Labiaplasty Surgeon Locations
Labiaplasty Before and After Photo Gallery
Videos - Meet Our Surgeons
Labiaplasty Patient Testimonials
Recovering from Surgery
Costs and Financing
Frequently Asked Questions (FAQ)
Before & After Photos -Dr. Stern, FL & VA
Before & After Photos - Dr. Stanton, CA
Before & After Photos - Dr. Goodman, CA
Before & After Photos - Dr. Gonzalez, KS
Before & After Photos - Dr. Placik, IL
Before & After Photos - Dr. Kolb, GA
Before & After Photos - Dr. Hardwick-Smith, TX
Before & After Photos - Dr. Rosenfield, OR
Before & After Photos - Dr. Jacobson, CT
Before & After Photos - Dr. Penmetsa, NY
Before & After Photos - Dr. Jason, NY
Before & After Photos - Dr. Alinsod, CA
Before & After Photos - Dr. Ekstrom, MA
Before & After Photos - Dr. Canales, CA
Before & After Photos - Dr. Chambers, NV
Enduring Constant Pain And Discomfort From Enlarged Labia
Vaginoplasty Before and After Photos
Vaginoplasty Frequently Asked Questions
Vaginoplasty Patient Success Story
Vaginoplasty Surgery Success
Labiaplasty and Vaginoplasty Combination Surgery
Labia Vagina Combination Surgery Before and After Photos
Combination Labiaplasty, Vaginoplasty, Clitoral Unhooding Patient Success Story
Female Sexual Enhancement Surgery
Patient Case Studies
Labiaplasty Patient Clinical Study
Selecting a Surgeon
What to Expect Before and After Your Surgery
Where to Stay
General Surgery FAQ
Tummy Tuck (Abdominoplasty)
Patient Resources Specific to Health, Diseases and Conditions Affecting Women and Girls
Women's Health Articles
Women's Health Books
Mucin Immunohistochemistry In The Diagnosis
And Mapping Of Extramammary Paget’s Disease
R. F. Smith, B. H. Stern, A. A. Smith, *
School of Nursing, Barry University,
Miami Shores, Florida, USA
Cosmetic Surgery, P.A., Ft. Lauderdale, Florida, USA
School of Graduate Medical Sciences, Barry University, Miami Shores, Florida,
Received: July 26, 2007; Accepted: November 23, 2007
Extramammary Paget’s disease (EMPD) is a rare skin cancer of the
genital region in which cancer cells with enlarged nuclei and pale cytoplasm
are scattered singly in the affected epidermis. These cancer cells, called
Paget cells, contain mucin, which is never found in normal epidermis.
The oligosaccharide side chains of Paget cell mucin end with sialic acid.
Sialic acid is easily detected by zirconyl haematoxylin or alcian blue.
The other sugars in the oligosaccharide chains can be detected by the
periodic acid-Shiff reaction. Rarely, the diagnosis of EMPD is complicated
by the absence of mucin from the Paget cells. We have examined such an
atypical case. The oligosaccharide side chains, including the sialic acids,
are absent. In both this case and a typical case, the Paget cells contain
epithelial membrane antigen mucin (MUC1) core protein and usually contain
gastric surface-type mucin (MUC5AC) core protein, which can be stained
by antibodies. Since neither core protein is found in normal epidermis,
epithelial membrane antigen core protein may be the most reliable diagnostic
marker for extramammary Paget’s disease. In both the atypical case
and the typical case of Paget’s disease, some cells that look like
keratinocytes contain mucin core proteins. These may be incipient Paget
cells. We suggest that using the epithelial membrane antigen core protein
as a marker for the true extent of extramammary Paget’s disease
could facilitate complete excision and reduce the rate of recurrence.
Keywords: apomucin • epithelial membrane
antigen • extramammary Paget’s disease • mucin •
core protein • MUC1 • MUC5AC • Paget cells
Extramammary Paget’s disease (EMPD) is a rare epidermal carcinoma
that most often appears in the anogenital region . It resembles Paget’s
disease of the nipple in appearing as isolated Paget cells or small groups
of Paget cells rather than as a continuous mass [2, 3].
Typical Paget cell morphology includes a large nucleus and pale cytoplasm.
