Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Negative Staining”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 1,459 records · Page 81Linked to original sources
PubMed1982

A biochemical and immunohistological study of collagen synthesis in Ewing's tumour.

The synthesis and localization of collagen have been studied on material from a total of 16 primary Ewing's tumours. The predominant collagen extracted from the tissues and synthesized in short-term cultures was type I. The proportion of type III collagen was relatively small and variable (0-8%) in the direct tumour extracts, but a higher proportion (29-38% of the total collagens) was synthesized in culture. Immunofluorescence studies showed that positive staining for all types of collagen tested (types I, III, IV and V) was restricted to stroma; there was no evidence of collagen either within the tumour cells or in their pericellular matrix, a finding endorsed by negative staining for reticulin in the same areas. The absence of any evidence for type IV or V collagen synthesis by Ewing's cells argues against an endothelial origin for the tumour, and indicates that collagen analysis is unlikely to be of value in the diagnosis of this particular sarcoma.

Bone Neoplasms↗
PubMed2001

S-Gal: an autoclavable dye for color selection of cloned DNA inserts.

Blue/white selection is the standard method for detecting a cloned DNA fragment. In the absence of an insert, uninterrupted expression of the vector-encoded alpha-complement of beta-galactosidase (beta-gal), results in the hydrolysis of X-gal (5-bromo-4-chloro-3-indolyl-beta-D-galactoside) and the subsequent blue staining of the host colony or bacteriophage plaque expressing the carboxyterminal portion of the beta-gal gene (lacZ). A white or clear colony or plaque indicates the presence of an insert. Because of its water insolubility, X-gal is dissolved in hazardous solvents such as dimethylformamide and then added to the medium following autoclaving. X-gal can be spread on previously plated medium, but this may result in an uneven color development. Also, incubation at 4 degrees C is frequently required for the distinction between a positive recombinant (unstained colony or plaque) and a stained negative. S-Gal (3,4-cyclohexenoesculetin-beta-D-galactopyranoside), a novel beta-gal substrate, is autoclavable and microwavable, allowing for dry-blending of the dye directly into the medium. Black S-Gal-stained colonies are visibly distinguishable from unstained colonies at an earlier time than X-gal. In addition, detection of the unstained signal over background is enhanced by 25% using S-Gal-containing medium, compared to medium containing X-gal. These characteristics offer convenience and better suitability for automated colony or plaque analyses.

Bacteriological Techniques↗
PubMed1996

Epithelial tumour cells in bone marrow of patients with pancreatic carcinoma detected by immunocytochemical staining.

In the present study, epithelial cells in the bone marrow of 42 patients with pancreatic carcinoma were identified immunocytochemically with monoclonal antibodies (MAbs) CK2, KL1 and A45-B/B3 directed to epithelial cytokeratins (CK), using the alkaline phosphatase anti-alkaline phosphatase method. The specificity of the MAbs was demonstrated by negative staining of marrow from 25 non-carcinoma age-matched control patients. Analysis of bone marrow aspirates from cancer patients revealed CK-positive cells in 14 (58.3%) of 24 cancer patients treated with curative intent and 10 (55.6%) of 18 patients with extended disease. After a median follow-up of 15.6 months (range 3-31 months), 5 (35.7%) out of 14 patients who underwent complete surgical resection but had tumour cells in bone marrow presented with distant metastasis and 6 (42.9%) with local relapse as compared to none of 10 corresponding patients without such cells (P < 0.05). The described technique may help to identify patients with pancreatic cancer and potential high risk of early metastatic relapse. The results promise to be of important assistance in determining prognosis and consequences in therapy of early stage pancreatic cancer.

Adenocarcinoma↗
PubMed1978

Reconstruction of glutamine synthetase using computer averaging.

The axial projection of the glutamine synthetase molecule has been reconstructed from electron micrographs of a stained preparation by using a new method of correlation search and averaging. The average over 50 individual molecules appears as a radial pattern with sixfold symmetry. The handedness evident in the average is attributed to nonuniformity of the negative stain.

Bacillus↗
PubMed1967

Filament ultrastructure and organization in vertebrate smooth muscle. Contraction hypothesis based on localization of actin and myosin.

