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Papillary and follicular neoplasms of the thyroid gland. Differential immunohistochemical staining with high-molecular-weight keratin and involucrin.

The morphologic distinction between papillary and follicular neoplasms of the thyroid gland can be difficult, especially on small biopsy specimens or in fine-needle aspirations. To determine whether immunohistochemistry could help in achieving the correct diagnosis, we characterized the staining pattern for a series of papillary and follicular neoplasms of the thyroid gland. A pilot study was performed using a panel of antibodies, including high-molecular-weight keratin (HMWK, 34 beta E12), cytokeratin (CK) 5/6, CK7, CK13, CK14, CK20, AE1/AE3, CAM5.2, involucrin, and villin. Of these antibodies, involucrin and HMWK showed strong differential staining between follicular and papillary neoplasms. HMWK stained 91% of papillary carcinomas, including follicular variants, with a median of 53% positive cells, and involucrin stained 72.5% of papillary neoplasms with a median of 45% positive cells. HMWK stained only 20% of follicular neoplasms, whereas involucrin stained 29% of cases. Papillary neoplasms showed strong, although patchy, staining with HMWK and involucrin, whereas those follicular neoplasms that did have staining showed a weak, diffuse pattern of staining. We believe that HMWK, and involucrin to a lesser degree, could be useful in differentiating papillary from follicular neoplasms, especially for cytologic cell block material or for cases in which the architectural pattern is follicular.

Adenocarcinoma, Follicular↗

Is immunostaining for Helicobacter pylori superior to the special stain thiazine in detecting small numbers of H. pylori in gastric biopsies?

Helicobacter pylori is implicated in the pathogenesis of gastritis and duodenal ulcers, gastric lymphoma of the mucosa-associated lymphoid tissue (MALT) type, and gastric adenocarcinoma. Eradication of H. pylori with antibiotic therapy therefore is essential, not only for the successful treatment of active gastritis, but also for the treatment and prevention of the MALT lymphoma. It has been suggested recently that immunostaining for H. pylori is more sensitive than special stains for the detection of the organism in the gastric biopsies after triple therapy. Fifty-five endoscopic mucosal biopsies from 38 patients, including 18 treated with H. pylori eradication therapy, were selected for immunostaining because they were either negative or contained rare H. pylori organisms by thiazine stain. Formalin-fixed, paraffin-embedded tissue sections were immunostained for H. pylori using a polyclonal antibody using standard immunoperoxidase technique. The results were compared with those obtained with the thiazine stain. Detection of H. pylori by immunostaining was easier and less time-consuming than by thiazine stain. There was complete agreement between immunostaining and thiazine stain in 48 (87%) cases. Of the 7 discordant cases, 3 (42%) were positive for H. pylori with thiazine only and 4 (48%) with immunostaining only. Given the nature of the selection of the study sample (absent to rare by thiazine stain), the discordance most likely represents a sampling error. The authors concluded that immunostaining for H. pylori did not appear to be more sensitive than special stains. Three cases with bacterial clumps were diagnosed previously as positive for H. pylori, but identified correctly as negative using both staining methods. Pathologists, however, should balance the added cost to patients of immunostaining against the time saved by the easier screening of the immunostained slides and the possibility of false positive results when special stains are interpreted by inexperienced pathologists.

Biopsy↗

Hep par 1 antibody stain for the differential diagnosis of hepatocellular carcinoma: 676 tumors tested using tissue microarrays and conventional tissue sections.

