Cell Blocks Vinod B.

Shidham,
In MD, FRCPath, FIAC
Cytopathology vshidham@med.wayne.edu

Cell Blocks
In Cytopathology
May 17 & 19, 2010 (Workshop# 5 & 43)
Vinod B. Shidham, MD, FRCPath, FIAC
Professor
Co-editor-in-chief & Executive editor, CytoJournal (www.cytojournal.com)
Vice-chair
Director of Cytopathology, Cytotechnology School, Cytopathology fellowship, &
GI fellowship
Dept of Pathology, Wayne State University Medical School
Detroit, MI 48201, USA
vshidham@med.wayne.edu
 Cell Blocks Vinod B. Shidham,
In MD, FRCPath, FIAC
Cytopathology vshidham@med.wayne.edu

Disclaimer
VS is co-editor of
‘Cytopathologic Diagnosis of
Serous Fluids’
Elsevier (W. B. Saunders Company)
cited for various methods of
cell block preparation (the
sketches and tables used are
from this reference).
 Cell Blocks Vinod B. Shidham,
In MD, FRCPath, FIAC
Cytopathology vshidham@med.wayne.edu

Disclaimer
VS has indirect financial interest (through spouse) in
AV marker mentioned in the workshop.

http://www.bioinnovationllc.com/Page_2.html
Cell Blocks Vinod B. Shidham,
In MD, FRCPath, FIAC
Cytopathology vshidham@med.wayne.edu

Outline
Cell block- microbiopsy-
Role in cytopathologic evaluation/patient management
Critical issues to be considered
Different methods of cell block preparation
Aligning the cells along the cutting surface
Depth of section cutting
Immunophenotyping and cell blocks-
Immunoreactivity interference
Marker for SCIP approach
A few study cases
Q/A session
Cell Blocks Vinod B. Shidham,
In MD, FRCPath, FIAC
Cytopathology vshidham@med.wayne.edu

Cell block of cytopathology specimen is

Equivalent to microbiopsy evaluation
the interface between cytology and histopathology
(bridge to histopathology/surgical pathology)

Routine example is Endocervical curretage (ECC),

But without cytology preparation

Cell block with cytology preparation has

Added benefit of excellent cytomorphologic details
in concert with architectural insight
Cyto-histo-pathology
Cell Blocks Vinod B. Shidham,
In MD, FRCPath, FIAC
Cytopathology vshidham@med.wayne.edu

Role of cell blocks in cytopathologic evaluation/patient management

1. Immunophenotyping
2. Special stains- Mucicarmine, Congo red, organism stains
3. Architectural evaluation-
Trabecular-sinusoidal pattern in HCC,
Hollow or solid proliferation spheres without cores
in carcinoma versus mesothelioma in effusions
Evaluate for invasion
Comparative evaluation with surgical pathology material
E.g- Peritoneal/pelvic washing
Quantification of some features such as mitotic figures
4. Enhanced sampling of FNAB rinses
5. Molecular test
e.g. FISH, CISH, In-situ PCR
6. Archival for future studies
 Cell Blocks Vinod B. Shidham,
In MD, FRCPath, FIAC
Cytopathology vshidham@med.wayne.edu

Critical issues to be considered

Depending on the primary indication,
the method of cell block preparation vary

Multiple variants should be considered

for selecting the method and modifying it as needed for
individual specimens
Specimen type- Fresh versus fixed cells
Cellularity of the specimen
Nature of cell distribution-
predominantly solitary cells versus microfragments/aggregates
Ancillary tests anticipated
Available resources/infrastructure in the lab
Institutional and regional biases
 Cell Blocks Vinod B. Shidham,
In MD, FRCPath, FIAC
Cytopathology vshidham@med.wayne.edu

Different methods of cell block preparation

A. Cell block from specimen with clot or significant sediment- FNAB

B. HistoGel method
C. Gelatin embedding
D. Agar embedding method
E. Plasma-thrombin method
F. Collodion (Celloidin) bag method

G. From scraped material from cytology smears

H. From Millipore filters
I. From cells lifted selectively from the cytology preparation
(Kaneko C et al. Diagn Cytopathol 2000;22:117–119)
Cell Blocks Vinod B. Shidham,
In MD, FRCPath, FIAC
Cytopathology vshidham@med.wayne.edu