Using a variety of preparative techniques for electron microscopy, we have obtained evidence for the disposition of actin and myosin in vertebrate smooth muscle. All longitudinal myofilaments seen in sections appear to be actin. Previous reports of two types of longitudinal filaments in sections are accounted for by technical factors, and by differentiated areas of opacity along individual filaments. Dense bodies with actin emerging from both ends have been identified in homogenates, and resemble Z discs from skeletal muscle (Huxley, 1963). In sections, short, dark-staining lateral filaments 15-25 A in diameter link adjacent actin filaments within dense bodies and in membrane dense pataches. They appear homologous with Z-disc filaments. Similar lateral filaments connect actin to plasma membrane. Dense bodies and dense patches, therefore, are attachment points and denote units analogous to sarcomeres. In glycerinated, methacrylate-embedded sections, lateral processes different in length and staining characteristics from lateral filaments in dense bodies exist at intervals along actin filaments. These processes are about 30 A wide and resemble heavy meromyosin from skeletal muscle. They also resemble heads of whole molecules of myosin in negatively stained material from gizzard homogenates. Intact single myosin molecules and dimers have been found, both free and attached to actin, even in media of very low ionic strength. Myosin can, therefore, exist in relatively disaggregated form. Models of the contraction mechanism of smooth muscle are proposed. The unique features are: (1) Myosin exists as small functional units. (2) Movement occurs by interdigitation and sliding of actin filaments.

Animals↗
PubMedDate not supplied

Immunologic profile of patients with fibromyalgia.

Fibromyalgia is a musculoskeletal disorder characterized by generalized myalgias, arthralgias widespread tender points in discreet areas on examination. It is frequently accompanied by fatigue, stiffness, and a nonrestorative sleep pattern. These patients generally have a normal blood count and chemistry profile. There is a subset of people with fibromyalgia (FM) who test positive for the antinuclear antibody (ANA) and have constitutional symptoms that resemble those of patients with early lupus. We studied the immunologic profile of patients with FM who are ANA-positive (+). A retrospective review of patient records in a university-based rheumatology practice was conducted. In a group of 66 FM patients, 30% (20) were ANA+, with a 75% preponderance of the speckled pattern and 20% diffuse pattern. The remaining 5% were equally split between diffuse-speckled and speckled-nucleolar patterns. All had negative staining for extractable nuclear antibodies. The Smart Index (SI), a ratio of the sedimentation rate to one-half the patient's age, was developed to characterize each patient's inflammatory response. The FM patients who were ANA negative (-) had a mean SI of 0.55, whereas the FM patient's who were ANA+ had a SI of 1.07. These ANA+ patients represent a subgroup of patients who have FM with an inflammatory response profile larger than that of the ANA-patients.

Adolescent↗
PubMed2003

Clinicopathological analysis of c-kit expression in carcinosarcomas and leiomyosarcomas of the uterine corpus.

OBJECTIVE: The purpose of this study was to evaluate the expression of the protooncogene, c-kit, in carcinosarcomas and leiomyosarcomas of the uterine corpus and determine the associations between c-kit expression and clinicopathologic factors, including clinical outcome. METHODS AND MATERIALS: Using a polyclonal anti-KIT-antibody, immunohistochemical staining was performed on formalin-fixed paraffin-embedded tissue blocks from 21 carcinosarcomas, 17 leiomyosarcomas, and 1 endometrial stromal sarcoma. KIT-positive tumors were defined as those tumors demonstrating immunopositivity in > or =30% of tumor cells examined. KIT-negative lesions demonstrated immunopositivity in <30% of tumor cells. Two authors independently scored the slides as positive or negative. Staining was repeated on all specimens and independently scored, and in the occasion of a mismatch, a third staining was performed. The carcinosarcomas were catalogued as to whether the sarcomatous and/or carcinomatous elements expressed c-kit. Clinical data were abstracted for those patients with uterine carcinosarcomas. The associations between clinicopathologic characteristics and c-kit expression were compared using univariate and multivariate analyses. Kaplan-Meier curves based on c-kit expression were plotted for progression-free and overall survival and compared using the log-rank test. RESULTS: Nine of 21 (43%) carcinosarcomas demonstrated immunopositivity for the KIT receptor, although staining was relatively weak. In contrast, only 1/17 (6%) leiomyosarcomas demonstrated KIT immunopositivity (P = 0.029). The solitary endometrial stromal sarcoma evaluated did not demonstrate significant KIT positivity. The majority of KIT-positive carcinosarcomas (6/9 (67%)) demonstrated KIT presence in the sarcomatous portion as compared to the carcinomatous portion (4/9 (44%)). No clinical factor had a statistically significant association with c-kit expression. The lack of c-kit expression was the only factor that was significantly associated with disease recurrence in univariate and multivariate analyses (P < 0.05), although there appeared to be a trend toward a low stage associated with kit positivity. The median progression-free interval for the KIT-negative cohort was 8 months, while it had not been reached for the KIT-positive cohort after median follow-up of 15 months (P = 0.0462). CONCLUSIONS: A significant proportion of carcinosarcomas of the uterine corpus display immunoreactivity for c-kit. Patients with KIT-positive carcinosarcomas may have an improved progression-free survival compared to KIT-negative tumors; however, further data are needed to determine whether this finding is confounded by stage.