A well-characterized positive marker for hepatocellular differentiation would be a useful tool for the diagnosis of hepatocellular carcinoma (HCC). The recently commercially available Hep Par 1 antibody (clone OCH1E5.2.10) has been reported to be a sensitive marker for HCC in paraffin embedded sections. Of non-hepatocellular tumors, occasional carcinomas have been reported to stain, most frequently gastric adenocarcinomas. This study further evaluated the staining of this antibody on a large number of neoplasms using tissue microarray technology as well as conventional tissue sections. Six hundred seventy-six tumors, including 19 cases of HCC, were tested. Eighteen of 19 cases of HCC were positive, 3 showing <5% staining. Two cases negative on the array showed focal staining when whole tissue sections from the same tumors were used. 16 of 34 cases of gastric carcinomas gave positive reactions, 4 of these showed less than 5% staining. Staining of gastric carcinomas was not limited to signet ring-type carcinomas or to areas of hepatoid differentiation. Only 1 of 11 cases of cholangiocarcinoma showed focal staining. We also noted several other tumors to stain occasionally, including adrenal cortical carcinoma (3/13), yolk sac tumor (2/9), colonic adenocarcinoma (8/106), lung carcinoma (3/52), ovarian carcinoma (5/48), and endocervical adenocarcinoma (1/5). We did not observe staining in pancreatic carcinoma (11), renal cell carcinoma (36), breast carcinoma (85), melanoma (25), or mesothelioma (5). This study supports Hep Par 1 as a useful marker in the differential diagnosis of HCC, but with significant limitations. Cautious use of this antibody in a panel with other positive (alpha fetoprotein, CD10, polyclonal carcinoembryonic antigen) and negative (epithelial membrane antigen, monoclonal carcinoembryonic antigen, CD15) markers of hepatocellular differentiation may aid in the accurate diagnosis of HCC.

Antibodies, Monoclonal↗

A pre-application drop containing carboxymethylcellulose can reduce multipurpose solution-induced corneal staining.

PURPOSE: Use of polyhexanide based multipurpose solutions (MPSs) for contact lens disinfection has been linked to low-grade corneal staining. In vitro data suggest that carboxymethylcellulose (CMC) may neutralize polyhexanides. The purpose of this investigation was to determine whether a pre-application drop of CMC reduces polyhexanide staining in vivo. METHODS: Thirty adapted soft contact lens (SCL) wearers participated in this investigator-masked, randomized, two-way cross-over study. Subjects wore a new Group II lens (alphafilcon A, 66% water) daily for 4 weeks and disinfected lenses using a MPS containing 0.0001% polyaminopropyl biguanide. A lens lubricant containing either CMC or povidone as the primary viscolyzer was applied to the lens each day before lens wear. Biomicroscopic signs and symptomatology were assessed. The difference in scores, 0 to 4 weeks and the difference between lubricants were analyzed. RESULTS: The cumulative fluorescein staining scores for combined eyes demonstrated a significant increase over time (e.g., cumulative staining score; p=0.004 and p<0.001 for CMC and povidone, respectively, matched pairs t-test, two-tailed), suggesting that for both lubricants the staining worsened with wear. This effect was expected and likely driven by the MPS. However, the mean cumulative staining scores for CMC and povidone were 2.8 and 2.6 out of 20 possible at baseline, increasing to 4.9 and 7.1 at 4 weeks, respectively. The increases were significantly different (p=0.003, matched pairs t-test, two-tailed) suggesting a greater increase in corneal staining for the povidone lubricant. The symptom scores were not significantly different, 0 to 4 weeks by regimen or between preinstillation drops. CONCLUSIONS: These results suggest that a CMC-containing preapplication drop can reduce corneal staining resulting from disinfection with a polyhexanide MPS. This result is consistent with a proposed mechanism for CMC to neutralize cationic disinfectants and may offer clinicians another means to reduce this type of corneal staining.

Adult↗

Screening of BRCA1 mutation using immunohistochemical staining with C-terminal and N-terminal antibodies in familial ovarian cancers.

