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Molecular profiling already exerts a profound influence on biomedical research and disease management. Microdissection technologies contribute to the molecular profiling of diseases, enabling investigators to probe genetic characteristics and dissect functional physiology within specific cell populations. Laser-capture microdissection (LCM), in particular, permits collation of genetic, epigenetic, and gene expression differences between normal, premalignant, and malignant cell populations. Its selectivity for specific cell populations promises to greatly improve the diagnosis and management of many human diseases. LCM has been extensively used in cancer research, contributing to the understanding of tumor biology by mutation detection, clonality analysis, epigenetic alteration assessment, gene expression profiling, proteomics, and metabolomics. In this review, we focus on LCM applications for DNA, RNA, and protein analysis in specific cell types and on commercially available LCM platforms. These analyses could clinically be used as aids to cancer diagnosis, clinical management, genomic profile studies, and targeted therapy. In this review, we also discuss the technical details of tissue preparation, analytical yields, tissue selection, and selected applications using LCM.

Loss of heterozygosity (LOH) has previously been demonstrated at multiple chromosome microsatellites in primary and metastatic melanomas. Epidermotropic metastases of melanoma are unique in their varied histopathologic appearance, which can mimic a primary lesion. Our objective was to compare LOH profiles in primary and epidermotropic metastatic melanoma to delineate their clonal relationship. We examined the pattern of allelic loss in the primary melanomas of nine patients in addition to the 21 corresponding epidermotropic metastatic melanomas (average 2.3 metastases per patient). DNA samples were prepared from formalin-fixed, paraffin-embedded tissue sections using laser capture microdissection. Eight DNA microsatellite markers on six different chromosomes were analyzed: D1S214 (1p), D6S305 (6q), D9S171 (9p), D9S157 (9p), IFNA (9p), D10S212 (10q), D11S258 (11q), D18S70 (18q). In addition, X-chromosome inactivation analysis was performed in tumors from four women. LOH was seen in 67% (6/9) of primary melanomas and 81% (17/21) of epidermotropic metastatic melanomas. The most frequent allelic losses in informative cases occurred at 10q (33%), 9p (22%), and 11q (22%) in primary melanomas, and at 10q (50%), 1p (44%), and 6q (39%) in epidermotropic metastatic melanomas. Primary lesions demonstrating LOH had concordant allelic loss in at least one locus in a corresponding epidermotropic metastatic melanoma in 83% (5/6) of cases. X-chromosome analysis showed nonrandom inactivation in 75% (3/4) and 71% (5/7) of primary melanoma and epidermotropic metastatic melanoma cases, respectively. Our LOH and X-chromosome inactivation analysis data suggest that epidermotropically metastatic melanomas are clonally related to their primary lesion in many cases. Our data also indicated that some cases diagnosed as epidermotropic metastatic melanoma might be divergent clones or new primaries rather than metastatic disease.Modern Pathology (2007) 20, 821–827; doi:10.1038/modpathol.3800833; published online 15 June 2007

Loss of heterozygosity (LOH) has previously been demonstrated at multiple chromosome microsatellites in primary and metastatic melanomas. Epidermotropic metastases of melanoma are unique in their varied histopathologic appearance, which can mimic a primary lesion. Our objective was to compare LOH profiles in primary and epidermotropic metastatic melanoma to delineate their clonal relationship. We examined the pattern of allelic loss in the primary melanomas of nine patients in addition to the 21 corresponding epidermotropic metastatic melanomas (average 2.3 metastases per patient). DNA samples were prepared from formalin-fixed, paraffin-embedded tissue sections using laser capture microdissection. Eight DNA microsatellite markers on six different chromosomes were analyzed: D1S214 (1p), D6S305 (6q), D9S171 (9p), D9S157 (9p), IFNA (9p), D10S212 (10q), D11S258 (11q), D18S70 (18q). In addition, X-chromosome inactivation analysis was performed in tumors from four women. LOH was seen in 67% (6/9) of primary melanomas and 81% (17/21) of epidermotropic metastatic melanomas. The most frequent allelic losses in informative cases occurred at 10q (33%), 9p (22%), and 11q (22%) in primary melanomas, and at 10q (50%), 1p (44%), and 6q (39%) in epidermotropic metastatic melanomas. Primary lesions demonstrating LOH had concordant allelic loss in at least one locus in a corresponding epidermotropic metastatic melanoma in 83% (5/6) of cases. X-chromosome analysis showed nonrandom inactivation in 75% (3/4) and 71% (5/7) of primary melanoma and epidermotropic metastatic melanoma cases, respectively. Our LOH and X-chromosome inactivation analysis data suggest that epidermotropically metastatic melanomas are clonally related to their primary lesion in many cases. Our data also indicated that some cases diagnosed as epidermotropic metastatic melanoma might be divergent clones or new primaries rather than metastatic disease.