Aged↗
PubMed1992

Immunohistochemical detection of estrogen and progesterone receptors in the normal urinary bladder and in pseudomembranous trigonitis.

According to recent studies showing the presence of estrogens receptors (ERs) in the human female lower urinary tract, we performed ER and progesterone receptor (PR) assays in fresh frozen and paraffin embedded biopsies taken from the urinary bladder. Fourteen females undergoing endoscopy during staging for gynecological cancer (endometrium, cervix, ovary) and 15 women complaining of recurrent abacterial cystitis (pseudomembranous trigonitis) were enrolled in the study as Group 1 and Group 2, respectively. After informed consent, they were submitted to cystoscopy, during which two biopsies were taken: one on the trigonum and the other on the bladder lateral wall. ERs were identified in both groups only in the trigonum (7/14 patients in Group 1 and 8/15 in Group 2), whereas the bladder lateral wall always stained negative. PRs were found at both sites in both groups (11/14 cases on the trigonum and 7/14 on the bladder lateral wall in Group 1; 11/15 and 2/15 respectively in Group 2). Morphological localization of PRs showed intense omogeneous staining in the nuclei of the stromal fibroblasts too. A clear correspondence between the presence of steroid receptors at the squamous metaplasia of the trigonum was observed. These data are discussed speculating about a possible endocrine pathogenesis of pseudomembranous trigonitis.

Adult↗
PubMed2007

Domains 16 and 17 of tropoelastin in elastic fibre formation.

Naturally occurring mutations are useful in identifying domains that are important for protein function. We studied a mutation in the elastin gene, 800-3G>C, a common disease allele for SVAS (supravalvular aortic stenosis). We showed in primary skin fibroblasts from two different SVAS families that this mutation causes skipping of exons 16-17 and results in a stable mRNA. Tropoelastin lacking domains 16-17 (Delta16-17) was synthesized efficiently and secreted by transfected retinal pigment epithelium cells, but showed the deficient deposition into the extracellular matrix compared with normal as demonstrated by immunofluorescent staining and desmosine assays. Solid-phase binding assays indicated normal molecular interaction of Delta16-17 with fibrillin-1 and fibulin-5. However, self-association of Delta16-17 was diminished as shown by an elevated coacervation temperature. Moreover, negative staining electron microscopy confirmed that Delta16-17 was deficient in forming fibrillar polymers. Domain 16 has high homology with domain 30, which can form a beta-sheet structure facilitating fibre formation. Taken together, we conclude that domains 16-17 are important for self-association of tropoelastin and elastic fibre formation. This study is the first to discover that domains of elastin play an essential role in elastic fibre formation by facilitating homotypic interactions.

Amino Acid Sequence↗
PubMed1985

A nucleus-basal body connector in Chlamydomonas reinhardtii that may function in basal body localization or segregation.

