Diagnosis of Metastatic Neoplasms
An Immunohistochemical Approach
Murli Krishna, MD
NofContext.It is important to determine the type and/or site
origin of metastatic tumors for optimal clinical manage-
ment.
Objective.To summarize the use of currently available
immunohistochemical markers in the evaluation of meta-
static tumors.
Data Sources.Review of relevant literature on immu-
nohistochemical evaluation of tumors and the authors
personal experience.
D etermination of the type and origin of metastatic
tumors is an important and potentially challenging
area in pathology. The site of origin is best determined by
correlating clinical and pathologic findings; however, in the
absence of a clinically known or suspected primary site,
morphologic and immunohistochemical evaluations are
key to determining the tumor lineage and origin.1 While
routine microscopy may reveal characteristics that are
diagnostic or suggestive of the lineage and/or origin (eg,
mucin, melanin, keratin, clear cell change, bile production),
absence of morphologically distinctive features often
necessitates the use of immunohistochemistry, particularly
in poorly differentiated malignancies. Even if a basic tumor
lineage is apparent on routine hematoxylin-eosin stain,
confirmation of a site of origin (eg, lung, colon) may be
clinically important. Tumor characterization is often
performed on limited tissue samples obtained by cytologic
specimens or core biopsies, underscoring the need for a
strategic approach to use of immunohistochemistry.2
This review outlines the use of immunohistochemistry
in the diagnosis of metastatic tumors, both in the context
of confirming a clinically suspected site of origin as well as
a tumor of unknown origin. It includes review of relevant
current markers that may be used and of differential
diagnostic considerations based on presence or absence of
staining. It must be emphasized that no marker is entirely
specific; thus, use of a carefully constructed panel of
markers is important. Immunohistochemistry and other
ancillary techniques should not diminish the role of
Accepted for publication September 3, 2009.
From the Department of Pathology, Mayo Clinic Florida, Jacksonville,
Florida.
The author has no relevant financial interest in the products or
companies described in this article.
Reprints: Murli Krishna, MD, Department of Pathology, Mayo Clinic
Florida, 4500 San Pablo Road, Jacksonville, FL 32224 (e-mail: krishna.
murli@mayo.edu).
Arch Pathol Lab MedVol 134, February 2010
Conclusions.Immunohistochemistry is an important
ancillary technique for evaluation of metastatic tumors and
should be used in the context of routine morphology and
clinical information. While a single marker may be used to
support a known or suspected site of origin, a carefully
constructed panel is strongly recommended, particularly
for tumors of morphologically uncertain lineage or origin.
(Arch Pathol Lab Med. 2010;134:207215)
clinical information and careful gross and routine histo-
logic evaluation.
SCREENING MARKERS FOR LINEAGE DETERMINATION
The first step in the evaluation of a metastatic tumor is
the determination of tumor lineage (eg, epithelial, mesen-
chymal, melanocytic). Markers for lineage determination
should include a panel with expected positive and
negative staining in the different lineages under consid-
eration. For example, to confirm the diagnosis of
metastatic carcinoma, a reasonable panel may consist of
epithelial markers such as keratins (expected positive) and
melanocytic markers such as S100 and HMB-45 (expected
negative).
Epithelial Markers
Markers useful in confirming epithelial differentiation
include low-molecular-weight cytokeratins recognized by
CAM 5.2 and 35BH11 and broad spectrum cytokeratins
(pankeratin) recognized by AE1/AE3 antibody. Cytoker-
atin (CK) expression may sometimes be seen in none-
pithelial tumors such as melanomas and sarcomas, and
CK alone does not distinguish a carcinoma from meso-
thelioma.3,4 This underscores the utility of a panel of
immunostains. Conversely, absence of CK does not
always exclude a carcinoma; for example, adrenal cortical
carcinomas are often negative for CK, and hepatocellular
carcinomas (HCCs) are often negative for pankeratin.
Epithelial membrane antigen (EMA) may be used in
conjunction with keratins; however, it is not specific to
epithelial differentiation and may be expressed in normal
and neoplastic hematolymphoid cells including plasma
cells and anaplastic large cell lymphomas.5 Epithelial
membrane antigen is usually absent in medullary thyroid
carcinoma and adrenal cortical carcinoma. MOC-31 stains
most adenocarcinomas, and CK5/6 stains most squamous
cell carcinomas and mesotheliomas.