Phyllodes tumor of the prostate is a rare neoplasm with an unpredictable clinical behavior. It may undergo early recurrence with sarcomatous transformation or may even metastasize. Because targeted therapies have shown great success against several malignancies, there is hope that these same therapies may show similar promise in the treatment of other neoplasms. This study was undertaken to investigate both amplification of the epidermal growth factor receptor (EGFR) gene by fluorescence in situ hybridization and the overexpression of EGFR, Her-2/neu, CD117 (c-kit), and androgen receptor by immunohistochemical staining in a series of 11 phyllodes tumors of the prostate. In the stromal elements, EGFR gene amplification was present in four of 11 tumors and polysomy chromosome 7 was present in two of 11 tumors. No amplification was present in the epithelial components. Only one of 11 tumors had polysomy of chromosome 7 in the epithelial components. Immunohistochemically, in the stromal components, EGFR expression was demonstrable in four of 11 tumors and androgen receptor was demonstrated in six of 10 tumors. Neither Her-2/neu nor c-kit expression was seen in the stromal components of any of the 11 tumors. In the epithelial components, EGFR expression was present in all 11 tumors with strong staining in the basal cell layers and weak or no staining in luminal epithelium; androgen receptor expression was seen in seven of 10 tumors; Her-2/neu was weakly positive in four of 11 tumors; and c-kit expression was present focally and weakly in two of 11 cases with only 2–5% of cells staining. The highest staining intensity and the highest percentage of positively staining cells were seen with EGFR immunostaining in both the stromal and epithelial (mainly basal cells) components. Androgen receptor staining showed the next highest staining intensity and percentage of positive cells in both components. Her-2/neu and c-kit were only weakly or infrequently expressed in the epithelial components of prostatic phyllodes tumors. Our data indicate that EGFR and androgen receptor are frequently and strongly expressed in both epithelial and stromal components of prostatic phyllodes tumors. EGFR gene amplification is frequently present in prostatic phyllodes tumors and may account for one of the mechanisms leading to protein overexpression in some but not all cases. Anti-EGFR and/or antiandrogen agents may be potentially useful for management of patients with tumors expressing EGFR and/or androgen receptor.Modern Pathology (2007) 20, 175–182. doi:10.1038/modpathol.3800724; published online 22 December 2006

Phyllodes tumor of the prostate is a rare neoplasm with an unpredictable clinical behavior. It may undergo early recurrence with sarcomatous transformation or may even metastasize. Because targeted therapies have shown great success against several malignancies, there is hope that these same therapies may show similar promise in the treatment of other neoplasms. This study was undertaken to investigate both amplification of the epidermal growth factor receptor (EGFR) gene by fluorescence in situhybridization and the overexpression of EGFR, Her-2/neu, CD117 (c-kit), and androgen receptor by immunohistochemical staining in a series of 11 phyllodes tumors of the prostate. In the stromal elements, EGFRgene amplification was present in four of 11 tumors and polysomy chromosome 7 was present in two of 11 tumors. No amplification was present in the epithelial components. Only one of 11 tumors had polysomy of chromosome 7 in the epithelial components. Immunohistochemically, in the stromal components, EGFRexpression was demonstrable in four of 11 tumors and androgen receptor was demonstrated in six of 10 tumors. Neither Her-2/neunor c-kitexpression was seen in the stromal components of any of the 11 tumors. In the epithelial components, EGFRexpression was present in all 11 tumors with strong staining in the basal cell layers and weak or no staining in luminal epithelium; androgen receptor expression was seen in seven of 10 tumors; Her-2/neuwas weakly positive in four of 11 tumors; and c-kitexpression was present focally and weakly in two of 11 cases with only 2–5% of cells staining. The highest staining intensity and the highest percentage of positively staining cells were seen with EGFRimmunostaining in both the stromal and epithelial (mainly basal cells) components. Androgen receptor staining showed the next highest staining intensity and percentage of positive cells in both components. Her-2/neuand c-kitwere only weakly or infrequently expressed in the epithelial components of prostatic phyllodes tumors. Our data indicate that EGFRand androgen receptor are frequently and strongly expressed in both epithelial and stromal components of prostatic phyllodes tumors. EGFRgene amplification is frequently present in prostatic phyllodes tumors and may account for one of the mechanisms leading to protein overexpression in some but not all cases. Anti-EGFR and/or antiandrogen agents may be potentially useful for management of patients with tumors expressing EGFR and/or androgen receptor.