We have isolated a nucleus-basal body complex from Chlamydomonas reinhardtii. The complex is strongly immunoreactive to an antibody generated against a major protein constituent of isolated Tetraselmis striata flagellar roots (Salisbury, J. L., A. Baron, B. Surek, and M. Melkonian, J. Cell Biol., 99:962-970). Electrophoretic and immunoelectrophoretic analysis indicates that, like the Tetraselmis protein, the Chlamydomonas antigen consists of two acidic isoforms of approximately 20 kD. Indirect immunofluorescent staining of nucleus-basal body complexes reveals two major fibers in the connector region, one between each basal body and the nucleus. The nucleus is also strongly immunoreactive, with staining radiating around much of the nucleus from a region of greatest concentration at the connector pole. Calcium treatment causes shortening of the connector fibers and also movement of nuclear DNA towards the connector pole. Electron microscopic observation of negatively stained nucleus-basal body complexes reveals a cluster of approximately 6-nm filaments, suspected to represent the connector, between the basal bodies and nuclei. A mutant with a variable number of flagella, vfl-2-220, is defective with respect to the nucleus-basal body association. This observation encourages us to speculate that the nucleus-basal body union is important for accurate basal body localization within the cell and/or for accurate segregation of parental and daughter basal bodies at cell division. A physical association between nuclei and basal bodies or centrioles has been observed in a variety of algal, protozoan, and metazoan cells, although the nature of the association, in terms of both structure and function, has been obscure. We believe it likely that fibrous connectors homologous to those described here for Chlamydomonas are general features of centriole-bearing eucaryotic cells.

Calcium↗
PubMed1989

Immunohistochemical differentiation of clear-cell carcinoma of the female genital tract and endodermal sinus tumor with the use of alpha-fetoprotein and Leu-M1.

The morphologic differentiation between clear-cell carcinoma and endodermal sinus tumors is difficult at times. To improve the accuracy of the diagnosis, the authors studied nine ovarian and eight vaginal clear-cell carcinomas and seven endodermal sinus tumors of the ovary by immunohistochemical methods with the use of antibodies to alpha-fetoprotein and Leu-M1. Sixteen (94.1%) of the 17 clear-cell carcinomas and two (28.5%) of the seven endodermal sinus tumors reacted for Leu-M1, whereas six (85.7%) of the seven endodermal sinus tumors and three (17.6%) of the 17 clear-cell carcinomas stained for alpha-fetoprotein. Three clear-cell carcinomas and two endodermal sinus tumors showed immunoreactivity for both markers. No reactivity for either of these markers was present in one endodermal sinus tumor and one clear-cell carcinoma. All 13 tumors that stained only for Leu-M1 proved to be clear-cell carcinomas, and the four that reacted exclusively for alpha-fetoprotein were endodermal sinus tumors. Therefore, the authors concluded that positive immunostaining for Leu-M1 and negative immunostaining for alpha-fetoprotein support the differential diagnosis of clear-cell carcinoma, whereas a positive reaction for alpha-fetoprotein and a negative reaction for Leu-M1 favor a diagnosis of endodermal sinus tumor. However, positive or negative staining for both markers appears to have no diagnostic value.

Adenocarcinoma↗
PubMedDate not supplied

Value of GCDFP-15 (BRST-2) as a specific immunocytochemical marker for breast carcinoma in cytologic specimens.

OBJECTIVE: The diagnosis of metastatic mammary carcinoma by morphologic criteria alone can be difficult, depending on the site of metastasis and state of cell differentiation. Numerous histopathologic studies have shown GCDFP-15 (BRST-2) to be a specific marker for breast cancer in surgical specimens. To date, no studies have been done to evaluate its utility in cytologic preparations. STUDY DESIGN: To evaluate the usefulness of GCDFP-15 as a marker in the cytologic diagnosis of breast carcinoma, we studied 23 cases of mammary carcinoma and compared them with 20 cases of tumors of nonmammary origin (lung, ovary, liver, colon, stomach and bladder). "Bench top" fine needle aspirates from unfixed surgical specimens of breast carcinoma, cytocentrifuge samples from body cavity fluids and cerebrospinal fluids with morphologically proven metastatic carcinoma were studied. RESULTS: Expression of BRST-2 was found in 56.5% of primary and recurrent or metastatic breast carcinomas. All the nonmammary carcinomas studied were negative. Staining was found to be strongly dependent on the means of cell fixation. Slides fixed in 10% formalin and Bouin's solution gave optimal results. Except in two cases, which showed focal immunostaining, all specimens fixed in alcohol were negative. CONCLUSION: Our results support the diagnostic value of GCDFP-15 in recognizing tumors of breast origin and suggest that in clinical situations in which metastatic breast carcinoma is suspected, a portion of the cytologic specimen should be fixed with an optimal fixative for BRST-2 detection.