Shidham & Epple

Ch 14 ‘Cytopathologic Diagnosis of Serous Fluids’
Elsevier (W. B. Saunders Company)
 Cell Blocks Vinod B. Shidham,
In MD, FRCPath, FIAC
Cytopathology vshidham@med.wayne.edu

Let the remaining Aspirate 10% formalin Gently and firmly Transfer the aspirated
aspirate clot in the from the container in remove the plunger of formalin with
syringe for 5 to 7 which the specimen is the syringe . dislodged cot in to the
minutes (slightly to be submitted for specimen container
longer than the cell block processing. with 10% formalin
clotting time). This dislodges the clot fixative
from syringe wall.

1 2 3 4

Processing of FNA aspirate to be submitted to laboratory for Cell block

Cell Blocks Vinod B. Shidham,
In MD, FRCPath, FIAC
Cytopathology vshidham@med.wayne.edu

Protocol for plasma-thrombin method for cell block preparation

Shidham & Epple

Ch 14 ‘Cytopathologic Diagnosis of Serous Fluids’
Elsevier (W. B. Saunders Company)
Cell Blocks Vinod B. Shidham,
In MD, FRCPath, FIAC
Cytopathology vshidham@med.wayne.edu

Protocol for cell block preparation with collodion bag

Shidham & Epple

Ch 14 ‘Cytopathologic Diagnosis of Serous Fluids’
Elsevier (W. B. Saunders Company)
Cell Blocks Vinod B. Shidham,
In MD, FRCPath, FIAC
Cytopathology vshidham@med.wayne.edu

Protocol for preparing cell blocks with HistoGel

Shidham & Epple

Ch 14 ‘Cytopathologic Diagnosis of Serous Fluids’
Elsevier (W. B. Saunders Company)
Cell Blocks Vinod B. Shidham,
In MD, FRCPath, FIAC
Cytopathology vshidham@med.wayne.edu

Aligning the cells along the cutting surface

Depth of section cutting

 Cell Blocks Vinod B. Shidham,
In MD, FRCPath, FIAC
Cytopathology vshidham@med.wayne.edu

Varsegi GM, Shidham V (2009)

Cell Block Preparation from Cytology Specimen with Predominance of Individually
Scattered Cells.
Journal of Visualized Experiments (JoVE) 2009 Jul 21;(29). pii: 1316.
doi: 10.3791/1316. PMID: 19623160
Cell Blocks Vinod B. Shidham,
In MD, FRCPath, FIAC
Cytopathology vshidham@med.wayne.edu
 Cell Blocks Vinod B. Shidham,
In MD, FRCPath, FIAC
Cytopathology vshidham@med.wayne.edu

Video of JoVE article (8 minutes 15 sec)

Video article FREE on web as open access at-
At this stage,
http://www.jove.com/index/Details.stp?ID=1316
please note
Or atthe URL
and observe the full FREE video in the new internet window
http://alturl.com/zkns

Video of JoVE article (8 minutes 15 sec)

Cell Blocks Vinod B. Shidham,
In MD, FRCPath, FIAC
Cytopathology vshidham@med.wayne.edu

From:
Varsegi GM, Shidham V (2009)
Journal of Visualized Experiments
(JoVE) 2009 Jul 21;(29). pii: 1316.
doi: 10.3791/1316. PMID: 19623160
Cell Blocks Vinod B. Shidham,
In MD, FRCPath, FIAC
Cytopathology vshidham@med.wayne.edu

From:
Varsegi GM, Shidham V (2009)
Journal of Visualized Experiments
(JoVE) 2009 Jul 21;(29). pii: 1316.
doi: 10.3791/1316. PMID: 19623160
Cell Blocks Vinod B. Shidham,
In MD, FRCPath, FIAC
Cytopathology vshidham@med.wayne.edu

Modified from:
Varsegi GM, Shidham V (2009)
Journal of Visualized Experiments
(JoVE) 2009 Jul 21;(29). pii: 1316.
doi: 10.3791/1316. PMID: 19623160
Cell Blocks Vinod B. Shidham,
In MD, FRCPath, FIAC
Cytopathology vshidham@med.wayne.edu

