Malignant melanoma of the conjunctiva

One hundred thirty-one cases of conjunctival melanoma in which biopsies had been performed were studied to determine potential factors that might affect outcome in patients with these lesions. Two groups of lesions were identified: those associated with primary acquired melanosis (melanoma with PAM, 98 cases, 74.8 per cent) and those without primary acquired melanosis (melanoma without PAM, 33 cases, 25.2 per cent). The overall mortality rate in the 131 cases was 26 per cent (34 of 131); the mortality rate due to melanoma with PAM was 25.5 per cent (25 of 98), and that due to melanoma without PAM was 27.3 per cent (9 of 33). If PAM was associated with the lesion, the presence of atypical melanocytes within the epithelium (pagetoid invasion) was a sensitive indicator of subsequent metastasis. Tumor thickness may also be useful for predicting subsequent metastases. None of the histologic parameters studied proved useful for predicting outcome in patients who had melanomas without PAM. The presence or absence of nevi had no effect on prognosis.     

Lentigo maligna and malignant melanoma in situ, lentigo maligna type.

Some authors have considered lentigo maligna to be an atypical melanocytic proliferation, whereas others have considered it to be melanoma in situ. We reviewed 50 cases of lentigo maligna. We have identified two subsets of lesions. The first has atypical melanocytic hyperplasia, which we postulate to be correctly designated lentigo maligna. The second subset has the following features in addition to the melanocytic hyperplasia: individual and nests of cells at varying layers of the epidermis, confluence of the melanocytes replacing the basilar region, uniformity of the cytological atypia, and nesting of uniformly atypical melanocytes. These lesions we designate as malignant melanoma in situ, lentigo maligna type. We are proposing that the lesions that have been termed lentigo maligna represent a spectrum of atypia and that the application of some of the traditional features for the diagnosis melanoma may permit the segregation of more and less aggressive lesions.     

Differentiating nonhigh-grade duct carcinoma in situ from benign breast lesions

This study was undertaken to determine the discriminating cytological features between nonhigh-grade duct carcinoma in situ (NHGDCIS) and benign breast lesions and to determine any histological characteristics which would influence the cytological categorization. Smears of 12 each of histologically confirmed NHGDCIS and benign breast lesions were reviewed with regard to cellularity, cell discohesion, nuclear atypia, crowding of cells, tubule formation, necrosis, and presence of bare atypical nuclei and regular bare bipolar nuclei, and statistically analyzed. Architectural pattern, presence of necrosis, and the size of the lesion assessed at histological examination were compared with the initial cytological categorization. NHGDCIS lesions showed more cell discohesion (P = 0.04), bare atypical nuclei (P = 0.05), necrosis (P = 0.03), and sparse bare bipolar nuclei (P = 0.02) than benign lesions. These differences were statistically significant. Cellularity (P = 0.8), nuclear atypia (P = 0.06), crowding of cells (P = 0.1), and tubule formation did not show a significant difference. Six (out of six lesions) with a solid architectural pattern and six (of seven) with necrosis could be cytologically categorized as suspicious or malignant. Size of the lesion did not influence this. We conclude that cell discohesion, bare atypical and bare bipolar nuclei, and necrosis are discriminating features between NHGDCIS and benign breast lesions and NHGDCIS lesions with a solid architectural pattern and necrosis are more likely to be satisfactorily categorized cytologically.     

The prevalence of carcinoma in situ in normal and cancer-associated breasts

Two hundred ninety-two human breasts were examined in toto by a subgross sampling technique with histologic confirmation. The samples consisted of 185 breasts from random autopsies, 63 cancer-containing breasts, and 44 breasts contralateral to cancer-containing breasts. The method permits the identification and enumeration of essentially all of the dysplastic, hyperplastic, and neoplastic lesions present in each breast. Emphasis was on the prevalence within each sample category of ductal carcinoma in situ (DCIS), lobular carcinoma in situ (LCIS), and epithelial proliferative lesions with severe atypia, previously termed ALA 4 and ALB 4, which correspond to the clinicopathologic entities atypical ductal hyperplasia and atypical lobular hyperplasia, respectively. Additional primary foci of DCIS (unrelated to invasive breast carcinoma, if present) were found in 52.5 per cent of cancer-containing breasts, and were seen in 47.7 per cent of contralateral and 5.9 per cent of the breasts from random autopsies. Lobular carcinoma in situ was generally seen only in association with infiltrating carcinoma, usually of the ductal type. No LCIS was seen in the breasts from random autopsies. These trends are the same if the proliferative lesions with severe atypia are included with carcinoma in situ. The numbers of lesions were also markedly greater in affected cancer-associated breasts than in affected breasts obtained from autopsies. These findings suggest that LCIS, although a rare lesion in the general population, may be a significant marker for clinical carcinoma. They support previous studies showing a small percentage of women with undetected DCIS of uncertain clinical and biological potential. The multicentric nature of preinvasive breast carcinoma is further substantiated. Finally, when the prevalence and number of lesions are considered in association with the ages of the patients, the lower prevalence of such lesions in the older patients in each sample suggests that at least some DCIS and LCIS may be dependent on a premenopausal hormonal milieu for their continuing existence.     