Intestinal metaplasia has been proposed to be a precursor lesion of adenocarcinoma in the urinary bladder. CDX2 is a transcription factor that is encoded by a homeotype gene that plays an essential role in the differentiation and proliferation of intestinal epithelial cells. Hepatocyte-specific antigen (Hep) has also been shown to be a useful marker of intestinal metaplasia. Tissues from 46 patients, including 22 cases of intestinal metaplasia of the urinary bladder, 11 cases of typical cystitis glandularis, and 13 cases containing both lesions, were selected and immunohistochemical stains for CDX2, Hep, cytokeratin 20 (CK20), and cytokeratin 7 (CK7) were performed. Nuclear staining for CDX2 was observed in 29 of 35 (83%) cases of intestinal metaplasia of the urinary bladder. In contrast, nuclear staining for CDX2 was not observed in any case of typical cystitis glandularis; however, seven of 24 (29%) cases showed aberrant cytoplasmic expression in a mean of 37% of cells. CK20 was expressed in 28 of 35 (80%) cases of intestinal metaplasia, but was observed in only one of 24 (4%) cases of cystitis glandularis in 15% of cells. CK7 was expressed in only six of 35 (17%) cases of intestinal metaplasia, whereas expression of CK7 was observed in all cases (100%) of typical cystitis glandularis with a mean percentage of positively staining cells of 63%. The mean percentages of positively staining cells in intestinal metaplasia with CDX2, CK20, and CK7 were 55, 49, and 53%, respectively. All examples of both intestinal metaplasia and typical cystitis glandularis were uniformly negative for Hep. In the urinary bladder, intestinal metaplasia and typical cystitis glandularis have sharply contrasting immunoprofiles. Additionally, the absence of Hep staining in intestinal metaplasia of the urinary bladder, despite its morphologic resemblance to normal colonic mucosa and intestinal metaplasia in other organs, may signify the presence of unique metaplastic pathways in the urinary bladder.Modern Pathology (2006) 19, 1395–1401. doi:10.1038/modpathol.3800670; published online 1 September 2006

Evidence of genetic heterogeneity within urothelial carcinomas of the bladder has raised questions about the clonal origin of urothelial carcinoma and its metastases. High‐grade urothelial carcinoma of the bladder frequently metastasizes to multiple regional lymph nodes in the pelvis. Whether or not these multiple lymph node metastases originate from the same tumor clone is uncertain. Molecular analysis of microsatellite alterations and X‐chromosome inactivation status of distinct tumor cell populations from the same patient may further our understanding of the genetic basis of carcinoma progression and metastasis.The authors examined 24 patients who underwent radical cystectomy for urothelial carcinoma. All patients had multiple (from two to four) lymph node metastases. Genomic DNA samples were prepared from formalin fixed, paraffin embedded tissue sections using laser‐assisted microdissection. Loss of heterozygosity (LOH) assays for 3 microsatellite polymorphic markers on chromosome 9p21 (D9S171, region of putative tumor suppressor gene p16), 9q32 (D9S177, putative tumor suppressor gene involved in urothelial carcinoma tumorigenesis), and 17p13 (TP53, the p53 locus) were performed. In addition, X‐chromosome inactivation analysis was performed in primary tumors and metastases from 10 female patients.In total, 79 tumors were analyzed. The overall frequency of allelic loss was 67% (16 of 24 tumors) in the primary urothelial carcinomas and 79% (19 of 24 tumors) in the metastatic carcinomas. The primary urothelial carcinoma showed LOH at the D9S171, D9S177, and TP53 loci in 39% (9 of 23 tumors), 30% (6 of 20 tumors), and 30% (7 of 23 tumors) of informative samples, respectively. LOH in ≥ 1 lymph node metastases was seen at the D9S171, D9S177, and TP53 loci in 35% (8 of 23 tumors), 45% (9 of 20 tumors), and 48% (11 of 23 tumors) of informative samples, respectively. Eleven tumors demonstrated identical allelic loss patterns at all DNA loci both in the primary carcinoma and in all corresponding lymph node metastases. Three tumors showed allelic loss in the metastatic carcinoma but not in its matched primary carcinoma. Six tumors demonstrated a different LOH pattern in each of its lymph node metastases. Clonality analysis showed the same pattern of nonrandom X‐chromosome inactivation both in the primary urothelial carcinoma and in all of the lymph node metastases in five of nine informative tumors studied. Four tumors showed a random pattern of X‐chromosome inactivation in both the primary carcinoma and in the metastases.LOH and X‐chromosome inactivation assays showed that multiple lymph node metastases and matched primary urothelial carcinomas of the bladder had the same clonal origin, suggesting that the capability for metastasis often arises in only a single clonal population in the primary tumor. The variable LOH patterns observed in some of the tumors likely reflect genetic divergence during the clonal evolution of urothelial carcinoma. Cancer 2005. © 2005 American Cancer Society.