We examined the subcellular localization of BRCA1 proteins using immunohistochemical staining with C-terminal (GLK-2 antibody) and N-terminal (Ab-2 antibody) monoclonal antibodies in 44 familial ovarian cancers. Among these, 24 cases were associated with 13 independent germ-line mutations of BRCA1, and loss of heterozygosity (LOH) at one or more BRCA1 microsatellite markers was found in all 21 informative tumors tested. With GLK-2 antibody, cytoplasmic staining was observed in 15 of 16 tumors (93.8%) with mutation in exon 11, and BRCA1 staining was absent in 8 of 8 tumors (100%) with mutation in exons other than exon 11. When immunohistochemical staining was performed with Ab-2 antibody, both nuclear and cytoplasmic staining were observed in 14 of 16 tumors (87.5%) with mutation in exon 11. Interestingly, nuclear staining was observed in 3 of 3 tumors (100%) with mutation downstream of exon 11, even though no staining was detected in 5 of 5 tumors (100%) with mutation upstream of exon 11. On the other hand, in familial ovarian cancers without BRCA1 mutations, nuclear staining or both nuclear and cytoplasmic staining was observed in 18 of 20 specimens (90%) and 20 of 20 specimens (100%) with GLK-2 antibody and with Ab-2 antibody, respectively. These results suggest that an immunohistochemical assay in combination with employing the C-terminal and the N-terminal antibodies appears to have potential as a reliable and useful technique for the screening of BRCA1 mutations, at least to predict the status of mutation, upstream or downstream of exon 11.

BRCA1 Protein↗

Staining for factor VIII related antigen and Ulex europaeus agglutinin I (UEA-I) in 230 tumours. An assessment of their specificity for angiosarcoma and Kaposi's sarcoma.

In this study we examined the staining reactivity of commercially available antisera to factor VIII related antigen (F VIII RAg) and Ulex europaeus agglutinin I (UEA-I) on sections from 230 formalin fixed paraffin embedded tumours. These included 196 sarcomas, 20 carcinomas and 14 angiomas. All angiomas showed positive staining for F VIII RAg; all carcinomas showed negative staining; the vasoformative areas of all angiosarcomas stained positively but only four of six angiosarcomas showed positive staining of their solid areas; of seven Kaposi's sarcomas, all showed positive staining of vessels and six showed positive staining of the spindle cell component. In the remaining 181 non-vascular sarcomas there was a false positive result in four tumours (2.2%), three of which had a history of irradiation. Pre-radiotherapy biopsies of these three tumours stained negatively with anti-F VIII RAg. UEA-I was demonstrated in all the angiomas studied, in all angiosarcomas (including the solid components) and in well-formed vessels of all Kaposi's sarcomas, but only in the spindle cell component of 3/6. However, there was an unacceptably high rate of false positive staining amongst the carcinomas and non-vascular sarcomas. In conclusion, F VIII RAg is a specific but not a sensitive marker of angiosarcomas; UEA-I is a sensitive but not a specific marker of angiosarcomas.

Adolescent↗

Comparison of the bisbiguanide antiseptics alexidine and chlorhexidine. II. Clinical and in vitro staining properties.

A blind cross-over trial was carried out to compare the tooth and tongue staining associated with the use of a 0.035% alexidine and a 0.2% chlorhexidine mouthrinse. Twenty-two volunteers were divided into two groups termed "tea drinkers" and "non-tea drinkers". All volunteers were requested to refrain from oral hygiene measures throughout two 10-day periods when they rinsed twice a day with the preparation randomly allocated for the respective period. During both periods the members of the groups excluded coffee, red wine and port from their diet. The tea drinking group consumed seven cups of tea per day. Tooth and tongue staining was recorded for extent and severity at the end of each period. The amount of stain accumulating in the two groups was similar following the use of chlorhexidine and alexidine. However, for both chlorhexidine and alexidine the extent and severity of tooth and tongue staining were significantly increased in the tea drinking group. An in vitro study of tea staining of perspex blocks exposed twice a day to 0.035% solutions of alexidine or chlorhexidine throughout a 5-day period demonstrated significantly more staining with alexidine when measured spectrophotometrically. Visually however, the differences in the specimens were minimal. Saliva treatment of the perspex did not significantly alter the staining by alexidine or chlorhexidine. The results provide further evidence for a dietary aetiology to the staining associated with cationic antiseptics. However, alexidine at the concentration used offered no advantage in reducing the side effect of staining when compared with chlorhexidine.