Cell growth and p53 expression in primary acquired melanosis and conjunctival melanoma.

AIMS: To evaluate cell growth and the pattern of p53 suppressor gene expression in atypical primary acquired melanosis (PAM) and in recurrent conjunctival melanoma. METHODS: Eighteen specimens of PAM with atypia and 24 specimens, comprising early and late lesions, from 12 patients with conjunctival melanoma were stained for the proliferating cell nuclear antigen using the PC10 antibody, and for the p53 gene product using the BP53-12-1, 1801 and DO7 clones. The immunoreactive cells were counted manually and the data evaluated statistically. RESULTS: Seven of nine PAM specimens progressing to melanoma expressed PC10. None of these lesions expressed the p53 gene product. The number of proliferating cells was higher in the late than in the early lesions of conjunctival melanoma. Four of the 12 recurrent melanomas displayed focal, but minimal, p53 expression. The proliferating cell count in the p53 positive tumours was very similar to that of the p53 negative conjunctival melanomas. CONCLUSION: Examination of the expression of proliferating cells in atypical PAM may be used as an adjunct to predict which lesions will progress to melanoma. The increase in the number of proliferating cells over time in recurrent conjunctival melanomas probably reflects more aggressive behaviour and may be used to monitor recurrence. The absence of p53 expression in PAM and minimal staining of conjunctival melanomas did not correlate with cell growth, suggesting that alterations in the p53 tumour suppressor gene are uncommon and late events in conjunctival melanoma, and that p53 expression is unlikely to be a useful prognostic indicator.     

Risk factors for breast cancer in women with proliferative breast disease

To assess the importance of various risk factors for breast cancer in women with benign proliferative breast lesions, we reevaluated 10,366 consecutive breast biopsies performed in women who had presented at three Nashville hospitals. The median duration of follow-up was 17 years for 3303 women, 1925 of whom had proliferative disease. This sample contained 84.4 per cent of the patients originally selected for follow-up. Women having proliferative disease without atypical hyperplasia had a risk of cancer that was 1.9 times the risk in women with nonproliferative lesions (95 per cent confidence interval, 1.2 to 2.9). The risk in women with atypical hyperplasia (atypia) was 5.3 times that in women with nonproliferative lesions (95 per cent confidence interval, 3.1 to 8.8). A family history of breast cancer had little effect on the risk in women with nonproliferative lesions. However, the risk in women with atypia and a family history of breast cancer was 11 times that in women who had nonproliferative lesions without a family history (95 per cent confidence interval, 5.5 to 24). Calcification elevated the cancer risk in patients with proliferative disease. Although cysts alone did not substantially elevate the risk, women with both cysts and a family history of breast cancer had a risk 2.7 times higher than that for women without either of these risk factors (95 per cent confidence interval, 1.5 to 4.6). This study demonstrates that the majority of women (70 per cent) who undergo breast biopsy for benign disease are not at increased risk of cancer. However, patients with a clinically meaningful elevation in cancer risk can be identified on the basis of atypical hyperplasia and a family history of breast cancer.     

Epidermal melanocytes adjacent to melanoma and the field change effect

The presence of a field change, affecting epidermal melanocytes in the skin surrounding melanomas, has been cited as a justification for performing radical excision of these lesions. Twenty-five consecutive re-excisions of melanoma specimens were examined and melanocytes per 100 keratinocytes counted continuously across the width of each, in order to ascertain whether melanocyte counts decreased from the centre to the periphery, as would be expected if there were a field change effect associated with the tumour. Melanocyte counts did not appear to diminish with increasing distance from the tumour site and were within the range seen in a previous study in which we demonstrated increased numbers of basal melanocytes and melanocyte atypia in sun-exposed skin from a control population. Melanocyte atypia of a mild degree was present in five of the 25 cases studied, but in no case was this greater than that observed in sun-exposed forearm skin. We therefore suggest that the 'field change effect' in epidermal melanocytes adjacent to melanoma could be a result of chronic sun exposure alone.     