Metastatic Neoplasms, ImmunohistochemistryKrishna 207
 Mesenchymal Markers
There is no reliable positive screening marker for
confirming mesenchymal differentiation. While vimentin
has been used to support a mesenchymal lineage in
tumors, it is coexpressed with keratins in some carcinomas
and mesothelioma, and it is also expressed in melano-
mas.6,7 Therefore, to confirm mesenchymal differentiation,
it is important to exclude other lineages by incorporating
appropriate markers in the panel together with vimentin.
Melanoma Markers
S100 is the most sensitive screening marker for primary
and metastatic melanomas (.95%).8,9 However, S100 is not
specific for melanomas and can also be seen in some
mesenchymal and epithelial tumors.10 Both nuclear and
cytoplasmic staining should be present for the staining
result to be interpreted as positive. To confirm the presence
of melanoma, S100 should be combined with 1 or more
markers with higher specificity, such as HMB-45, Melan-
A/MART-1, tyrosinase, or microphthalmia transcription
factor protein.1115 Compared to S100, these markers are less
sensitive for the diagnosis of melanomas and stain less than
10% of desmoplastic/spindle cell melanomas.16 Melan-A
also stains adrenal cortical neoplasms and other steroid-
producing tumors.17 Melan-A and HMB-45 commonly
stain perivascular epithelioid cell tumors (PEComas), with
less frequent staining for tyrosinase and microphthalmia
transcription factor protein.18
Mesothelioma Markers
Distinguishing mesothelioma from metastatic adenocar-
cinoma is important and can be successfully achieved by
using a panel of markers for both tumor types. There are no
consensus-based guidelines on the choice of markers.
Positive markers that are useful for mesothelioma but not
entirely specific include calretinin, CK5/6, and Wilms tumor
gene product (WT1). Calretinin is a sensitive marker,
frequently expressed (.80%) in epithelial and sarcomatoid
subtypes.19 Staining is both nuclear and cytoplasmic. Other
tumor types may also be immunoreactive for calretinin,
including sex cordstromal tumors, adrenal cortical tumors,
and a minority of adenocarcinomas and squamous cell
carcinomas.19 Cytokeratin 5/6 and WT1 both stain more
than 75% of epithelial mesotheliomas but are frequently
absent in sarcomatoid mesotheliomas.20 Cytokeratin 5/6 also
stains most squamous cell carcinomas, and WT1 stains most
ovarian serous carcinomas, Wilms tumors, and desmoplas-
tic small round cell tumors. Other positive markers that can
be used for mesothelioma include HBME-1, thrombomodu-
lin, mesothelin, D2-40, and podoplanin.21,22
Markers for adenocarcinoma include carcinoembryonic
antigen (CEA), MOC-31, thyroid transcription factor 1
(TTF-1), Ber-EP4, B72.3, and CD15 (Leu-M1). These
markers have a high specificity for adenocarcinomas
relative to mesotheliomas but may also variably stain
squamous cell carcinomas.19 Thyroid transcription factor 1
is highly specific for adenocarcinoma of lung if thyroid
carcinoma is excluded, and both CD15 and TTF-1 have
lower sensitivity compared to the other markers.
Hematolymphoid Markers
Leukocyte common antigen (CD45) is a useful screening
marker for hematolymphoid neoplasms, with a sensitivity
and specificity greater than 95%. Rare examples of
208 Arch Pathol Lab MedVol 134, February 2010
undifferentiated or neuroendocrine tumors have been
reported to be positive for CD45. Additional markers that
could be used include terminal deoxynucleotidyl trans-
ferase, CD3, CD20, CD30, CD43, CD21, or CD23 and
myeloperoxidase, depending on the morphologic fea-
tures.23,24 Subclassification of metastatic hematolymphoid
malignancies is beyond the scope of this review.