Biguanides↗

The use of an oxidising mouthwash to reduce staining associated with chlorhexidine. Studies in vitro and in vivo.

Discolouration of the teeth, tongue and dental restorative materials associated with the use of cationic antiseptics is still the limiting factor in long-term usage of antiplaque agents such as chlorhexidine. Oxidising agents have been used successfully to remove dental stain in animals, but products and regimens have not been evaluated in humans. These pilot studies in vitro and in vivo assessed a long-established oxidising mouthwash, based on peroxyborate, for stain removal from teeth and acrylic. The laboratory studies measured chlorhexidine tea stain removal from tooth and acrylic specimens. The oxidising agent markedly reduced staining on specimens. In a short-term clinical experiment, dental staining was forced over a 2 1/2-day period by reciprocal chlorhexidine and tea rinsing. 5 rinses with the oxidising mouthwash during the next 1 1/2 days dramatically reduced staining compared with water after rinses. Additionally, a single rinse with peroxyborate following the 5 water after rinses, again considerably reduced the residual staining. Effects on tongue staining were less impressive. Both experimental models produced findings consistent with those from the animal model. The results suggest proprietary oxidising mouthwashes could have a place in the control of dental stain associated with the use of chlorhexidine. Further studies to evaluate such products in a more conventional chlorhexidine regimen are indicated.

Adult↗

The comparative tea staining potential of phenolic, chlorhexidine and anti-adhesive mouthrinses.

Staining of teeth and mucous membranes is a well-known side-effect with chlorhexidine mouthrinses in which dietary chromogens play an important rôle. The purpose of this study was to determine whether a co-polymer anti-adhesive agent would prevent staining by a low concentration chlorhexidine solution. Additionally, the possibility that an essential oil/phenolic rinse product may cause staining was investigated. The rinses studied were the anti-adhesive alone and combined with 0.02% chlorhexidine and the essential oil/phenolic rinse. These were positioned against a positive control rise, 0.2% chlorhexidine, and a negative control rinse, water. The study was a single blind 5-treatment, randomised Latin square cross-over design, incorporating balance for carry-over effects. 15 volunteers participated and on Day 1 of each study period were rendered stain free by scaling and polishing of the teeth. Oral hygiene was suspended and 8 x per day subjects rinsed under supervision, firstly with the allocated formulation and then with 10 ml of warm black tea. On Day 4, tooth and tongue staining was scored by area and intensity (colour). A washout period of at least 3 1/2 days was permitted between treatment periods when oral hygiene was resumed. Before the study and during washouts, volunteers practised tongue brushing. Tooth and tongue staining was significantly increased with 0.2% chlorhexidine compared to the essential oil/phenolic rinse which in turn was significantly increased compared to the other 3 rinses. The antiadhesive/chlorhexidine rinse produced no more staining than the anti-adhesive or water rise. However, the parallel plaque regrowth study suggests this inhibition of staining resulted from the vitiation of the chlorhexidine activity by the antiadhesive. The methodology would appear a simple and quick way of assessing the propensity of mouthrinses to cause extrinsic staining.

Anti-Infective Agents, Local↗

Application of immunoperoxidase staining to more rapid detection and identification of rubella virus isolates.

Efforts were made to shorten the time required for detection of rubella virus in clinical materials through the use of immunoperoxidase (IP) staining. Comparative studies were performed in which specimens were inoculated in parallel into BHK-21 hamster kidney cells, which were examined by IP staining at 5 days, and into BK-13 and BS-C-1 cells, which were examined in two ways, viz., by subpassage at 7 days into BHK-21 cells and IP staining 3 days later and by subpassage at 7 days into BS-C-1 cells followed by interference testing and immunofluorescence (IF) staining on positive materials (standard method). Direct inoculation into BHK-21 cells with IP staining at 5 days permitted detection and identification of 59% of the 63 positive specimens. Toxicity of some specimens preserved with sorbitol and of certain tissue specimens reduced the number of satisfactory examinations which could be performed in this system. Virus detection and identification by IP staining on subpassaged RK-13 and BS-C-1 materials, requiring a total of 18 days, was comparable to the longer interference-IF method, requiring 17 days. Results obtained by IP staining and interference-IF showed 98% correlation on RK-13 materials and 97% correlation on BS-C-1 materials. IP staining on inoculated BHK-21 cells can be a useful method for rapid identification of a relatively high proportion of rubella-positive specimens, particularly if sorbitol-preserved specimens are avoided, and IP staining on subpassaged RK-13 and BS-C-1 materials is a highly satisfactory alternative to the longer interference-IF method.

