Evaluation of micrometastases in sentinel lymph nodes of cutaneous melanoma: higher diagnostic accuracy with Melan-A and MART-1 compared with S-100 protein and HMB-45.

Accurate diagnosis of micrometastases in sentinel lymph nodes of cutaneous melanoma is critical for proper clinical management. S-100 protein and HMB-45 are the traditional immunomarkers widely used for this purpose. However, the interpretation of micrometastases by these markers is difficult with significant reduction in the diagnostic accuracy. S-100 protein demonstrates immunoreactivity for other nonmelanoma cells and obscures nuclear details, which are crucial for the interpretation of single cell metastases. We compared the new melanoma markers, Melan-A (clone A103) and MART-1 (clone M2-7C10), with S-100 protein and HMB-45, by examining 77 formalin-fixed paraffin-embedded sections of sentinel lymph nodes from 13 cases of primary cutaneous melanoma. CD68 (PG-M1) and hematoxylin-eosin-stained sections were also studied. Four pathologists interpreted the staining pattern after concealing the identity of each immunomarker. Az values (area under receiver operating characteristic curve) with receiver operating characteristic curve were higher with Melan-A (0.9742) and MART-1 (0.9779) compared with S-100 protein (0.8034) and HMB-45 (0.8651), demonstrating a higher diagnostic accuracy with Melan-A and MART-1 with superior detection of melanoma micrometastases. Melan-A and MART-1 showed sharp cytoplasmic immunoreactivity, almost exclusively restricted to the melanoma cells. Therefore, Melan-A and MART-1 are recommended for the evaluation of micrometastases in sentinel lymph nodes of cutaneous melanoma as a routine alternative to S-100 protein and HMB-45.

Routine air drying of all smears prepared during fine needle aspiration and intraoperative cytology studies. An opportunity to practice a unified protocol offering the flexibility of choosing a variety of staining methods.

OBJECTIVE: To evaluate the possibility of routine use of air-dried smears (ADS) instead of wet-fixed smears (WFS). STUDY DESIGN: Intraoperative cytology (IC) smears from 293 specimens and fine needle aspiration cytology (FNAC) smears from 118 cases were studied. Cytomorphology of ADS processed with our protocol for hematoxylin-eosin (HE) and Papanicolaou (PAP) staining after saline rehydration and postfixation in 95% ethanol with 5% acetic acid were compared with respectively stained WFS. Additional ADS were stored up to 72 hours at room temperature prior to HE, PAP and Diff-Quik (DQ) staining to evaluate the effects of postponing rehydration and postfixation. Special stains for fungi were also studied in four cases. RESULTS: ADS were easy to prepare without air-drying artifact in the final HE- and PAP-stained smears. ADS were more cellular than WFS. Erythrocyte interference was frequent in WFS. HE and PAP staining of ADS stored up to 72 hours showed cytomorphology comparable to that of the similarly stained fresh smears. However, DQ staining was better if ADS were processed before 24 hours. ADS stained with special stain for fungi showed good morphology, similar to that in WFS. CONCLUSION: All ADS showed results comparable to or better than WFS. ADS could be stored up to 72 hours before staining with HE and PAP. ADS offers the flexibility of selecting a variety of staining methods and is a practical alternative to WFS.

Methods of cytologic smear preparation and fixation. Effect on the immunoreactivity of commonly used anticytokeratin antibody AE1/AE3.

OBJECTIVE: To evaluate the effect of fixation and methods of cytologic smear preparation on the immunoreactivity of commonly used anticytokeratin antibody AE1/AE3. STUDY DESIGN: Scrape cytology smears and formalin-fixed, paraffin-embedded tissue sections (FPTS) of 20 unfixed, fresh specimens submitted for intraoperative consultation were studied by the immunoperoxidase method. In addition to the morphologic examination, the smears and FPTS were evaluated for intensity and proportion scores. For each specimen, two scrape cytology smears were wet fixed in 95% ethanol, and 12 smears were air dried without fixation. Air-dried smears were either postfixed after rehydration in saline or fixed directly without rehydration by one of the three fixatives: alcoholic formalin, 95% ethanol with 5% acetic acid or 95% ethanol. RESULTS: Both intensity and proportion scores were higher with rehydrated, air-dried smears as compared to those without rehydration and were comparable to those with wet-fixed smears and FPTS. In the rehydrated group, the optimum results were achieved when the smears were postfixed with alcoholic formalin. CONCLUSION: The method of preparation and fixation had variable effects on the immunoreactivity of anticytokeratin antibody AE1/AE3. The optimum results were achieved with saline-rehydrated, air-dried smears post-fixed in alcoholic formalin. To evaluate the role of inter-sample variation, further, larger studies are recommended on this and other antibodies before applying them to different types of cytologic smears.