From:
Varsegi GM, Shidham V (2009)
Journal of Visualized Experiments
(JoVE) 2009 Jul 21;(29). pii: 1316.
doi: 10.3791/1316. PMID: 19623160
Cell Blocks Vinod B. Shidham,
In MD, FRCPath, FIAC
Cytopathology vshidham@med.wayne.edu

Immunophenotyping and cell blocks-

Factors affecting immunoreactivity-
Loss, reduction, or enhancement of antigen immunoreactivity
Exposure to different reagents and fixative(s)
Temperature
Storage of specimen with or without fixative

Subtractive Coordinate Immunoreactivity Pattern (SCIP) approach

Shidham & Atkinson
Ch 5. Immunocytochemistry of effusion fl uids: introduction to SCIP approach.
‘Cytopathologic Diagnosis of Serous Fluids’
Elsevier (W. B. Saunders Company)
 Mesothelial & X Metastasis Y Metastasis Z
inflammatory cells (carcinoma) (non-carcinoma)

vimentin 1 2 1 2 1 2
A
3 3
4 3 4
6 5 6
5 4

7 6
8 7 5
7
In

1 2 1 2 2
B Pan CK 1
(Mixture of AE1/AE3 3
3
Cell Blocks
Cytopathology

& CAM5.2) 4 3
6 5 4 6
5 4

7 6
8 7 5
7

1 2 1 2 2
C LCA(CD45) 1
[or PGM1(CD68)
3
3
or mixture of LCA 4 3
6 5 4 6
& PGM1] 4
5
7 6
8 7 5
7

1 2 1 2 2
D Calretinin 1
3
3
4 3
6 5 4 6
5 4

7 6
8 7 5
7

1 2 1 2
E 1
WT-1 3 3
4 3
6 5 4 6
5 4

7 6
8 7 5
7
(continued)

SCIP
Vinod B. Shidham,

of effusion fluids

approach
vshidham@med.wayne.edu
MD, FRCPath, FIAC

Immunocytochemistry
Cell Blocks Vinod B. Shidham,
In MD, FRCPath, FIAC
Cytopathology vshidham@med.wayne.edu

Shidham & Atkinson

Ch 5. ‘Cytopathologic
Diagnosis of Serous Fluids’
Elsevier (W. B. Saunders
Company)
Cell Blocks Vinod B. Shidham,
In MD, FRCPath, FIAC
Cytopathology vshidham@med.wayne.edu

Shidham & Atkinson

Ch 5. ‘Cytopathologic
Diagnosis of Serous Fluids’
Elsevier (W. B. Saunders
Company)
 Cell Blocks Vinod B. Shidham,
In 35
MD, FRCPath, FIAC
Cytopathology
A. Vimentin vshidham@med.wayne.edu
Non-immunoreactive NC
NC
10X 40X

B. Pan-cytokeratin
Immunoreactive
NC
NC
10X 40X
SCIP
C. LCA (CD45)
Non-immunoreactive
approach
NC
NC
10X 40X

D. Calretinin
Non-immunoreactive
(Inset {2}- RM
Mesothelial cell NC
immunoreactive
nuclear-cytoplasmic) RM NC
Metastatic colonic
10X 40X
adenocarcinoma,
E. WT-1
Non-immunoreactive RM (peritoneal fluid).
HE (Arrow 2 with inset:
stained Mesothelial cell- NC
cell block immunoreactive RM NC
section nuclear-cytoplasmic) 10X 40X

NC
F. CDX2
Immunoreactive
nuclear NC
NC
40X 10X 40X 100X
Cell Blocks Vinod B. Shidham,
In MD, FRCPath, FIAC
Cytopathology vshidham@med.wayne.edu

Study cases
 Cell Blocks Vinod B. Shidham,
In MD, FRCPath, FIAC
Cytopathology vshidham@med.wayne.edu

A B C D
A. Pap smear dx – LSIL,
B. H&E cell block sections,
C. p16 stained cell block sections,
D. biopsy showing CIN II-III
 Cell Blocks Vinod B. Shidham,
In MD, FRCPath, FIAC
Cytopathology vshidham@med.wayne.edu