Antibodies, Monoclonal↗
PubMed2005

Expression of human telomerase reverse transcriptase in thyroid follicular neoplasms: an immunohistochemical study.

Differential diagnosis of follicular adenoma (FA) and follicular carcinoma (FC) of the thyroid can be difficult in the routine practice of surgical pathology because the diagnosis of FC is strictly defined and determined by the presence of capsular and/or vascular invasion by the tumor. These features may be equivocal in the histologic sections. On the other hand, telomerase is expressed in many human cancers and is thought to contribute to their immortality. Human telomerase reverse transcriptase (hTERT) is the major determinant of human telomerase activity, and its expression is suggestive of capacity for unlimited replication. This case-control study examined the expression of hTERT using immunohistochemistry in 36 thyroid FC and 36 FA from patients who were matched by age and sex. The aim was to investigate the value of immunohistochemical staining for hTERT in the differential diagnosis of follicular neoplasms. The results revealed 23 cases of FC and 14 cases of FA that showed high expression of hTERT, with moderate to strong immunoreactivity. The remaining cases showed weak or negative staining. The difference between FA and FC was statistically significant (p < 0.05). In conclusion, immunohistochemical staining for hTERT can be considered an ancillary marker for differential diagnosis of FA and FC.

Adenocarcinoma, Follicular↗
PubMed2004

[Studies on expression and location of VEGF protein in rat testis and epididymis].

In order to investigate the function of vascular endothelial growth factor (VEGF) in the male reproductive system, we examined the expression and localization of VEGF protein in adult rat testis and epididymis with the immunohistochemical method and PAS stain and western blots. The results showed that the VEGF protein (about 45 kD) was present in the rat testis and epididymis. The VEGF immunoreactive particles were localized in the cytoplasm of spermatids, Sertoli cells and Leydig cells with negatively stained nuclei. The expression pattern of the spermatids was different in the 14 stages of epithelial cycle, and the strong immunopositive reaction was mainly in the forming/formed acrosome of round/elongated spermatids and spermiogenic residual bodies. In the epididymis, the region-specific and cell-specific expression and localization of VEGF protein was shown in the epididymal epithelium. In the initial segment, the VEGF protein was presented in the whole epithelial cells and the immunoreactive particles were localized in the supranuclear region of the cells. Except for a few principal cells and basal cells, counter-staining of PAS stain proved that the immunoreactive cells were clear cells in the caput, corpus, and cauda epididymis. And the immunoreactive particles were localized in the different region of the cytoplasm of clear cells, some were in the supranuclear region, some were filled in the whole cytoplasm, and some were secreted out of the cells along the free border. The positive acrosomes of the spermatozoa were only seen in the lumen of the proximal epididymis. No expression was detected in the interstitial blood vessels and the capillaries of the testis and epididymis. The present results indicate that VEGF protein may play an important role in the process of the spermatogenesis and spermiogenesis, especially in the acrosomal developing process. VEGF may also be related to the sperm maturation in the epididymis.

Animals↗
PubMed1989

[McAb LC-1 against human lung cancer].

Recently Ge et al reported LAC-122, LAC-210 and LAC-163 McAbs against Human non-small cell lung cancer and LSC McAbs against human lung small cell carcinoma. The immunotoxin, composed of McAb LAC-122 conjugated with Ricin A chain has been reported to have the significant cytotoxic effect in vitro on lung adenocarcinoma cell line SPC-A-1 by Tan et al. The LAC-122 alone has no effect on this target cell in the presence of complement from human, rabbit or guinea pig. The tumor associated antigens of human lung cancer have been recognized for many years, but only few reports deal with the common antigens or common epitopes of the lung cancer. From one fusion, 20 hybrids had been observed, all of these culture supernatants could react with target cell by IF. One of them after 4 th cloning, immunoglobulin isotype of the monoclonal antibody thus far obtained belonged to IgM and named to LC-1. Table 1 showed the results of ABC staining of LC-1 with a variety of tumors, normal adult and fetal tissues. From 12 non-small cell lung cancers, including 7 lung adenocarcinoma, 2 lung squamous carcinoma, 3 lung giant cell carcinoma, only one adenocarcinoma gave negative staining. As for 6 small cell lung cancers, all of them showed the positive reaction. It could be also reacted with 11 other tumors.(ABSTRACT TRUNCATED AT 250 WORDS)