Markers for Germ Cell Tumors
Placental alkaline phosphatase is a membrane-bound
isoenzyme that is produced by placental syncytiotropho-
blasts and many neoplasms.2527 Among germ cell tumors,
it is expressed in nearly all (.95%) seminomas and
embryonal carcinomas, most yolk sac tumors, and
variably in choriocarcinomas. Placental alkaline phospha-
tase is also expressed in some nongerm cell carcinomas,
including serous carcinomas, and is therefore not specific
for germ cell tumors.27,28
a-Fetoprotein is an oncofetal glycoprotein that is a
useful marker for yolk sac tumors and hepatocellular
carcinomas. Most yolk sac tumors are positive with this
marker, although staining may be patchy.27 Pure semino-
mas are negative for a-fetoprotein, and positivity in mixed
germ cell tumors identifies yolk sac differentiation.
Human chorionic gonadotropin is a glycoprotein
composed of a and b subunits, the latter produced by
syncytiotrophoblasts.29 It is therefore a useful marker for
choriocarcinoma and also identifies multinucleated syn-
cytiotrophoblastic cells in seminomas, embryonal carci-
nomas, and yolk sac tumors.
Other markers that are useful include OCT3/4 and
CD30. OCT3/4 is a sensitive and specific marker for
seminoma and embryonal carcinoma.30 CD30 stains most
embryonal carcinomas and can be used as part of a panel
of markers to distinguish metastatic embryonal carcinoma
from somatic carcinoma.31
DETERMINATION OF ORGAN OR TISSUE OF ORIGIN
FOR METASTATIC CARCINOMAS
Determination of site of origin of carcinomas is often
possible and necessary for adequate clinical management
of patients. After a metastasis is determined to be a
carcinoma on the basis of screening immunostains, a panel
of tissue- or organ-specific markers can be used in an
attempt to determine or suggest the origin. Most tumor
or organ-specific markers may variably react with other
tumor types also; hence, for metastatic tumors of
unknown origin, the use of a panel of markers is strongly
encouraged. For example, to determine whether an
adenocarcinoma is of lung or colonic origin, a combination
of CK7, CK20, TTF-1, and CDX-2 markers is likely to
establish a higher degree of certainty than TTF-1 alone.
Table 1 summarizes the key markers for commonly
encountered carcinomas and mesothelioma.
Cytokeratin Profile
A combination of CK7 and CK20 has been shown to
provide discrimination between subsets of carcinomas
and is often used as part of a panel to establish a primary
site (Table 1). Cytokeratins 7 and 19 may be used to
distinguish between cholangiocarcinoma (CK7+/CK19+)
and hepatocellular carcinoma (CK72/CK192). AE1/AE3
stains most carcinomas; however, hepatocellular carcino-
ma is commonly negative, and AE1/AE3 may be used as
part of a panel to distinguish between HCC and a
Metastatic Neoplasms, ImmunohistochemistryKrishna
 Table 1. CK7/20 Profile and Other Relevant Markers Markers for Lung Carcinoma
for Select Carcinomas and Mesothelioma Thyroid transcription factor 1 is a nuclear transcription
CK7/20 Profile and Tumors Markers and Immunoreactivity factor that regulates gene expression in the thyroid, lungs,
+ +
and diencephalon during embryogenesis. It is expressed by
CK7 /CK20
most adenocarcinomas (72%), large cell neuroendocrine
Urothelial carcinoma Uroplakin+,
carcinomas (75%), and small cell carcinomas of the lung
thrombomodulin+, p63+,
CK5/6+ (.90%) (Figure 1, A and B). Squamous cell and large cell
Pancreatic ductal CEA+, CA19-9+, carcinomas are less frequently positive for TTF-1, in 7% and
adenocarcinoma CDX-2+/2 20% of cases, respectively.32 While most pulmonary small
Ovarian mucinous carcinoma CDX-2+/2 cell carcinomas are positive for this marker, TTF-1 may also
Adenocarcinoma of bladder Thrombomodulin+, stain small cell carcinomas from other sites, including those
CDX-2+/2
Gastric adenocarcinoma
arising from prostate, urinary bladder, and uterine cervix.33
(subset) Other markers for lung primary tumors include napsin
Cholangiocarcinoma (subset) A, which stains most pulmonary adenocarcinomas, and a
CK7+/CK202 novel marker, ES1.32 These markers are currently not
Lung adenocarcinoma TTF-1+, CEA+, MOC-31+, widely used.