Endocervical reactive atypia: a histologic-cytologic study

Of 1,500 cervical tissue specimens, 27 cases showed histologic changes of reactive glandular atypia which we defined as endocervical cells with large hyperchromatic, often irregular nuclei, which did not fulfill the criteria for endocervical adenocarcinoma. Eighteen of these 27 cases had preceding or concurrent cervico-vaginal smears. Six of these showed cells which were similar to those seen in histologic sections. The cytologic characteristics of these cells are defined. To determine if atypia is related to inflammatory-regenerative changes, 29 cases of endocervical polyps were examined, of which 11 showed histologic changes of endocervical reactive atypia; 4 showed these changes cytologically as well. Twenty-eight cases of routine hysterectomy specimens were examined, of which 2 cases showed endocervical reactive atypia, which indicated that the atypical changes were indeed reactive. Nine out of 27 cases were associated with hormonal usage. Fourteen cases were associated with squamous intraepithelial lesions or evidence of human papilloma virus. Follow-up of our 27 index cases revealed no progression to adenocarcinoma. These findings indicate that atypia, as we define it, of the endocervix can be due to inflammatory-reparative changes or possibly related to hormonal usage, and permit its separation from precursor lesions of endocervical adenocarcinoma.     

Biology of tumor progression in human melanocytes

Tumor progression in the human melanocyte system can be delineated into 6 sequential stages. The first three steps represent nonmalignant melanocyte lesions from focal proliferations of structurally normal melanocytes to lesions with architectural and cytologic atypia. Primary melanoma may be divided into radial growth phase without competence for metastasis and vertical growth phase with metastatic competence. Melanocytes isolated from normal skin, nonmalignant pigmented lesions, and melanomas and maintained in culture have properties that are characteristic for each stage of tumor progression. Cytogenetic studies revealed nonrandom chromosomal abnormalities of advanced melanomas involving chromosomes 1, 6, and 7. Recent progress in tissue culture techniques has allowed studies of growth regulation of normal and malignant cells. Six growth factor receptor-growth factor systems seem to be of biologic significance in the melanocyte system: EGF, NGF, FGF, PDGF, insulin, and beta-TGF. Monoclonal antibodies have characterized a large number of antigens on melanocytes of the various stages of tumor progression, making melanoma one of the most widely studied human tumor systems.     

Architectural and nuclear morphometrical features together are more important prognosticators in endometrial hyperplasias than nuclear morphometrical features alone

Previous studies have shown that 80-90 per cent of cases of atypical hyperplasia of the endometrium do not progress to cancer. Criteria to predict the outcome in an individual patient with hyperplasia are lacking, and hysterectomy is the usual (over)treatment in order to avoid a 10-20 per cent chance of confrontation with cancer later on in the course of the disease. A recent study has shown that using a nuclear morphometric classification rule, 15 per cent of patients without progression can be accurately separated from patients with progression. However, as it is unlikely that nuclear morphometrical features are the only morphological factors reflecting the outcome of the disease, other quantitative parameters describing the architecture of the glands have also been studied for their potential value in selecting patients who will progress to cancer. In total, 10 nuclear features and 12 glandular architectural features were studied in 39 cases of atypical endometrial hyperplasia. Among these cases, seven (18 per cent) progressed to cancer. Using linear stepwise regression analysis and discriminant analysis, the volume percentage stroma and the standard deviation of the shortest nuclear axis are the best discriminators, although the outer surface density of the glands also adds to the discriminating power. The volume percentage stroma is the best single prognosticator; this feature is highly reproducible. In total, using these combined architectural and nuclear morphometrical features, 20 of the 32 cases without progression were separated from those who subsequently progressed (62.5 per cent). This is a considerable improvement over nuclear morphometrical features alone (15 per cent separated).     

