graphics
White cell cytograms

Erythrograms
Size
Platelets
Retics
Refractive index or density

Staining intensity
This presentation is for educational purposes only; cytogram patterns do not represent the full range of information needed for diagnosis.

White cell technology

ADVIA 2120i System — white cell technology
• Dual WBC count – internal QC

• Cytochemical classification according to:
• peroxidase activity
• nuclear density
• size and granularity
• Myeloperoxidase:
excellent stability and differentiation
• Expanded linearity of 400 x 109/L
• Accurate flagging and lower review rates

Quality of results – WBC
Two methods
Each method measures 2 cell characteristics
Method 1 — Size and perox staining
Method 2 — Size and nuclear density
Unique Siemens Solution

ADVIA 2120i — Peroxidase cytochemistry
Eosinophil RBC
Neutrophil Heat/SDS
formaldehyde
(Dil 1)
Monocyte
4-Chloro-1
Basophil Naphthol (Dil 2)
H2O2 (Dil 3)
Lymphocyte

WBC analysis
Size
Absorption

Quality of results — Baso channel — WBC
High angle detector

measures nuclear
density / complexity
Laser
Zero angle
Low angle detector

measures size
Primary WBC and baso count

Basophil / nuclear lobularity channel
1. Noise
2. Blast cell nuclei
3. Mononuclear WBCs
Cell Size
4. Basophils
5. Baso Suspect
6. Saturation
7. Polymorphonuclear WBCs
Nuclear Shape and Density
Accurate WBC counts
No lyse-resistant red cells
No cellular interference
Reduced manual reviews

Basophil/Nuclear lobularity channel
Eosinophil RBC
Neutrophil
Heat
Monocyte Phthalic Acid
Surfactant
Basophil
Lymphocyte

WBC analysis
Size
Complexity

Quality of results — Baso channel — WBC
High angle detector

measures nuclear
density / complexity
Laser
Zero angle
Low angle detector

measures size
Primary WBC and baso count

Basophil / nuclear lobularity channel
1. Noise
2. Blast cell nuclei
3. Mononuclear WBCs
Cell Size
4. Basophils
5. Baso Suspect
6. Saturation
7. Polymorphonuclear WBCs
Nuclear Shape and Density
Accurate WBC counts
No lyse-resistant red cells
No cellular interference
Reduced manual reviews

White cell cytograms
Normal
M1 AML
M3 AML

Quality of results — WBC
Dual WBC methodologies
Fewer repeats
Lower review rate
Faster TAT
Diagnostic scattergrams

PANDA
Peroxidase Activity and Nuclear Density Analysis
Courtesy of G. d’Onofrio, MD and G. Zini, MD

Peroxidase patterns
P2 P2
P0 P1 P3
P5
P0 P2
P1P3P4 P6
Courtesy of G. d’Onofrio and G. Zini

Nuclear density patterns — blasts present?
• D0
Normal rounded MNC cluster shape and no
blast flag
• D1
The MNC cluster is shifted down and left to D0
blast area (usually with flag)
D1 D1

Preclassification grid
P0 P1 P2 P3 P4 P5 P6
D0 CLL severe partial CML CML AIDS
PLL MPO-def. MPO-def. MDS
HCL CMML
ALL (L1-L3)
total MPO-def.
D1 ALL (L1-L3) M1 M2 M2 M2 M3v M3

NHL M5a M4 M4 M3v at. CML
M0 M2 M5a CML-BC M4
M5a M4 M5b
M6 M1
M7

Peripheral blood
P2
P2-D1

Preclassification grid
P0 P1 P2 P3 P4 P5 P6
D0 CLL severe partial CML CML AIDS
PLL MPO-def. MPO-def. MDS
HCL CMML
ALL (L1-L3)
total MPO-def.
D1 ALL (L1-L3) M1 M2 M2 M2 M3v M3

NHL M5a M4 M4 M3v at. CML
M0 M2 M5a CML-BC M4
M5a M4 M5b
M6 M1
M7

Peripheral blood
P2
M5/AML

ADVIA 2120i System — Body fluid software
Cerebrospinal fluid
Pleural fluid
Peritoneal fluid
Dialysate

Body fluid assay selection

ADVIA 2120i System CSF Assay Flow Chart
Combine equal volume 5 mins

CSF + reagent
Patient CSF Prepared

CSF – 300mL reagent sample
Run
Patient CSF result: sample
RBC/WBC count
and WBC Diff
TAT
5-10 minutes

Cerebrospinal fluid result
ADVIA® 2120i System Manual

CSF RBC 16 1
CSF WBC 3838 3089
CSF %MN 31 22
CSF %PMN 69 78
CSF %Neut 69 78
CSF %Lymph 3 3
CSF %Mono 28 19
CSF %EOS* 0 0
Example of bacterial meningitis