CK5/62
Lung small cell carcinoma TTF-1+, Chr+, Syn+, CK202 Markers for Thyroid Carcinomas
Breast carcinoma ER/PR+, GCDFP+, Thyroglobulin is a marker that is highly sensitive and
Mammaglobin+, CEA+ specific for follicular and papillary thyroid carcinomas,
Ovarian serous carcinoma WT1+, ER/PR+, mesothelin+,
CEA2
staining more than 95% of these tumors. Immunoreactiv-
Endometrial adenocarcinoma Vimentin+, ER/PR+, CEA2 ity is also maintained in these tumors at metastatic sites.34
Cholangiocarcinoma CEA+, CK19+, CA19-9+, Thyroid transcription factor 1 is also a useful marker for
CDX-2+/2, Hep Par 12 thyroid tumors, being expressed in more than 90% of
Thyroid carcinoma TTF-1+, Thyr+ (follicular, follicular cellderived tumors and medullary carcino-
papillary) mas.35 As described above, TTF-1 also stains carcinomas of
TTF-1+, Thyr2, Calcitonin+,
Chr+, Syn+ (medullary) lung origin, and rarely, carcinomas from other sites. It is
Mesothelioma Calretinin+, CK5/6+, D2- therefore a reliable marker for thyroid differentiation
40+, mesothelin+, WT1+ when a lung primary tumor can be excluded, or when
CEA2, MOC-312, Ber- used in conjunction with thyroglobulin. Poorly differen-
EP42, TTF-12 tiated (insular) carcinomas are focally and/or weakly
Pancreatic carcinoma (subset)
Gastric carcinoma (subset)
positive for both thyroglobulin and TTF-1. Anaplastic
thyroid carcinomas are usually negative for both markers
CK72/CK20+ but commonly coexpress keratin and vimentin.
Colorectal adenocarcinoma CDX-2+ Calcitonin is a protein secreted by thyroid C cells and is a
Merkel cell carcinoma TTF-12, Chr+, Syn+
useful marker for medullary thyroid carcinomas (MTCs). It
CK72/CK202 can also be used to evaluate for C-cell hyperplasia
Prostate adenocarcinoma PSA+, PAP+, CEA2, associated with familial MTC.36 Although highly specific,
CK5/62
Renal cell carcinoma (CC) Vimentin+, RCC+, CD10+, calcitonin staining may be patchy in medullary carcinomas
CEA2, EMA+ and 5% of these tumors may be negative for this marker.37
Hepatocellular carcinoma Hep Par 1+, pCEA+ In tumors with no staining and still suspected to be MTC,
(canalicular), CD10+ calcitonin and calcitonin generelated peptide mRNA can
(canalicular), MOC-312, be demonstrated by in situ hybridization.38 Carcinoem-
CK192
Squamous cell carcinoma p63+, CK5/6+
bryonic antigen, chromogranin, and synaptophysin are also
Adrenal cortical carcinoma Inhibin+, calretinin+, expressed by most MTCs, while follicular and papillary
Melan-A+, vimentin+, carcinomas are negative for these markers. However, they
CEA2, EMA2 are not specific to MTC and may also stain other
neuroendocrine tumors, and thus are less helpful in the
Gastric adenocarcinoma setting of metastases. A reasonable panel to confirm
(subset)
metastatic MTC would be chromogranin, synaptophysin,
Abbreviations: CC, clear cell; CEA, carcinoembryonic antigen; Chr, TTF-1, and calcitonin (Figure 2, A through D).