The extracellular matrix in pigmented skin lesions: an immunohistochemical study

In recent years the interaction between tumour cells and the surrounding extracellular matrix in the process of tumour development, invasion and metastasis has been a focus of interest. We studied frozen sections of nine naevocellular naevi (junctional, compound and intradermal), 40 dysplastic naevi, six pagetoid in situ melanomas and 12 superficial spreading melanomas in order to determine the expression of: the basement membrane proteins collagen type IV and laminin, the interstitial collagen types I, III and VI, and fibronectin and tenascin. An indirect immunoperoxidase technique was used. In the various stages of melanocytic tumour progression we observed: 1 loss of type IV collagen and laminin within dermal melanocytic cell nests; 2 de novo expression of basement membrane type IV collagen and increased expression of the interstitial collagen types I, III and VI, as well as tenascin and fibronectin in the dermal stroma surrounding dysplastic naevus cells and melanoma cells; 3 presence of extracellular matrix components in close association with intra-epidermally located invading atypical melanocytes. These data demonstrate the complex alterations of the composition of the extracellular matrix from bland naevi through lesions with progressive atypia to invasive melanoma. The changes described result in a molecular environment which melanocytes with an altered adhesion molecule profile are able to invade.     

Histopathological characteristics of small diameter melanocytic naevi

BACKGROUND/AIMS: The clinical definition of an atypical naevus ("dysplastic naevus" or "naevus with architectural disorder and cytological atypia of melanocytes") stresses size larger than 5 mm in diameter as a major diagnostic criterion. Because malignant melanomas and their precursors may arise in smaller lesions, a histological study of melanocytic lesions smaller than 4 mm in diameter was conducted to evaluate their histological appearance. METHODS: Two hundred and sixty one naevi smaller than 4 mm in diameter were collected and characterised by histological examination into benign naevi without architectural disorder and naevi with architectural disorder and mild, moderate, and severe atypical melanocytes according to criteria used on larger lesions. RESULTS: Small melanocytic naevi covered the same complex histological spectrum from benign naevi to severely atypical naevi when compared with larger lesions. A high proportion of small naevi (72%) exhibited features diagnostic for naevi with architectural disorder and cytological atypia. CONCLUSION: There is a discrepancy between histological and clinically defined atypical naevi. The same generally accepted criteria for the histological diagnosis of atypical naevi should be used for small melanocytic naevi in addition to large ones. Thus, small naevi exhibiting atypical features on histological examination should be categorised as atypical naevi, regardless of their small diameter.     

The approach to the patient with a difficult melanocytic lesion

Although most histological diagnoses are made with relative ease and with great specificity and reproducibility, there is a subset of cases in which a specific and reproducible diagnosis is difficult or even impossible to render. In the melanocytic system, these cases can be divided into two broad categories. The first category, 'superficial atypical melanocytic proliferations of uncertain significance' (SAMPUS), includes predominantly junctional melanocytic proliferations, and melanocytic proliferations that are confined to the epidermis and papillary dermis, without evidence of tumorigenic proliferation or mitotic activity there. The prognosis for cure of these lesions is excellent if they are completely excised. Such lesions may include, for example, dysplastic naevi, Spitz naevi or pigmented spindle cell naevi with a few atypical melanocytes above the dermal-epidermal junction, or with greater than average cytological atypia, or with mitoses, where the differential diagnosis of melanoma in situ is difficult or impossible to rule out. The other category, 'melanocytic tumours of uncertain malignant potential' (MELTUMP), is comprised of melanocytic proliferations that form tumours in the dermis, and are therefore potentially capable of metastasis. Examples of such lesions may include atypical Spitz naevi, deep penetrating naevi, possible naevoid melanomas, or cellular blue naevi, where because of increased mitotic activity or cytologic atypia, a diagnosis of invasive or tumorigenic melanoma cannot be ruled out. In managing such lesions, we follow two main principles. The first is to manage each lesion with therapy designed to be adequate for management of the most significant consideration in the differential diagnosis. The second principle is to make clinicians and patients specifically aware of the diagnostic difficulty in their lesion, so that management can be undertaken on a true informed consent basis.     

Lentiginous melanoma: a histologic pattern of melanoma to be distinguished from lentiginous nevus.

Atypical lentiginous melanocytic proliferations in elderly patients continue to pose a diagnostic dilemma with lesions variably categorized as dysplastic nevus, atypical junctional nevus, melanoma in situ (early or evolving) and premalignant melanosis. We present pigmented lesions from 16 patients (seven male and nine female) and with the exception of one case, all were older than 50 years of age. The anatomical sites included trunk (7), head and neck (6) and upper extremity (3). The clinical diagnosis was variable and included lentigo maligna, atypical nevus, pigmented basal cell carcinoma, seborrheic keratosis and lentigo. The initial biopsies mimicked lentiginous nevus or dysplastic nevus and were characterized by a lentiginous proliferation of melanocytes at the dermoepidermal junction both as single cells and as small nests with areas of confluent growth, extending to the edges of the biopsy. The retiform epidermis was maintained and pagetoid spread of melanocytes was not prominent in hematoxylin- and eosin- stained sections. Dermal fibrosis was variably present and the melanocytic proliferation demonstrated cytological atypia. The subsequent re-excisions demonstrated similar atypical melanocytic proliferation occurring over a broad area flanking the prior biopsy sites. The diagnosis of melanoma was more easily recognized in the complete excision specimens. Immunohistochemical stains for Mitf and Mart-1 highlighted the extent of the basalar melanocytic proliferation as well as foci of pagetoid spread by melanocytes. Familiarity with this pattern of early melanoma should facilitate proper classification of lentiginous melanocytic proliferations in biopsies from older adults.     