Animals↗

Kinetics of white blood cell staining by intravascular administration of rhodamine 6G.

Rhodamine 6G is a vital dye accumulating in the mitochondria of cells. It is used in intravital fluorescence microscopy for contrast enhancement of white blood cells (WBC), enabling visualization of WBC in the microvasculature even at high center flow velocity. The aim of this study was to examine the kinetics of WBC staining after intravascular administration of rhodamine 6G in Lewis rats, Syrian golden hamsters and BALB/c mice. For this purpose, WBC were isolated from whole blood and the percentage of cells stained positively as well as their fluorescence intensity were measured by flow cytometry 5, 15, 30 and 60 min after dye administration. Injection of 0.06-0.2 mg/kg body weight of rhodamine 6G resulted in staining practically all granulocytes and monocytes over the entire observation period of 60 min. Fluorescence intensity of WBC was adequate to be detected in an experimental setup for intravital fluorescence microscopy in the hamster dorsal skinfold chamber. The degree of WBC staining was different in the species studied, yielding a higher percentage of stained lymphocytes in rats than in mice and hamsters. Staining of lymphocytes declined within 60 min after rhodamine application, the loss of fluorescent label being most pronounced in hamster cells. After 15-30 min, relative fluorescence intensity of stained lymphocytes had decreased considerably, indicating the need for reinjection of the dye or limiting microscopic analysis to approximately 15 min after rhodamine 6G administration. While the intravascular injection of rhodamine 6G results in adequate staining of granulocytes and monocytes, only a fraction of lymphoid cells are stained.

Animals↗

Immunoperoxidase staining in the differential diagnosis of parathyroid from thyroid origin in fine needle aspirates of suspected parathyroid lesions.

OBJECTIVE: To determine whether immunoperoxidase staining could be used for differential diagnosis of parathyroid from thyroid origin in fine needle aspirates of suspected parathyroid lesions. STUDY DESIGN: Immunoperoxidase staining of parathyroid hormone and thyroglobulin was performed on aspirated smears from 10 patients with parathyroid lesions (6 of 10 patients also associated with thyroid lesions) and 10 patients with thyroid lesions. All of them had surgical tissue proof of the diagnosis. RESULTS: Aspirated smears of six patients with parathyroid lesions had strong to moderate staining for parathyroid hormone and negative staining for thyroglobulin. Another four patients with parathyroid lesions had equivocal staining for parathyroid hormone and negative staining for thyroglobulin. All aspirated smears from the 16 thyroid lesions had negative staining for parathyroid hormone and positive staining for thyroglobulin. CONCLUSION: Immunoperoxidase staining of parathyroid hormone and thyroglobulin, done for each suspected parathyroid lesion, was helpful in the differential diagnosis of parathyroid vs. thyroid origin.

Adenoma↗

Morphological changes in the lungs of meconium-stained piglets.