* Research use only

Cerebrospinal fluid result
ADVIA 2120i System Manual

CSF RBC 2 0
CSF WBC 8 6
CSF %MN 83 81
CSF %PMN 17 19
CSF %Neut 17 19
CSF %Lymph 21 30
CSF %Mono 62 51
CSF %EOS* 0 0
Example of ALL
* Research use only

CSF Assay
• Reduces labor, specialized staffing needs, and turnaround time

• Provides quality control, consistency of results, and 24 hour
testing availability
• White cell linearity to zero
• Increases prepared sample stability to enable sample transportation
and batch testing

Red cell and reticulocyte technology

Quality of results — RBC
RBC accuracy
Measures size and hgb concentration
Quantitative morphology results

Red cell technology
%Macro 120fl
60fl
%Micro
Hgb.conc. 28g/dl 41g/dl
%Hypo %Hyper

Red blood cells — clinical utility
% hypo, micro etc.
Aids thalassemia diagnosis

120fl
Detects spherocytosis
60fl
B12 / folate vs. liver / chemo
Monitoring of EPO therapy

Hgb.conc. 28g/dl 41g/dl
Quality of results — RBC
Lipemic samples
HGB 13.0 Plasma replacement

HGB = 12.1
MCHC 37.0
CHCM 34.7 Unique to ADVIA®

Direct HGB 12.2
Eliminates plasma replacement
Not effected by lipemia

Differential diagnosis:
Iron deficiency and b-thalassemia
MCV = Normal MCV = 61.1 fL MCV = 66 fL

M/H ratio = 0.63 M/H ratio = 2.05
Diagnosis = Diagnosis =
Iron Deficiency b-Thalassemia

Typical erythrograms
Normal Iron deficiency Iron treatment Hemoglobin S+C
Megaloblastic Cryoglobulins Hemolytic Uremia Spherocytosis
Elliptocytosis Sickle Cell Anemia a Thalassemia β Thal Major

Reticulocyte method
• RBC and platelets are isovolumetrically sphered

• Oxazine 750 differentially stains the reticulocytes
based on their RNA content

Reticulocyte parameters
• Reticulocytes slightly larger than mature

red cells (fL)
• Reticulocyte hemoglobin concentration
proportionately less (g/dL)
• Normal reticulocyte hemoglobin content
similar to mature red cells (pg)

Functionality of %Hypo and CHr
% hypochromic cells
Provides history of hemoglobin concentration
over previous 8-12 weeks
CHr
Provides snapshot of current hemoglobin
content in developing red cells

%Hypo/Hyper and CHr assays
• Provides screening capabilities for iron deficiency, especially

pediatrics
• Allows for better treatment management of EPO and iron
• Only method validated for monitoring hemodialysis patients in
NICE and European Guidelines
Beginning After 4 days After 2 weeks After 1 month

Cost effective
sTfR % Hypo
100
80
Er-ZPP T Sat
60
Sensitivity
40
Ferritin
20
0
0 20 40 60 80 100
Specificity
ref: Tessitori et al 1997

Conclusions (Dr. Iain Macdougall,
Consultant Nephrologist, Kings College, UK)
• EPO has revolutionized the management of renal anemia over the last decade.
The use of EPO for other clinical situations is becoming more widespread,
especially oncology
• It has, however, cast light on the concept of functional iron deficiency
• Serum ferritin and TSAT had major limitations in the detection of this condition
• Red cell and reticulocyte assessments using ADVIA 120, 2120, 2120i
technology provide very useful diagnostic information in this context

ADVIA 2120i System — platelet technology
Large Platelets
RBC Ghosts
Platelets
Debris
RBC Fragments

Impedance method
Platelet? Platelet?

ADVIA 2120i optical method
Platelet?
Platelet
Platelet?
Red Blood Cell X
Quality of results – PLT
Two cell characteristics
Size and RI (color)
Includes large platelets
Excludes micro RBCs, ghosts, fragments

Platelet method
• Provides two-dimensional analysis:

Large Platelets Volume and refractive Index
• Linearity – platelets 0 to 3500 x 109/L
RBC Ghosts
Platelets
• Size range - 1 to 60 fL
Debris • Large platelets included

RBC Fragments
• RBC fragments excluded
• Refractive Index - 1.35 to 1.40
• Indication of platelet activation

Platelets
Cat sample
RBC: 8.2 x 1012/L

MCV: 44.8 fL
PLT: 447 x 109/L
MPV: 17.1 fL

Evaluation of ADVIA 120 System platelet method
“ADVIA 120 very close to immuno method”

Advances in Routine Laboratory Platelet Counting
S.J.Machin, P.Harrison, D. Grant and C.Briggs.
British Journal of Haematology. 2000,108, 228-235