Paget cells usually contain sialomucins [4, 5]. The presence of sialomucin
is one way of distinguishing malignant Paget cells from benign Toker cells
 and from the malignant cells of Bowen’s disease [1, 7, 8]. All
three cell types appear as groups of 1–50 large cells with enlarged
nuclei and pale cytoplasm in H&E or trichrome preparations; they can
be confused if the diagnosis ismade on the basis of morphology alone.
An immunohistochemical re-evaluation of morphological diagnoses of extramammary
Paget’s disease, Bowen’s disease and superficial spreading
malignant melanoma found a 5% error rate in the original diagnoses .
The risks of such a mistake are serious: Toker cells are a common benign
anomaly [6, 10, 11], and Bowen’s disease can usually be treated
with topical chemotherapy alone [12, 13], but EMPD usually requires surgery
[14, 15] or prolonged radiotherapy .
Sialomucins are easily stained with zirconyl haematoxylinor alcian blue.
All mucins are stained by the periodic acid Schiff (PAS) reaction.
The occasional absence of mucin in EMPD has led to the suggestion that
mucin staining should be supplemented by at least one immunohistochemical
stain in all cases of suspected EMPD [17, 18]. The presence of cytokeratin
7 usually distinguishes EMPD from Bowen’s disease [19, 20], but
not from Toker cells .
The recent availability of antibodies to human mucin core proteins has
led to a search for specific mucin markers to distinguish EMPD from similar
skin lesions and to determine the extent of EMPD. Mucous neck cell-type
mucin (MUC6) has never been found in Paget cells [5, 22, 23]. Intestinal
type mucin (MUC2) has only rarely been found in Paget cells [5, 23]. Gastric
surface-type mucin (MUC5AC) is often found in EMPD [5, 22, 23].
Epithelial membrane antigen (EMA), also known as episialin or MUC1, has
the chemical structure of a mucin, but it is normally a transmembrane
glycoprotein rather than a secreted glycoprotein [24, 25, 26]. Paget cells
usually contain sialylated intracellular EMA in both extramammary and
mammary Paget’s disease . EMA is absent from Toker cells [10,
11]. EMA is weakly expressed in Bowen’s disease, and it is usually
confined to the cell membrane [27, 28]. Unlike the sialylated EMA usually
found in EMPD, the EMA found in Bowen’s disease usually has little
or no sialic acid and does not stain with Alcian blue .
Rarely, the diagnosis of extramammary Paget’s disease is complicated
by the absence of sialomucins from the Paget cells . Finding a case
of non-mucin-secreting EMPD led us to ask if the mucin core proteins might
be present without their oligosaccharide side chains.
Materials and methods
This protocol was approved by Barry University’s Institutional Review
Slides of formalin-fixed paraffin-embedded sections of two cases of EMPD
of the vulva were obtained from the Co-operative Human Tissue Network.
Several patients undergoing cosmetic surgery donated their tissues, which
served as normal controls. Pieces of labia minora from two patients, pieces
of perineal skin from two female patients and a fragment of skin from
the medial thigh from one male patient were fixed in 10% formalin, embedded
in paraffin, and cut at 7 ?m. A slide from each
case was stained with Ehrlich’s haematoxylinand eosin Y, zirconyl
haematoxylinand methylene green , alcian blue at pH 2.5 and kernechtrot
 and the PAS reaction .
Two sections from each case of EMPD were de-paraffinized, heated to 95_C
for 40 min. in 0.10 M pH 6.0 citrate buffer to retrieve the antigen, treated
with 0.3% hydrogen peroxide in methanol for 30 min. to quench endogenous
peroxidase activity, followed by 1.5% normal horse serum (Vector) in PBS
for 20 min., incubated 6 hr at 23°C in a 1:200 dilution of mouse monoclonal
antibodies (clone MAB 2011, Chemicon, Temecula, CA) to MUC5AC in 1.5%
normalhorse serum, washed in PBS, incubated 30 min in 0.5% biotinylated
horse anti-mouse immunoglobulin (Vector) in PBS, washed in PBS, treated
with avidin-conjugated horseradish peroxidase (Vector ABC kit) for 30
min. and washed in PBS. One section was stained with Vector’s Nova
Red for 5 min. and counterstained in haematoxylin; the second section
was stained with Vector’s VIP for 2 min. and counterstained in kernechtrot
. One section from each of the five normal control tissues was incubated
in the same way and stained with Nova Red. All sections were dehydrated,
cleared and mounted in Permount (Fisher, Atlanta, GA). A control section
from each case of EMPD was treated similarly with the omission of the
mouse antibodies to MUC5AC and stained with Nova Red.