Expression of interleukin-8 detected by in situ hybridization correlates with worse prognosis in primary cutaneous melanoma

Previous in vitro studies have demonstrated that endogenously produced human interleukin-8 (IL-8) can act as an important growth factor for human melanoma cells in vitro. The present study, has investigated whether IL-8 mRNA expression in primary melanomas may be of prognostic relevance with regard to melanoma progression and metastatic spread. In order to evaluate the clinical significance of IL-8 mRNA expression of melanoma cells in vivo, 59 melanocytic tissue specimens (37 primary melanomas and 22 melanocytic naevi) were studied using a semiquantitative in situ hybridization technique. Significant mRNA expression of IL-8 was found in 59 per cent (22/37) of melanomas. In 19 per cent (7/37) of the malignant melanomas, additional hybridization signals were noted within keratinocytes of the overlying epidermis. In contrast, paralesional normal-appearing epidermis and melanocytes in non-malignant lesions (melanocytic naevi) showed no IL-8 mRNA. Analysis of the relationship between IL-8 expression and clinico-histopathological features showed a significant association between IL-8 mRNA expression and the histological melanoma subtype (IL-8 mRNA: 14/19 in superficial spreading melanoma versus 4/12 in nodular melanoma, p< 0.05). Furthermore, IL-8 expression in primary tumours could be correlated with the patients' clinical course, with time to progression being significantly reduced in primary tumours expressing IL-8 in either the tumour cells or keratinocytes of the overlying epidermis. These results demonstrate for the first time that IL-8 expression, as detected by in situ hybridization in primary tumours, may serve as a significant prognostic factor for tumour progression in human malignant melanoma.     

Dysplastic naevi: an update

Elder D E
(2010) Histopathology 56 , 112–120
Dysplastic naevi: an update Dysplastic naevi are clinically atypical and histologically are characterized by architectural disorder and cytological atypia. Their diagnosis is reproducible if criteria and thresholds are agreed upon. They are significant only in relation to melanoma, as simulants of melanoma, as markers of individuals at increased risk of developing melanoma, and as potential and occasional actual precursors of melanoma. Morphologically and biologically, they are intermediate between common naevi and melanoma. Individuals with dysplastic naevi may have deficient DNA repair, and dysplastic naevi lesions are associated with overexpression of pheomelanin, which may lead to increased oxidative damage and increased potential for DNA damage and tumour progression.     

Basic fibroblast growth factor and ultraviolet B transform melanocytes in human skin

Ultraviolet (UV) light is an epidemiological risk factor for melanoma, but its specific contribution to melanoma induction is not known. The first critical step of melanoma development, ie, the uncontrolled proliferation of melanocytes, may be induced by a combination of UV damage and an imbalance of growth factor production by cells in the immediate area of the melanocyte. Among several candidates, basic fibroblast growth factor (bFGF) is the major autocrine growth factor in melanoma and associated with tumor progression. Overexpression of bFGF via adenoviral gene transfer in human skin xenografted to severe combined immunodeficiency mice led to black-pigmented macules within 3 weeks of treatment. Immunofluorescence analysis demonstrated pathological hyperpigmentation, proliferation and hyperplasia of activated melanocytes, but no malignant transformation. Similar changes were observed in skin reconstructs. When bFGF was combined with UVB, pigmented lesions with hyperplastic melanocytic cells were detected, including a lesion with high-grade atypia resembling lentiginous forms of malignant melanoma. Donor-matched control grafts revealed no melanocytic changes. bFGF was overexpressed in dermal fibroblasts demonstrating the co-carcinogenic influence of paracrine-acting growth factors by cells of the microenvironment. This is the first report suggesting that an imbalance of physiological growth factor production in the skin may cause melanoma in combination with UVB.     