chromogranin; CK, cytokeratin; EMA, epithelial membrane antigen; ER/
PR, estrogen and progesterone receptors; GCDFP, gross cystic disease Markers for Hepatic Carcinomas
fluid protein15; Hep Par 1, hepatocyte paraffin 1; PAP, prostatic acid
phosphatase; pCEA, polyclonal carcinoembryonic antigen; PSA, Hepatocyte paraffin 1 (Hep Par 1) stains a cytoplasmic
prostate-specific antigen; RCC, renal carcinoma marker; Syn, synapto- antigen and is highly sensitive for hepatocyte differenti-
physin; Thyr, thyroglobulin; TTF-1, thyroid transcription factor 1; WT1, ation, thus is useful in the diagnosis of hepatocellular
Wilms tumor gene product. carcinoma (Figure 3, A and B). Expression is particularly
strong and diffuse in well-differentiated HCC but de-
creases in higher-grade tumors. However, Hep Par 1 is not
metastatic carcinoma. Cytokeratin 5/6 is a useful marker entirely specific for hepatocyte differentiation and may
for squamous cell carcinomas, epithelioid mesotheliomas, also be seen in other carcinomas, such as gastric signet
and urothelial carcinomas. Because of overlap in expres- ring cell carcinomas.39
sion, cytokeratin stains should always be combined with Canalicular staining with polyclonal CEA (pCEA) can
more specific markers if available. be used as a specific marker for the diagnosis of
Arch Pathol Lab MedVol 134, February 2010 Metastatic Neoplasms, ImmunohistochemistryKrishna 209
Figure 1. A, Metastatic adenocarcinoma (hematoxylin-eosin). B, Thyroid transcription factor 1 immunostain is positive, confirming pulmonary
origin (original magnifications 3200).

hepatocellular carcinomas. This pattern results from canalicular pattern of staining. Another marker that is
cross-reactivity with biliary glycoprotein I and is demon- specific for hepatocyte differentiation is albumin, which is
strable in 90% of HCCs and is not seen with monoclonal produced exclusively by hepatocytes. Immunohistochem-
CEA.40 Similar to pCEA, villin and CD10 also show a ical staining for albumin is difficult to interpret because of

Figure 2. A, Metastatic medullary thyroid carcinoma, amphicrine variant, with signet ring cells (hematoxylin-eosin). Chromogranin (B), calcitonin
(C), and thyroid transcription factor 1 (D) immunostains are positive (original magnifications 3400).

210 Arch Pathol Lab MedVol 134, February 2010 Metastatic Neoplasms, ImmunohistochemistryKrishna
Figure 3. A, Metastatic hepatocellular carcinoma (A; hematoxylin-eosin). Arrows indicate bile production. B, Hepatocyte paraffin 1 immunostain
is positive (original magnifications 3400).
the abundance of this protein in the serum; however,
albumin mRNA can be demonstrated by using in situ
hybridization. Both canalicular pCEA and albumin are
more specific for HCC than Hep Par 1, but like the latter,
tumors arising at other sites with hepatoid differentiation
(such as hepatoid gastric adenocarcinomas) may also be
positive for these markers.41
A specific marker for cholangiocarcinoma is currently
not available, and a diagnosis is established by exclusion
of HCC and metastatic carcinoma. This can be achieved by
the presence of appropriate clinical and radiographic
features and the use of markers that exclude carcinomas
from other sites (eg, lung and breast).
Markers for Gastrointestinal Carcinomas
CDX-2 is a nuclear transcription factor that is used as a
marker for gastrointestinal tract adenocarcinomas and is
expressed in most (.95%) colorectal and duodenal
adenocarcinomas, with nuclear staining. Variable expres-
sion is also seen in adenocarcinomas arising in the
stomach, esophagus, pancreas, and biliary tract. Extrain-
testinal carcinomas that are frequently positive for CDX-2
include urachal adenocarcinomas arising in the urinary
bladder and ovarian mucinous carcinomas.42 CDX-2
staining may also be seen in a minority of gastrointestinal
and extragastrointestinal neuroendocrine tumors and is
therefore less specific in this setting than it is for
adenocarcinomas.43
Villin is a cytoskeletal protein associated with microvilli
of the intestine and proximal renal tubular epithelium. It is
a marker of gastrointestinal adenocarcinomas, stains most
(82%100%) primary and metastatic colonic adenocarci-
nomas, and does not appear to be decreased in metasta-
ses.23 The specificity of villin is limited by its expression in
extraintestinal adenocarcinomas including those arising in
lung, ovary, endometrium, kidney, and bladder.44 Like
pCEA, villin may also stain HCC with a canalicular
pattern.40
As with primary hepatobiliary adenocarcinomas, a
specific marker is not currently available for pancreatic
ductal adenocarcinoma. The rare acinar cell carcinoma of
the pancreas can be confirmed with stains for pancreatic
enzymes such as trypsin, chymotrypsin, lipase, and
Arch Pathol Lab MedVol 134, February 2010
elastase. It is also commonly negative for CEA, unlike
ductal adenocarcinomas.45
Markers for Breast Carcinoma
Gross cystic disease fluid protein 15 (GCDFP-15; BRST-
2), also known as prolactin-inducing protein, is a glyco-
protein present in various body fluids including saliva,
milk, and seminal fluid.46,47 It is expressed in cells with
apocrine features including those present in breast, axillary
and anogenital skin apocrine glands, salivary glands, and
Paget disease of skin.48 For a metastatic carcinoma, GCDFP-
15 has a high (.95%) specificity for breast primary tumor if
the other mentioned sites are clinically excluded. The utility
of this marker, however, is somewhat limited by a lower
(50%74%) sensitivity. Therefore, absence of staining does
not exclude a breast primary tumor.