A B C D

A. Pap smear dx – HSIL ,

B. H&E cell block section containing “microbiopsies”,
C. p16 stained cell block section showing true nuclear positivity,
D. biopsy showing invasive squamous cell carcinoma .
 Cell Blocks Vinod B. Shidham,
In 14
MD, FRCPath, FIAC
Cytopathology vshidham@med.wayne.edu

2
1

a Calretinin b Calretinin
Calretinin immunoreactivity pattern (epithelioid mesothelioma, pleural fluid).
Mesothelioma cells (arrow in a) show nuclear (arrowhead 1) immunoreactivity usually with
cytoplasmic immunostaining (arrowhead 2) imparting the so called „fried-egg‟ appearance.
 Cell Blocks Vinod B. Shidham,
In MD, FRCPath, FIAC
Cytopathology vshidham@med.wayne.edu

Cell block- Fat

pad aspiration

a b

Positive control

c d
Congo red stained 10 micron thick sections: Orange yellow birefringence under polarized light.
The color changes to apple green when the axis of polarizer (blue arrows) is changed by 90 degree
 ‘Subtractive coordinate immunoreactivity pattern’
(SCIP) in cell block sections
Cell Blocks Vinod B. Shidham,
In 38
MD, FRCPath, FIAC
A. Vimentin vshidham@med.wayne.edu
Cytopathology
Non-immunoreactive
(Mesothelial &
inflammatory cells are
immunoreactive)
20X 40X

B. CD68 (PGM1)
Metastatic
Non-immunoreactive
(inflammatory cells are
mammary adenocarcinoma,
immunoreactive) (pleural effusion).
20X 40X

RM
C. Calretinin
Non-immunoreactive
(Rare mesothelial
cell [blue arrow] is
immunoreactive
nuclear-cytoplasmic)
20X 40X

D. BerEP4
Immunoreactive NC SCIP
20X 40X approach
NC (continued)
E. Estrogen
receptors
NC
Immunoreactive

20X 40X
 Cell Blocks Vinod B. Shidham,
In MD, FRCPath, FIAC
Cytopathology vshidham@med.wayne.edu

H&E CD177 vimentin

GIST
WS cytokeratin S-100 protein SMA
 Cell Blocks Vinod B. Shidham,
In MD, FRCPath, FIAC
Cytopathology vshidham@med.wayne.edu

Cell block section of FNA of From: Çoban S et al. BMC Cancer 2004, 4:89
Open access article from:
Hepatocellular Carcinoma http://www.biomedcentral.com/1471-2407/4/89
 Cell Blocks Vinod B. Shidham,
In MD, FRCPath, FIAC
Cytopathology vshidham@med.wayne.edu

Study cases- Blocks & slides

Blocks 1-4
1. Peritoneal fluid –solitary tumor cells- with & without AV marker.
2. Peritoneal fluid – Metastatic pancreatic adenoca.
3. Liquid based cervical cytology specimen- Cell blocks A & B not cut yet.
4. Metastatic adenoca with solitary cells – cell block from scant clot
Slides (I - VI)
I. Lymphoma (serous cavity effusion)-
II. Peritoneal fluid (metastatic pancreatic carcinoma) (See with cell blocks – set #1)
III. p16 immunostaining of cervical cytology LBC specimen
IV. Metastatic non-small cell carcinoma (Compare cellularity)
V. Metastatic non-small cell carcinoma (pleural fluid) (SCIP approach)
VI. Compare H&E stained sections of cell blocks.
Cell Blocks Vinod B. Shidham,
In MD, FRCPath, FIAC
Cytopathology vshidham@med.wayne.edu

Q/A
Cell Blocks Vinod B. Shidham,
In MD, FRCPath, FIAC
Cytopathology vshidham@med.wayne.edu

Peer-reviewed,
open access,
teaching material with many pictures.
Hard copy and online availability.
Opportunity for frequent updates
 Cell Blocks Detroit Vinod B. Shidham,
In MD, FRCPath, FIAC
Cytopathology vshidham@med.wayne.edu

After June 1, 2010

vshidham@med.wayne.edu

Milwaukee Art Museum

vshidham@mcw.edu