The mouse antibodies to MUC5AC were raised against a synthetic polypeptide
of the consensus tandem repeat of human MUC5AC core protein: threonine-threonine-serinethreonine-
threonine-serine-alanine-proline [33, 34, 35].
One section from each case was de-paraffinized, blocked with 0.3% hydrogen
peroxide in methanol for 30 min. followed by 1.5% normal horse serum (Vector,
Burlingame, CA) in PBS for 20 min., incubated 30 min. at 23°C in a
1:100 dilution of mouse monoclonal antibodies (clone ZCE 113, Zymed, San
Francisco, CA) to human EMA in 1.5 % normal horse serum (Vector), washed
in PBS, incubated 30 min. in 0.5 % biotinylated horse antimouse immunoglobulin
(Vector), treated with avidin-conjugated horseradish peroxidase (Vector
ABC kit) for 30 min., stained with Vector’s Nova Red for 15 min.,
counterstained in haematoxylin, dehydrated, cleared and mounted in permount.
One section from each of the five normal control tissues was treated the
same way. Another section from each case of EMPD was treated similarly
with the omission of the mouse antibodies to EMA.
The nuclei stain with methylene green.The mouse antibodies to EMA were
raised against cream from human milk, which contained membrane bounded
fat globules. Thus, the antibodies were raised against the glycosylated
Both cases of EMPD showed many cells with typical Paget cell morphology
(enlarged cells with an enlarged nucleus and pale cytoplasm) in the epidermis
of sections stained with haematoxylinand eosin The Paget cells in slides
from the typical case stained with zirconyl haematoxylin (Fig. 1), strongly
with the PAS reaction, and brilliantly with alcian blue. The Paget cells
in slides from the other case did not stain at all with zirconyl haematoxylin
(Fig. 2) or alcian blue, establishing the absence of sialomucins. They
did not stain with the PAS reaction, showing the absence of any kind of
mucin. Normal keratinocytes in both cases of EMPD and in the control tissues
did not stain with zirconyl haematoxylinor alcian blue, but they did stain
faintly with PAS.
Sialomucin in typical Paget cells stains purple with zirconyl haematoxylin.
Paget cells in mucin-negative extramammary Paget’s disease fail
to stain with zirconyl haematoxylin. The nuclei and rough ER stain
with methylene green.
Many Paget cells in the typical sialomucin-positive (Fig. 3) and the atypical
sialomucin-negative (Fig. 4) case reacted with monoclonal antibody to
MUC5AC mucin core polypeptide. In both cases, a few cells that did not
have a Paget cell morphology reacted with antibody to MUC5AC polypeptide.
Controls slides of EMPD with the antibody omitted did not stain.
Paget cells in mucin-positive extramammary Paget’s disease contain
MUC5AC core protein (red–brown reaction product).
Many Paget cells in mucin-negative extramammary Paget’s disease
contain MUC5AC core protein (red–brown reaction product). A
morphologically normal cell (arrow) also contains MUC5AC core protein
(red–brown reaction product).
Live keratinocytes in the control tissues never stained
with antibody to MUC5AC, but light background staining was often seen
in the stratum corneum (Fig. 5). Rarely, a few sebaceous glands stained
Almost all Paget cells in both cases reacted strongly with monoclonal
antibody to EMA (Figs. 6–7). In both cases, a few cells that did
not have Paget cell morphology reacted with antibody to EMA. Control slides
of Paget’s disease with the antibody omitted did not stain.
No cells in the control skin from the thigh or from the perineum reacted
with antibody to EMA. Sebaceous glands in both control labia minora reacted
strongly with antibody to EMA; keratinocytes did not react (Fig. 8). Even
the epithelium around a microscopic condyloma in one labium minus did
not react with antibodies to EMA. (The patient was referred to her gynaecologist
for treatment of the underlying human papilloma virus infection.)