Mammaglobin is a marker that is overexpressed in 48%
to 84% of breast carcinomas. It is more sensitive but less
specific than GCDFP-15 for diagnosis of a breast primary
tumor.23,49
Estrogen and progesterone receptors are not specific to
breast and cannot be used as primary markers to support
evidence of a breast primary tumor, but may be used in
conjunction with GCDFP-15 and mammaglobin.
Markers of Prostatic Carcinoma
Prostate-specific antigen (PSA) and prostatic acid
phosphatase (PAP) are 2 commonly used markers for
prostatic carcinoma. Essentially, all low- to intermediate-
grade prostatic adenocarcinomas stain for both markers,
with reactivity decreasing by 10% to 20% in high-grade
carcinomas.50 A combination of PSA and PAP detects most
metastatic prostatic carcinomas. Prostate-specific antigen
is more specific as a marker for prostate than PAP,
although both have also been reported in extraprostatic
sites.51,52 Prostate-specific membrane antigen is another
marker that is less widely used but demonstrates high
specificity for prostate carcinoma and better sensitivity for
higher-grade carcinomas than PSA and PAP.53 While a-
methylacyl coenzyme A racemase is useful for the
diagnosis of carcinoma in the prostate, it is not useful as
a specific marker for prostatic carcinoma, as it is expressed
in many normal tissues and nonprostatic tumors.
Metastatic Neoplasms, ImmunohistochemistryKrishna 211
Figure 4. A, Metastatic gastrointestinal stromal tumor (hematoxylin-eosin). B, CD117 immunostain is positive (original magnifications 3400).

Potential future markers for prostatic carcinoma include Renal cell carcinomas are also commonly positive for
NKX3.1 and prostein. NKX3.1 is a nuclear marker that CD10 and vimentin and negative for CEA; however, these
more frequently stains high-grade carcinoma than PSA. It markers are not specific and should always be used in
may also be expressed in some other tissues, such as conjunction with other markers.
testicular germ cells and lobular carcinoma of breast.
Prostein is a transmembrane protein with a high specific- Markers for Urothelial Carcinoma
ity for benign and malignant prostatic epithelium.54 Uroplakins (Ia, Ib, II, and III) are transmembrane
proteins that are specific to urothelial cells. A monoclonal
Markers for Renal Cell Carcinoma antibody for uroplakin III is highly specific for urothelial
Renal cell carcinoma (RCC) marker antibody is directed differentiation and expressed in urothelial carcinomas and
against a normal proximal tubular brush border glyco- Brenner tumors.57 Use of this marker, however, is limited
protein. It stains most clear cell and papillary renal cell by low sensitivity, staining up to 60% of urothelial
carcinomas, but the sensitivity decreases in high-grade carcinomas at primary sites. Metastatic urothelial carci-
carcinomas and metastatic carcinomas. Only a minority of nomas are less frequently positive for this marker.