Meconium staining of the skin is a common event associated with fetal hypoxia, stillbirths, weak-born piglets, and neonatal mortality. Aspiration of meconium leads to meconium aspiration syndrome (MAS). This study was undertaken to assess the relationship between the degree of meconium staining of the skin at birth, meconium aspiration, and pulmonary changes in porcine neonates. A total of 353 farrowing sows and 3,693 born piglets were monitored during parturition and for 15 days after delivery. Umbilical cords were classified as normal or ruptured. Meconium staining in the skin was graded as nonstained, mildly, moderately, and severely stained. Mortality from birth to 15 days of age was 8.4%. The lungs from 60 meconium-stained piglets and 60 lungs from nonstained piglets were collected and microscopically examined for meconium aspiration and inflammation. Rupture of the umbilical cord was significantly higher (P < 0.01) in meconium-stained piglets. Microscopically, 32% and 40% of the lungs had evidence of meconium for the stained and nonstained groups, respectively. The microscopic grade of meconium aspiration and inflammatory cells was not different between nonstained and meconium-stained piglets. Aspiration of meconium induced a granulomatous response in the lungs. It was concluded that the grade of meconium staining is a good indicator of fetal hypoxia, but not a good predictor for meconium aspiration and MAS in piglets.

Animals↗

Selective staining of nucleic acids by osmium-ammine complex in thin sections from lowicryl-embedded samples.

Osmium-ammine (OA)/SO2 selectively contrasted RNA- and DNA-containing structures in thin sections from Lowicryl-embedded samples. No cell structures were stained after Epon embedding. RNAse and DNAse digestion experiments demonstrated that only RNA and DNA were stained in Lowicryl thin sections. Protease digestion did not modify the staining reaction. The very fine end-reaction produced a very high resolution of the stained structures. The staining reaction was not due to the presence of SO2 but to the low pH of the solution (ranging from 1.5-2.2). OA in glycine buffer, pH 1.5, selectively contrasted nucleic acids. Electrostatic bonds between nucleic acids and OA complex were probably involved in the staining reaction. Increasing the pH value of the staining medium resulted in loss of OA specificity for nucleic acids. The high electrolyte concentration of the staining medium hindered the staining reaction.

Acrylic Resins↗

Metabolic toxicity of fluorescent stains on thawed cryopreserved bovine sperm cells.

Several fluorescent probes, including derivatives of carboxyfluorescein, carbocyanine, ethidium, and rhodamine, have been used to assess sperm viability. However, the effects of these fluorescent dyes on the metabolic activity of sperm cells have not been systematically examined. This study was conducted to determine the effect of specific fluorescent stains on the metabolic processes of sperm. Cryopreserved bovine sperm cells were thawed, fluorescently stained, and examined using metabolic and flow cytometric techniques. Sperm were stained with either rhodamine 123 (Rhod-123), the aliphatic cell-tracking compound PKH2-GL, dihydro-ethidium (HED), the bisbenzimide stain Hoechst 33342 (Ho33342), or left unstained. The stained samples were compared for metabolic activity, cell staining pattern, and fluorescent intensity over a 180-min period. Samples stained with HED, Ho33342, and PKH2-GL had less oxygen uptake when compared with the unstained sperm samples (p greater than 0.05). Unstained samples and samples stained with Rhod-123 had similar oxygen consumption. The carbon dioxide produced during the 180 min was not different between controls and stained samples. Therefore, some fluorescent probes inhibit the oxygen metabolism of thawed, cryopreserved bovine sperm cells.

Animals↗

Histopathological evaluation of ocular microsporidiosis by different stains.

BACKGROUND: There is limited data on comparing stains in the detection of microsporidia in corneal biopsies. Hence we wanted to evaluate various stains for their ability to detect microsporidia in corneal tissue sections. METHODS: Four cases diagnosed with microsporidiosis on Hematoxylin and Eosin and Periodic Acid Schiff's stained sections of the corneal button between January 2002 and December 2004, were included. Further sections were prospectively stained with calcofluor white, Gram, Giemsa, Masson's trichrome, acridine orange, Gomori's methenamine silver, Gram's chromotrope and modified acid fast stain. The stained sections were analyzed for the spore characteristics in terms of size, shape, color contrast, cell wall morphology, waist band in cytoplasm and ease of detection. RESULTS: All sections showed microsporidial spores as 3-5 microm, oval bodies. 1% acid fast, Gram's chromotrope and GMS stains provided a reliable diagnosis of microsporidia as diagnostic waist band could be identified and good contrast helped distinguish the spores from inflammatory debris. CONCLUSION: Considering the ease of performance, cost effectiveness and rapidity of the technique, 1% acid fast stain and Gram's chromotrope stain are ideal for the detection of microsporidia.