Antibodies, Monoclonal↗
PubMed2000

[Remodeling of basement membrane in association with cancer invasion].

Type IV collagen the major component of basement membrane (BM), is composed of six genetically distinct alpha chains. In normal breast tissue, benign breast tumors, and in the intraductal components of invasive ductal carcinoma, alpha 1 (IV) and alpha 2 (IV) chains were stained in all BM, whereas alpha 5 (IV) and alpha 6 (IV) chains were restrictively localized in a linear pattern in the epithelial BM. However, in invasive ductal carcinoma, alpha 1 (IV) and alpha 2 (IV) chains were discontinuously or negatively stained in the cancer cell nest, and the assembly of alpha 5 (IV) and alpha 6 (IV) chains into the BM was completely inhibited. The results indicate that the mammary gland forms a second network of BM composed of alpha 5 (IV)/alpha 6 (IV) chains, in addition to the classic network of alpha 1 (IV)/alpha 2 (IV) chains. Remodeling of type IV collagen alpha chains during the development of invasive breast cancer seems to be differentially regulated, and to be associated with modification of histopathological findings.

Basement Membrane↗
PubMed2002

Persistence of solitary mammary carcinoma cells in a secondary site: a possible contributor to dormancy.

Tumors can recur years after treatment, and breast cancer is especially noted for long periods of dormancy. The status of the cancer during this period is poorly understood. As a model to study mechanisms of dormancy, we used murine D2.0R mammary carcinoma cells, which are poorly metastatic but form occasional metastases in liver and other organs after long latency. Highly metastatic D2A1 cells provided a positive, metastatic control. Our goals were to learn how the cell lines differ in survival kinetics in a secondary site and to seek evidence for the source of D2.0R dormancy. In spontaneous metastasis assays from mammary fat pad injections, we found evidence for dormancy because of a persistence of large numbers of solitary cells in the liver. To quantify the fate of cells after arrival in liver, experimental metastasis assays were used. To permit identification of cells that had not divided, cells were labeled before injection with fluorescent nanospheres, which were diluted to undetectable levels by cell division. Cancer cells were injected i.v. to target them to the liver and coinjected with reference microspheres to monitor cell survival. Dormancy was defined as retention of nanosphere fluorescence in vivo, as well as negative staining for the proliferation marker Ki67. A large proportion of D2.0R cells persisted as solitary dormant cells. No metastases formed, but viable cells could be recovered from the liver 11 weeks after injection. Large numbers of solitary, dormant, Ki67-negative D2A1 cells were also detected against a background of progressively growing metastases. Thus, this study identified a possible contributor to tumor dormancy: solitary, dormant cells that persist in tissue. If such cells are present in patients, they could contribute to tumor recurrence and would not be susceptible to current therapeutic strategies targeting proliferating cells.

Animals↗
PubMedDate not supplied

Differential (Ha-, K- and N-) ras p21 expression in benign and malignant human thyroid tumors: an immunohistochemical study.

An immunohistochemical study of 24 thyroid carcinomas, 10 thyroid adenomas and 10 normal thyroids was performed using monoclonal antibodies directed against each of the three members of ras p21s; Ha-, K- and N-ras p21. Normal thyroids showed a negative staining for all of the three ras p21s. The expressional levels of the three ras p21s were various in each benign and malignant thyroid tumor, but a higher expression of Ha- and N-ras p21 and a lower or negative expression of K-ras p21 was a common finding in thyroid tumors of all histologic categories. There were no correlations between the expressional levels of ras p21s and the histologic subtypes, tumor size or lymph node status in the thyroid carcinomas.

Adenocarcinoma, Follicular↗