Immunophenotypic profile of myeloid cells in granulocytic sarcoma by immunohistochemistry. Correlation with blast differentiation in bone marrow.

The present study was designed to evaluate the lineage differentiation (particularly monocytic differentiation) of immature myeloid cells in granulocytic sarcoma (GS) by immunohistochemistry and correlate the results with lineage differentiation of blasts in the bone marrow and to determine the degree of maturation of the infiltrating myeloid cells in GS by immunohistochemistry using CD34 and HLA-DR. Immunohistochemical stains were performed on paraffin-embedded tissue from 17 GS lesions with lineage-associated markers: myeloperoxidase, CD68 (KP1), CD68 (PG-Ml), glycophorin A, factor VIII, and CD56; and with markers for blasts and immature myeloid cells: CD34 and HLA-DR. Our results show that positive staining with PG-M1, but not KP1, suggests monocytic differentiation of myeloid cells in GS and correlates with the monocytic differentiation of blasts in the bone marrow. Expression of CD56 is frequent in GS, especially when the marrow blasts have monocytic differentiation, and should not be interpreted as a primary natural-killer cell process. The immature myeloid cells in GS are frequently HLA-DR positive. However, CD34 positivity of the immature myeloid cells is relatively uncommon, except in cases with underlying myelodysplastic syndrome or chronic myelogenous leukemia.

Evaluation of apoptotic leukocytes in peripheral blood smears.

OBJECTIVES: Apoptosis is a phenomenon of physiological cell death in which each step is regulated, similar to the process of mitosis. We observed apoptosis in leukocytes during the review of peripheral blood smears. This study was undertaken to evaluate the morphologic features of apoptotic leukocytes in peripheral blood smears and to ascertain their clinical significance. DESIGN: Sixty cases (23 males and 37 females, aged newborn to 92 years) exhibiting apoptotic leukocytes in peripheral blood smears were studied. Medical records for each case were reviewed, and patients were categorized according to their clinical diagnoses. RESULTS: Neutrophils were the most common apoptotic leukocytes identified (85%), followed by lymphocytes (18%) and eosinophils (2%). The diagnosis most frequently associated with the presence of apoptotic leukocytes was infection (55%). Apoptosis in lymphocytes was comparatively less common, but when present, the most common associations were diabetes mellitus, glucocorticoid administration, and neoplastic diseases. CONCLUSIONS: Our findings suggest that the presence of apoptotic leukocytes in peripheral blood smears may help in the differential diagnosis and may be related to the severity of disease. We recommend further evaluation using additional special techniques for detecting apoptotic leukocytes.

Cervical cytology and immunohistochemical features in endometrial adenocarcinoma simulating microglandular hyperplasia. A case report.

BACKGROUND: The histology of a few cases of adenocarcinoma simulating cervical microglandular hyperplasia (MGH-AdCa) has been reported. However, the cytologic features of MGH-AdCa in cervical smears and the immunohistochemical profile have not been described. CASE: A 73-year-old female presented with vaginal bleeding. The cervical Pap smear was initially interpreted by the cytotechnologist as "reactive endocervical cells" and was referred for cytopathologist review. The final interpretation was atypical glandular cells of undetermined significance (AGUS), probably neoplastic. Endometrial biopsy and total abdominal hysterectomy with bilateral salpingo-oophorectomy showed International Federation of Gynecologists and Obstetricians grade 1 endometrial carcinoma. The superficial component of the tumor resembled cervical microglandular hyperplasia (MGH); the deeper component had an endometrioid pattern. The Pap smear predominantly showed a glandular component with features of MGH. However, the presence of scattered single cells with hyperchromatic nuclei, one to three nucleoli, easily detectable mitotic figures, randomly scattered apoptotic bodies and focal, watery diathesis suggested a neoplastic process. Immunohistochemistry was studied on paraffin sections. In addition to other markers, the tumor cells were immunoreactive for carcinoembryonic antigen (CEA). CONCLUSION: Although the cervical Pap smear in this case had an MGH-like pattern, some features were atypical enough to suggest a diagnosis of AGUS, probably neoplastic. CEA immunoreactivity of MGH-AdCa could also help to differentiate it from MGH.