sarcomatoid renal cell carcinomas are positive for RCC Thrombomodulin is a marker that is expressed by most
marker.23 urothelial carcinomas; however, it lacks specificity and
Staining has also been reported in some breast therefore should not be used alone as confirmatory
carcinomas and embryonal carcinoma.55 Among the evidence for urothelial differentiation. It is also expressed
subtypes of RCC, a-methylacyl coenzyme A racemase is by squamous cell carcinomas, endothelial tumors, and
a marker for papillary RCC.56 mesothelioma.58,59
Markers for Adrenal Cortical Carcinoma
Table 2. Diagnostically Relevant Markers for A specific marker for adrenal cortical carcinoma (ACC) is
Select Sarcomasa not commercially available. However, ACCs show a
Sarcoma Markers and Immunoreactivity relatively specific immunoprofile when a combination of
Leiomyosarcoma SMA+, desmin+, calponin+,
markers are used. They are commonly positive for vimentin,
caldesmon+ Melan-A, inhibin, and calretinin and negative or weakly
Pleomorphic Myogenin+, MyoD1+, positive for keratin.60,61 Adrenal cortical carcinomas may be
rhabdomyosarcoma myoglobin+, desmin+/2 positive for synaptophysin but are negative for chromogra-
Angiosarcoma CD31+, CD34+, factor VIII+, nin. When included as part of a panel, these markers are
FLI1+ effective in distinguishing ACC from the usual differential
Synovial sarcoma EMA+, CK+ (epithelial
component), CD99+ diagnostic considerations, such as pheochromocytoma,
Gastrointestinal stromal tumor CD117+, SMA+/2, hepatocellular carcinoma, and RCC. Melan-A has been
desmin+/2, S100+/2 primarily used in the diagnosis of melanoma but also reacts
Endometrial stromal sarcoma CD10+, ER/PR+ with steroid-producing cells of the adrenal cortex. Inhibin A
Malignant peripheral nerve S100+/2 identifies steroid-producing cells, and thus is a sensitive
sheath tumor
Pleomorphic liposarcoma MDM2/CDK4+
marker for ACC, ovarian granulosa cell tumor, and testicular
Malignant fibrous histiocytoma No specific marker Sertoli-Leydig cell tumor. Calretinin is a calcium-binding
Fibrosarcoma No specific marker protein that is also commonly used as a marker for
Abbreviations: CDK4, cyclin-dependent kinase 4; CK, cytokeratin; EMA, mesothelioma and ovarian sex cordstromal tumors.61
epithelial membrane antigen; ER/PR, estrogen and progesterone recep-
tors; FLI1, Friend leukemia virus integration 1; SMA, smooth muscle actin. TUMORS OF THE FEMALE GENITAL TRACT
a
Carcinoma and melanoma should be excluded with appropriate Carcinomas arising in the female genital tract have
markers. overlapping morphologic and staining characteristics, and
212 Arch Pathol Lab MedVol 134, February 2010 Metastatic Neoplasms, ImmunohistochemistryKrishna
Figure 5. Metastatic pulmonary small cell carcinoma and Merkel cell carcinoma can be distinguished with a panel including thyroid transcription
factor 1 (TTF-1) and CK20 immunostains. Small cell carcinoma (A) is positive for TTF-1 (B) and negative for CK20 (C). Merkel cell carcinoma (D) is
negative for TTF-1(E) and positive for CK20, with a paranuclear dotlike accentuation (F) (original magnifications 3400).

reliable site-specific markers are not available. A combi-
nation of markers may be used to suggest origin in the
female genital tract, and differential expression of markers
may suggest a site of origin or tumor type. Markers for
germ cell tumors are discussed separately in this review.