Approximately 5% of clear cell renal cell carcinomas contain components with sarcomatoid differentiation. It has been suggested that the sarcomatoid elements arise from the clear cell tumors as a consequence of clonal expansions of neoplastic cells with progressively more genetic alterations. Analysis of the pattern of allelic loss and X‐chromosome inactivation in both the clear cell and sarcomatoid components of the same tumor allows assessment of the genetic relationship of these tumor elements.The authors of the current study examined the pattern of allelic loss in clear cell and sarcomatoid components of renal cell carcinomas from 22 patients who had tumors with both components. DNA samples were prepared from formalin‐fixed, paraffin‐embedded renal tissue sections using laser‐capture microdissection. Five microsatellite polymorphic markers for putative tumor suppressor genes on 5 different chromosomes were analyzed. These included D3S1300 (3p14), D7S522 (7q31), D8S261 (8p21), D9S171 (9p21), and TP53 (17p13). In addition, X‐chromosome inactivation analysis was performed in 14 tumors from female patients.The clear cell components showed loss of heterozygosity (LOH) at the D3S1300, D7S522, D8S261, D9S171, and TP53 loci in 18% (4/22), 18% (4/22), 50% (10/20), 15% (3/20), and 20% (4/20) of informative cases, respectively. LOH in the sarcomatoid components was seen at the D3S1300, D7S522, D8S261, D9S171, and TP53 loci in 18% (4/22), 41% (9/22), 70% (14/20), 35% (7/20), and 20% (4/20) of informative cases, respectively. Six cases demonstrated an LOH pattern in the clear cell component that was not seen in the sarcomatoid component. Different patterns of allelic loss were seen in the clear cell and sarcomatoid components in 15 cases. Clonality analysis showed the same pattern of nonrandom X‐chromosome inactivation in both clear cell and sarcomatoid components in 13 of the 14 cases studied. One case showed a random pattern of X‐chromosome inactivation.X‐chromosome inactivation analysis data suggest that both clear cell and sarcomatoid components of renal cell carcinomas are derived from the same progenitor cell. Different patterns of allelic loss in multiple chromosomal regions were observed in clear cell and sarcomatoid components from the same patient. This genetic heterogeneity indicates genetic divergence during the clonal evolution of renal cell carcinoma. Cancer 2005. © 2005 American Cancer Society.

In most cases, small-cell carcinoma of the urinary bladder is admixed with other histological types of bladder carcinoma. To understand the pathogenetic relationship between the two tumor types, we analyzed histologically distinct tumor cell populations from the same patient for loss of heterozygosity (LOH) and X chromosome inactivation (in female patients). We examined five polymorphic microsatellite markers located on chromosome 3p25-26 (D3S3050), chromosome 9p21 (IFNA and D9S171), chromosome 9q32-33 (D9S177), and chromosome 17p13 (TP53) in 20 patients with small-cell carcinoma of the urinary bladder and concurrent urothelial carcinoma. DNA samples were prepared from formalin-fixed, paraffin-embedded tissue sections using laser-assisted microdissection. A nearly identical pattern of allelic loss was observed in the two tumor types in all cases, with an overall frequency of allelic loss of 90% (18 of 20 cases). Three patients showed different allelic loss patterns in the two tumor types at a single locus; however, the LOH patterns at the remaining loci were identical. Similarly, the same pattern of nonrandom X chromosome inactivation was present in both carcinoma components in the four cases analyzed. Concordant genetic alterations and X chromosome inactivation between small-cell carcinoma and coexisting urothelial carcinoma suggest that both tumor components originate from the same cells in the urothelium.

We estimate the mass function of void galaxies in the second public data release of the Sloan Digital Sky Survey from a sample of 1000 galaxies with local density contrasts of dv < -0.6. The galaxy sample is split into ellipticals and spirals using a color-Sersic index criterion. We estimate the virial masses of ellipticals using the measured spectral line widths along with the observed size. Projection effects and uncertainties in halo properties make mass estimates of spirals more difficult. We use an inversion of the Tully-Fisher relation to estimate the isothermal rotational velocity and introduce a scaling factor to estimate the halo extent. We then fit the measured mass function against a theoretical Press-Schechter model and find that the distribution of galaxies in voids appears to be nearly unbiased compared to the mass.

The article examines the inaccuracies of real property category codes (CATCODE) used by the U.S. military services to classify property according to the use and functionality. It cites several examples that illustrate the inaccuracies of CATCODEs, facility sizes and occupancies. It adds that misclassification of properties may result to an installation's over or under-reporting of its facility capabilities, assets and space needs.