Platelet evaluation results
Impedance 1
Lipemic
200 Impedance 2
Impedance 3
160 ADVIA 120
120
80
40
ITP
0
0 20 40 60 80 100
Immunophenotypic count (x 109/L)

ADVIA 2120i System — nucleated red cell method

NRBC analysis algorithm
Three independent NRBC counts are determined for each sample:
• Two counts are based on the unstained region of the perox histogram
• The 3rd count is a combination of baso and perox counts (barox count)
• MN + baso from baso lobularity (= ba count)
• Neuts + eos from perox (= rox count)

WBC BAROX
An ‘uncontaminated’ BAROX WBC is created by merging “selected” clusters

of the BAso and peROX channels
WBC BA = MN + B
M N
L E Baso
MN PMN NRBC
NRBC
WBC ROX = N + E
Noise
WBC BAROX = (MN + B) + (N + E )

# NRBC = WBCB – BAROX WBC
Histogram analysis of perox unstained region
Red Raw histogram data of unstained

region of peroxidase cytogram
Blue Log-normal fit of noise region
Green NRBC events region
Yellow Gaussian fit of lymphocyte

region events
Amber Gaussian fit of NRBC events

ADVIA 2120i System — run screen
Areas where
NRBCs are
located

Summary NRBC method
The ADVIA 2120i System NRBC enumeration method is:
• Accurate and specific

• Automatically provides corrected WBC counts and differentials
• Does not rely on an NRBC flag
• Does not impact throughput
• Includes all stages of NRBC
• Every sample at no extra cost

Multispecies capability
• Random access of over 50 species

• Species-specific WBC differentials
• Species-specific normal ranges
• Superior differentiation between
platelets and RBCs
• Customizable species-specific flagging
• Playback feature for sample reanalysis

Versatile species selection
• Companion animals
• Farm animals
• Racing stables
• Laboratory animals
• Industry standard for clinical trials

Benefits of the ADVIA 2120i System
QUALITY results from ONE sample

• Dual WBC methods • Reticulocyte hemoglobin content (CHr)
• Peroxidase differential method • Effective renal patient EPO management
• Dual HGB methods (measured • FDA-approved CSF cell counts
and calculated) and differentials
• Quantitative RBC differential • Clinically useful scattergrams
• 2-D optical platelet count • Practical automation solutions with autoslide,
middleware,
• Reduced film review rates
and multidisciplinary options
Overall cost savings and confidence in results!

Why Siemens Hematology?
• Results right the first time

• Low film review rates
• Rapid turn around times
• Clinically useful parameters
• Confidence in results
• Cost effective quality
Practical Automation: Improved Productivity

Questions?


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ADVIA 2120i System:

Nucleic acid staining for reticulocyte counting

Unrestricted © Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.

White cell cytograms

Refractive index or density

Unrestricted © Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.

Unrestricted © Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.

• Dual WBC count – internal QC

Unrestricted © Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.

Each method measures 2 cell characteristics

Method 1 — Size and perox staining

Method 2 — Size and nuclear density

Unique Siemens Solution

Unrestricted © Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.

Unrestricted © Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.

Unrestricted © Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.

High angle detector

Low angle detector

Primary WBC and baso count

Unrestricted © Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.

Accurate WBC counts

No lyse-resistant red cells

Reduced manual reviews

Unrestricted © Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.

Unrestricted © Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.

Unrestricted © Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.

High angle detector

Low angle detector

Primary WBC and baso count

Unrestricted © Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.

Accurate WBC counts

No lyse-resistant red cells

Reduced manual reviews

Unrestricted © Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.

Unrestricted © Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.

Dual WBC methodologies

Lower review rate

Unrestricted © Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.

Courtesy of G. d’Onofrio, MD and G. Zini, MD

Unrestricted © Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.

Courtesy of G. d’Onofrio and G. Zini

Unrestricted © Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.

Courtesy of G. d’Onofrio and G. Zini

Unrestricted © Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.

D1 ALL (L1-L3) M1 M2 M2 M2 M3v M3

Courtesy of G. d’Onofrio and G. Zini

Unrestricted © Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.

Unrestricted © Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.

D1 ALL (L1-L3) M1 M2 M2 M2 M3v M3

Courtesy of G. d’Onofrio and G. Zini

Unrestricted © Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.

Unrestricted © Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.

Unrestricted © Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.

Unrestricted © Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.

Combine equal volume 5 mins

Patient CSF Prepared

Unrestricted © Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.

ADVIA® 2120i System Manual

Example of bacterial meningitis

Unrestricted © Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.

ADVIA 2120i System Manual

Unrestricted © Siemens Healthcare Diagnostics Inc. 2013 All rights reserved.

• Reduces labor, specialized staffing needs, and turnaround time