p16ink4a expression in benign and malignant melanocytic conjunctival lesions

Acquired conjunctival melanocytic lesions include nevi, primary acquired melanoses (PAMs), and melanomas. Conjunctival melanoma is a malignant melanocytic neoplasm with a high metastasis and mortality rate. Usually, the diagnosis can be achieved only with routine microscopic analysis, but in some cases, the samples are small or have artifacts. In these cases, complementary studies will be helpful, but currently, there are no well-understood or studied complementary methods. Objective. To analyze the immunohistochemical expression of p16 in conjunctival melanocytic lesions and to assess its potential for differentiating between benign and malignant melanocytic lesions. Methods. Immunohistochemical study against p16ink4a (p16) was performed on paraffin-embedded sections on 45 melanocytic lesions (9 melanomas, 19 nevi, and 2 PAMs with atypia and 15 without atypia). Expression was scored according to the German immunoreactive score (IRS). Results. Expression of p16 IRS differed between nevi, PAMs, and melanomas. The mean IRS for melanomas was 3.3 ± 1.8 and was lower than those for nevi (7.63 ± 3.24; P < .05), PAM with atypia (12 ± 0; P < .05), and PAM without atypia (11 ± 1.69; P < .05). Lesions with infiltration depths lower than 2 mm showed higher levels of p16. There were no differences between favorable and unfavorable locations. Conclusion. p16 Expression in conjunctival melanocytic lesions showed an expression similar to that in skin and seems to be a good marker to differentiate nevi and PAMs from melanomas. However, additional studies of larger series and follow-up are needed to confirm these findings.     

Accuracy of core needle biopsy diagnosis in assessing papillary breast lesions: histologic predictors of malignancy.

The purpose of this study was to determine the accuracy of core needle biopsy (CNB) diagnosis of papillary breast lesions and to identify histologic features that can predict malignancy. We retrospectively reviewed 2876 CNB performed at MD Anderson Cancer Center (01/95-08/02) and identified 50 papillary lesions: 30 papillomas, eight atypical papillomas and 12 papillary carcinomas. Histopathological parameters were evaluated and radiographic findings were reviewed. When available, the CNB was compared with the excisional biopsy (EB) material. Carcinoma was confirmed by EB in 11/12 cases and invasion was correctly assessed in 67% of them. In EB, 6/8 (75%) atypical papillomas revealed carcinoma in situ or atypia and the remaining two (25%) were benign, six out of 30 (20%) papillomas had been excised and none had shown atypia; the remaining patients had clinical and radiological follow-up with no evidence of disease progression. We conclude that CNB is effective for assessing papillary breast lesions and that EB is more accurate in determining invasion. Cellular monotony, lack of myoepithelial cells, and cytologic atypia are more accurate predictors of malignancy (P<0.0001) than is the presence of mitoses (P<0.053). A diagnosis of carcinoma or atypical papilloma by CNB should warrant an EB, whereas benign papillomas may be followed if imaging findings are concordant.     

Differential immunoreactivity of melanocytic lesions of the conjunctiva

AIMS: To evaluate the expression of S100NKI/C3 and HMB45 antigens in melanocytic lesions of the conjunctiva and the ability of HMB45 to aid assessment of neoplasia. METHODS AND RESULTS: Stored formalin-fixed specimens of conjunctival melanomas and primary acquired melanosis were considered as participants and conjunctival naevi and racial melanosis as controls. Ninety-seven conjunctival melanocytic lesions were analysed using formalin-fixed paraffin-embedded material. These included 20 melanomas arising in the context of primary acquired melanosis (PAM), 22 melanomas arising without evidence of pre-existing PAM, seven cases of PAM with atypia, nine cases of PAM with no atypia, 35 conjunctival naevi and four cases of racial melanosis. S100 and NKI/C3 were similarly expressed in all lesions, with at least one of these markers positive in 100% of the lesions examined. HMB45 was expressed in 72.7% of primary melanomas and 85% of melanomas in the context of PAM; 42.8% of PAM with atypia expressed HMB45 while it was expressed in 11.1% of PAM without atypia and 8.5% of naevi. Racial melanosis cases did not express HMB45. S100 and NKI/C3 were expressed to a similar extent in all groups. CONCLUSIONS: S100 and NKI/C3 are useful markers to assess the extent of melanocytic lesions in the conjunctiva. HMB45 immunoreactivity can act as a useful aid to histopathology for the distinction of benign from malignant conjunctival lesions, particularly in the context of primary acquired melanosis.