The role of constitutive NF-kappaB activity in PC-3 human prostate cancer cell invasive behavior.

The purpose of this study was to determine if increased NF-kappaB activity of highly invasive PC-3 cells contributed to their invasive behavior. Increased NF-kappaB activity has been observed in several malignant tumors and it may have an important role in tumorigenesis, progression and chemotherapy resistance. By serial selection, we obtained invasion variant PC-3 cell sublines. The PC-3 High Invasive cells invade readily through a Matrigel reconstituted basement membrane while PC-3 Low Invasive cells have low baseline invasion activity. In these studies, we discovered that NF-kappaB DNA binding activity was increased in PC-3 High Invasive cells when compared to PC-3 Low Invasive cells by electrophoretic mobility shift assay (EMSA). Gel supershift assays showed a 4-fold increase in p65 containing complexes and a 2.2-fold increase in the p50 containing complexes in the PC-3 High Invasive cells. Luciferase reporter assays showed that NF-kappaB dependent transcription activity was increased 10.2 +/- 2.5-fold in the highly invasive cells (P < 0.002). The PC-3 High Invasive cells showed a constitutive increase in phospho-IkappaB alpha and introduction of the super-repressor IkappaB alpha S32/36A inhibited NF-kappaB activity to 19.2 +/- 2.5 percent of control transfected cells (P < or = 0.001). The IkappaBa super-repressor reduced the basement membrane invasion of PC-3 High Invasive cells from 6.2 +/- 1.1 to 3.8 +/- 0.4 percent (P < 0.002) with no decrease in cell viability or proliferation. These results demonstrate that increased NF-kappaB activity contributed directly to the invasive behavior of PC-3 High Invasive prostate cancer cells.

Prostate-specific antigen expression and lipochrome pigment granules in the differential diagnosis of prostatic adenocarcinoma versus seminal vesicle-ejaculatory duct epithelium.

BACKGROUND: Lipochrome pigment granules (LPGs) and prostate-specific antigen (PSA) localization have been cited as helpful adjuncts in differentiating atypical histologic patterns of seminal vesicle-ejaculatory duct (SVED) from prostatic adenocarcinoma. However, LPGs have been described in both benign and neoplastic prostatic acini, and PSA expression within the intraprostatic SVED has not been fully explored. DESIGN: Fifty radical prostatectomy specimens were studied for LPGs and 9 cases for PSA expression. RESULTS: Two morphologic types of LPGs (type 1 and type 2) were observed. The reproducibility in classifying LPGs was evaluated by kappa statistics, which demonstrated a strong agreement between 4 observers. Type 1 was restricted to SVED in all 50 specimens. Type 2 was subclassified into 2A and 2B. Type 2 LPGs were observed in prostatic acini of different zones, high-grade prostatic intraepithelial neoplasia, prostatic adenocarcinoma, and occasionally with type 1 LPG in SVED. Focal reactivity for PSA in the distal portion of SVED near urethra was noted in 1 of 9 cases. CONCLUSION: Awareness about morphologic differences between the 2 types of LPGs could help to avoid a potential diagnostic pitfall of misinterpreting SVED epithelium for adenocarcinoma. Caution is recommended in interpreting PSA expression, since rare focal PSA reactivity was observed in the distal SVED.

Low-grade fibromyxoid sarcoma: clinicopathologic case report with review of the literature.

Low-grade fibromyxoid sarcoma is a rare, benign-appearing soft tissue neoplasm with an aggressive clinical course characterized by multiple local recurrences over several years, with ultimate spread to lung and occasionally to bone. Thus far, a total of 24 cases of low-grade fibromyxoid sarcoma have been reported in the literature. The authors present an additional case that grossly and microscopically emphasizes a pronounced lobular pattern of contrasting areas of cellularity showing high proliferative activity, as demonstrated by a proliferation marker, Ki 67 with MIB-1, and hypocellular areas with prominent myxoid component and abundant collagen fibrils. There was predominance of delicate capillary-sized stromal vessels with collagenized walls in both cellular and myxoid areas. The unusual features in this case were osseous metaplasia, prominent intranuclear pseudoinclusions, DNA tetraploidy, and membrane-bound intracytoplasmic fat vacuoles. The immunoprofile and cytologic and ultrastructural features are described. After the excision of the tumor, the patient was treated with radiotherapy without chemotherapy. The patient has been observed for 26 months and is alive without the evidence of disease. The postoperative follow-up with axial computed tomography at 24 months showed no evidence of disease, except postsurgical fibrotic changes.