Journal Article↗

Poorly differentiated carcinoma of unknown primary site: clinical usefulness of immunoperoxidase staining.

To assess the clinical utility of immunoperoxidase tumor-cell staining in patients with poorly differentiated carcinoma of unknown primary site, we performed a battery of stains on tumors from 87 patients treated between August 1978 and April 1983. Poorly differentiated carcinoma or poorly differentiated adenocarcinoma was diagnosed on the basis of light microscopic examination, and all patients were treated before the technology of immunoperoxidase staining was routinely used. Therefore, results of immunoperoxidase staining can be correlated with clinical outcome in this group of similarly treated patients with a long median follow-up. Immunoperoxidase staining confirmed the diagnosis of poorly differentiated carcinoma in 49 patients (56%) and yielded other diagnoses in 14 patients (16%): melanoma, eight; lymphoma, four; prostatic carcinoma, one; and yolk sac carcinoma, one. In 24 patients (28%) the immunoperoxidase staining pattern was inconclusive; electron microscopy was usually helpful in clarifying the diagnosis in these patients. Seventy-five patients (86%) received combination chemotherapy with a cisplatin-based regimen, and 24 patients (28%) had a complete response. Nine of these patients were later given specific diagnoses by immunoperoxidase staining (lymphoma, four; melanoma, four; yolk sac tumor, one). All patients with an immunoperoxidase diagnosis of lymphoma also had clinical features compatible with lymphoma and are long-term survivors. Patients with immunoperoxidase features suggesting melanoma were surprisingly responsive to chemotherapy, with three of seven complete responses and two long-term survivors. Patients with melanoma diagnosed by immunoperoxidase staining should not be excluded from a trial of cisplatin-based therapy. Immunoperoxidase staining is useful in the routine evaluation of metastatic poorly differentiated carcinoma of unknown primary site, as it can occasionally suggest the lineage of the tumor and have specific therapeutic implications.

Adenocarcinoma↗

Evaluation of Coomassie blue staining of the acrosome of equine and canine spermatozoa.

OBJECTIVE: To evaluate Coomassie blue staining of the acrosome of equine and canine spermatozoa. SAMPLE POPULATION: Spermatozoa of 5 mixed-breed male dogs and 3 Thoroughbred stallions. PROCEDURE: Various proportions of intact and acrosome-damaged spermatozoa were fixed in 2% phosphate-buffered formaldehyde or 4% paraformaldehyde, smeared onto glass slides, and stained with Coomassie blue stain. Acrosomal status (damaged vs intact) was also assessed by use of flow cytometry after staining with fluorescein isothiocyanate-conjugated Pisum sativum agglutinin (FITC-PSA) and propidium iodide. Comparisons were made between percentages of expected and observed acrosome-intact spermatozoa in different proportions of live and flash-frozen samples; the percentages of acrosome-intact spermatozoa as determined by use of Coomassie blue staining and flow cytometry were also compared. RESULTS: Strong correlations were found between the expected and observed distributions of acrosome-intact spermatozoa when fixed in 4% paraformaldehyde (r2 = 0.93 and 0.89 for canine and equine spermatozoa, respectively) as well as between Coomassie blue-stained cells and those stained with FITC-PSA and assessed by use of flow cytometry (r2 = 0.96 and 0.97 for canine and equine spermatozoa, respectively). However, in canine samples that were fixed in 2% phosphate-buffered formaldehyde, these correlations were weak. CONCLUSIONS AND CLINICAL RELEVANCE: Staining with Coomassie blue stain was a simple and accurate method to evaluate the acrosome in equine and canine spermatozoa after fixation in 4% paraformaldehyde. This assay should be useful in routine evaluation of semen samples from these species.

Acrosome↗