No cells in a control labium minus stain for MUC5AC core protein.
Melanin in the stratum basale is grey–brown unlike the red–brown
of Nova Red. The sebaceous gland (arrow) is unstained.
Mucin-positive extramammary Paget's disease. The Paget cells and two
morphologically normal cells (arrows) contain epithelial membrane
antigen (red-brown reaction product).
Mucin-negative extramammary Paget’s disease. The Paget cells
and two morphologically normal cells (arrows) contain epithelial membrane
antigen (red–brown reaction product).
Normal labium minus incubated with antibody to epithelial membrane
antigen. Note the difference between the red–brown stain in
the sebaceous gland and the grey–brown melanin in the stratum
It is common knowledge that normal epidermis does not contain sialomucins
[1, 4, 5]. Where normal skin has been used as a control, neither MUC5AC
and EMA core proteins have been noticed in the epidermis . This study
searched for MUC5AC and EMA core proteins in normal epidermis and found
neither. The staining of normal keratinocytes with PAS is presumably due
the presence of glycogen. It is notable that the non-mucin-secreting Paget
cells contained less glycogen than the surrounding normal keratinocytes.
We started our staining of MUC5AC core protein by following Yoshii et
al. . Our antibody (from Chemicon) and their antibody (from Novocastra)
were both monoclonal antibodies to synthetic polypeptides with the same
amino acid sequence.
Nevertheless, our antibody bound more quickly and less specifically than
theirs, forcing us to use a shorter incubation time to eliminate background
staining. Liegl et al. , using twice our antibody concentration, found
that their antibody (from Eubio) bound to Paget cells in less than half
their cases. The wide variation in the experience of different research
groups suggests that MUC5AC core protein is not a reliable marker for
the diagnosis of extramammary Paget’s disease. The failure of some
Paget cells in our cases to bind MUC5AC antibody also suggests that MUC5AC
core protein would not be a reliable marker for mapping the extent of
The staining of mucin-negative Paget cells with antibodies to EMA, that
is MUC1, raised against human milk fat globules confirms previous observations
[36, 37] that many antibodies generated against the complete glycoprotein
bind to the core protein. It is believed that the most antigenic portion
of the complete glycoprotein is the 20 amino acid tandem repeat polypeptide
. (It has been suggested that Paget cells may be malignantly transformed
Toker cells [10, 11]. The appearance of antigens typical of Paget cells
in cells that resemble keratinocytes rather than Toker cells, suggests
that Paget cells arise from keratinocytes rather than Toker cells.) Although
neither sialic acid residues nor the distribution pattern of EMA core
protein are infallible markers for the diagnosis of EMPD, each is useful
when used separately and, when used together, greatly enhance the accuracy
of diagnosis. The almost universal presence of EMA in Paget cells (5,
22) makes it a good marker for determining the extent of the disease.
It would be especially useful for scouting biopsies  and Mohs surgery.
The presence of EMA and MUC5AC core protein in a few cells that did not
have the morphology of Paget cells suggests the possibility that they
may be incipient Paget cells. If so, (extramammary Paget’s disease
arises from many cells and) removal of all cells with Paget cell morphology
often leaves some incipient Paget cells behind. This would account for
the high rate of recurrences after surgical excision of EMPD [40, 41,
42]. If our surmise is correct, mapping the margins of extramammary Paget’s
disease with antibodies to EMA should reduce the rate of recurrence after
(By detecting incipient Paget cells before they attain Paget cell morphology,
immunohistochemical detection of epithelial membrane antigen could make
surgical margins more accurate and sharply reduce the rate of recurrence.)
The authors thank the Co-operative Human Tissue Network for the two cases
of extramammary Paget’s disease. The authors also thank the five
patients who consented to the research use of tissue removed during cosmetic
surgery. They thank Barry University School of Graduate Medical Sciences
for providing all of the financial support of the work. The authors have
no financial interest in this work.
© 2007 The Authors
Journal compilation © 2007 Foundation for Cellular and Molecular
Medicine/Blackwell Publishing Ltd
Contact Dr. Stern @ (954) 981-3223
For more information: email@example.com