Arch Pathol Lab MedVol 134, February 2010
Endocervical adenocarcinomas are usually positive for
CEA but negative for vimentin and for estrogen and
progesterone receptors, in contrast to endometrial adeno-
carcinomas of endometrioid type.62 Use of p16 as a
surrogate marker for high-risk human papillomavirus
Metastatic Neoplasms, ImmunohistochemistryKrishna 213
 Table 3. Diagnostically Relevant Markers for Select
Small Round Cell Tumors (SRCTs)
SRCT Markers and Immunoreactivity
Small cell carcinoma CK , CK202, TTF-1+,a Chr+, Syn+
+
Merkel cell carcinoma CK+, CK20+, TTF-12, Chr+, Syn+
Poorly differentiated CK+/2, EMA+, desmin2, CD99+
synovial sarcoma
DSRCT CK+, desmin+, WT1+
ES/PNET CK2, S1002, desmin2, CD452,
CD99+, FLI1+
Neuroblastoma CK2, desmin2, S1002, CD452,
CD99+, Chr+, Syn+
Lymphoma CK2, CD45+, TdT+/2b
Rhabdomyosarcoma CK2, S1002, desmin+, myogenin+,
MyoD1+
Abbreviations: CK, cytokeratin; Chr, chromogranin; DSRCT, desmo-
plastic small round cell tumor; EMA, epithelial membrane antigen; ES/
PNET, Ewing sarcoma/primitive neuroectodermal tumor; FLI1, Friend
leukemia virus integration 1; Syn, synaptophysin; TTF-1, thyroid
transcription factor 1; TdT, terminal deoxynucleotidyl transferase;
WT1, Wilms tumor gene product.
a morphologic evaluation.
Positive in most pulmonary and some nonpulmonary small cell
carcinomas.
b
Lymphoblastic lymphomas are commonly Tdt+ and may be CD452.
infection to prove endocervical origin has been contro-
versial, as it may also be expressed in endometrial
carcinomas, especially endometrial serous carcinomas.63
Distinction between high-grade endometrioid and serous
carcinomas is not reliably achieved with the use of
markers because of significant overlap.64
Ovarian nonmucinous carcinomas are positive for CK7
but usually negative for CK20 and monoclonal CEA.
Mucinous carcinomas are positive for CK7, CK20, and
monoclonal CEA.65,66 Inhibin and calretinin are markers
for sex cordstromal tumors, and as mentioned above,
also stain adrenal cortical tumors. Wilms tumor gene
product (WT1) stains most serous ovarian carcinomas
but usually not breast, gastrointestinal tract, and pancrea-
ticobiliary carcinomas.67 It also commonly stains meso-
theliomas and therefore should be used as part of a panel
of markers. Other tumors that are positive for this
marker include Wilms tumor and desmoplastic small
round cell tumor.68,69 Metastatic choriocarcinomas can be
confirmed with positive staining for human chorionic
gonadotropin.
METASTATIC SARCOMAS
Sarcomas are composed of a complex and heteroge-
neous group of tumors that are classified by cell lineage.
The first step in the approach to diagnosis of these tumors
is to confirm the mesenchymal lineage, followed by an
attempt to define the subtype. Subtyping will require a
panel of markers and may not be achieved with the
currently available markers in all cases. The importance of
precise subtyping will also depend on its clinical relevance
and available therapeutic options, such as therapy with
imatinib mesylate for gastrointestinal stromal tumors
(Figure 4, A and B).70 Metastatic sarcomas are commonly
high grade, variable in histomorphology, and include
tumors with epithelioid, spindle cell, pleomorphic, and
small round cell appearance. The immunohistochemical
characteristics of a select group of sarcomas are summa-
rized in Table 2, with emphasis on reactivity with key
markers.70,71
214 Arch Pathol Lab MedVol 134, February 2010
METASTATIC SMALL ROUND CELL TUMORS
Small round cell tumors are a heterogeneous group of
tumors that contain morphologically undifferentiated
small cells with high nuclear to cytoplasmic ratio and
inconspicuous nucleoli. Considerations in the differential
diagnosis include small cell carcinoma, Merkel cell
carcinoma, Ewing sarcoma/primitive neuroectodermal
tumor, poorly differentiated synovial sarcoma, hemato-
poietic malignancies, rhabdomyosarcoma, desmoplastic
small round cell tumor, and neuroblastoma. Metastases
from these tumors are often difficult to classify without
the use of a panel of markers (Figure 5, A through F).
Typical immunoprofiles of these tumors are summarized
in Table 3.23,72
Characterization of type and origin of metastatic tumors
requires judicious use of lineage and organ-specific tissue
markers. No marker is entirely specific, and for optimal
patient care, this tool should be used in the context of the
clinical and radiographic findings and careful routine
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