Pheochromocytoma. Cytologic findings on intraoperative scrape smears in five cases.

BACKGROUND: There have been few studies describing the cytology of adrenal pheochromocytoma (PC). Although fine needle aspiration (FNA) for a preoperative diagnosis of PC is generally considered a contraindication, this tumor can be an unsuspected finding in adrenal FNA performed for other reasons. STUDY DESIGN: Scrape cytology smears prepared in five cases of PC were examined for different cytomorphologic features. The results were correlated with the corresponding permanent histologic sections. RESULTS: Previously described features, like cellular smears showing cells with abundant, poorly defined fragile cytoplasm, bare nuclei, anisonucleosis, "salt and pepper" chromatin, variable nucleoli and few ganglion cell-like cells, were noted. In addition, several previously unreported cytologic features were observed: (1) loosely cohesive PC cells along a ramifying, delicate central core; (2) intracytoplasmic microvesicular (not hyaline/homogeneous) globules; and (3) different arrangements of capillary-stroma and PC cells (Zellballen pattern; empty capillary rings; stroma with adherent, intact PC cells or fragments of disrupted PC cell cytoplasm). CONCLUSION: The cytologic appearance of PC may resemble that of other neuroendocrine tumors; however, it can be diagnostic when combined with proper clinical data and ancillary tests.

Fine needle aspiration cytology of gastric solitary fibrous tumor: a case report.

BACKGROUND: Solitary fibrous tumors (SFT) occur mainly in the pleura and other serosal sites. However, they have been found in extraserosal sites and should be considered in the differential diagnosis (DDx) of any spindle cell lesion, including those in the gastrointestinal tract. In this report, we describe fine needle aspiration (FNA) cytologic evaluation of a gastric SFT, emphasizing the role of immunocytochemistry in the DDx. CASE: Computerized tomography-guided FNA of a subserosal gastric mass in a 77-year-old female was performed. The moderately cellular smears showed neoplastic cells arranged in interlacing fascicles and in a "patternless" pattern. There was variable collagenous stroma. The cell block revealed a similar pattern, with a single mitotic figure. Nuclear atypia and necrosis were absent. The neoplastic cells were strongly reactive for vimentin and CD34, with weak focal reactivity for smooth muscle actin, suggestive of vessels in tangential section. They were nonreactive for muscle specific actin, desmin, S-100 and pancytokeratin. Other immunocytochemical markers were also studied. CONCLUSION: SFT should be considered in the DDx of spindle cell lesion of the stomach. Cell block and immunocytochemical markers, especially CD34, were extremely useful in the diagnosis of SFT on FNA.

A rapid, economical, and simple method for concentration of Schistosoma mansoni ova in feces.

An easy, rapid, and economical method for concentration of Schistosoma mansoni ova in feces is described. The basic procedure involves 20 minutes of gravity sedimentation of a fecal suspension sieved through gauze and suspended in a 5% (volume/volume) solution of glycerol in tap water with Ig/L benzoic acid. There is excellent recovery of S. mansoni ova. It can be applied under field conditions and can be implemented by any laboratory with routine facilities. It also allows detection of other ova, larva, and to some extent, cysts.

Role of scrape cytology in rapid intraoperative diagnosis. Value and limitations.

The diagnostic applications of frozen sections and imprint cytology are well known. A study was conducted to evaluate the role of scrape cytology, which is a modification of imprint cytology, in the rapid intraoperative diagnosis of tumors. For 249 surgical specimens studied by scrape cytology, the diagnostic accuracy was 98.4% and the false-positive rate was 0.4%. The method is rapid, easy, reliable and does not require special instruments or set ups. The diagnostic accuracy was better than reported for imprint cytology or frozen sections alone. Scrape cytology is advocated as a routine intraoperative procedure, especially where frozen section facilities are not available. While not meant as an alternative to frozen sections, scrape cytology can be an excellent adjunct to that technique.

Application of specific red blood cell adherence test to the human cornea and conjunctiva.

We modified the specific red blood cell adherence test used in the study of blood group antigens to apply to the human cornea and conjuctiva. For the first time this test was applied to these ocular tissues with considerably specificity and sensitivity. The test revealed a concentration of blood group antigens in the epithelium of cornea and conjunctiva and along the corneal endothelium, whereas it was absent in the stroma. The test is useful in studying ABO compatibility in corneal transplants.