A feasible method for llama embryo paraffin embedding

Maternal recognition of pregnancy in llamas would be related to a still-unknown signal secreted by the embryo to extend the corpus luteum lifespan. In order to study llamas reproductive physiology and early pregnancy aspects, research on embryos between 8 and 12 days post-mating is essential. However, there is a lack of information on llama embryo paraffin embedding, a procedure that exhibits difficulties, mostly for embryo manipulation. This work presents an accessible, easy and rapid protocol for embedding llama blastocysts. Uterine flushings from four superovulated llamas were performed to recover embryos 8 days post-mating. Each embryo was fixed in 1 ml of 10 % neutral buffered formalin for 24 h, at 4 °C, and transferred to 70 % ethanol. Later, they were submitted to a series of baths: 500 µl PBS with 5 % bovine foetal serum solution (PBS+BFS) for 1 min; zona pellucida permeabilization with 500 µl PBS with 0.2 % Triton X-100 for 20 min; 500 µl PBS+BFS for 1 min; dehydration on baths with ascendent grades of ethanol (70 %, 96 %, and 100 % twice) for 10 min each; and bath of butanol for 5 min. Finally, embryos were embedded in a pellet of paraffin and afterwards in a paraffin block. Complete sections from 8 of 13 grade I and II embryos were obtained. In conclusion, by applying this method, good quality sections with preserved embryo morphology suitable for histological, histochemical or immunohistochemical approaches would be possible to acquire.     

Acute unintentional intoxication with paraffin in a 25-year old patient – Clinical case report

“Fire-breathing” or “fire-eating” is a special kind of street art where the acts are always stunning, spectacular and amazing. People exhibiting this kind of show are professionals, not rare amateurs, who use different kind of fuels, usually hydrocarbons, in order to produce a pillar of fire. Intoxications caused by ingestion or inhalation of liquid paraffin, used as a fuel while performing, are numerous and various. We present a clinical case report of a 25-year old, previously healthy, amateur “fire-breather”. During October, 2010 this young men arrived at the Emergency Unit of the University Clinic for toxicology and Urgent Internal Medicine in a severe clinical condition, after his unsuccessful attempt to perform real “fire-breathing”. He had fever, strong headaches, mild abdominal and chest pain and he also had difficulties breathing and persistent dry cough. The patient was extremely dyspneic with peripheral cyanosis and shortness of breath.“Fire-breathers” must be viewed as a population at risk of paraffin-induced pneumonia, which has low mortality rate, but still is related with numerous and various chronic complications. Our patient was first in a life threatening, extremely serious clinical condition which was urgently treated with appropriate vigorous and effective therapy. This therapeutic protocol led to successful full recovery of these young men, who luckily didn't exhibit any chronic complications.     

吞咽石蜡油置胃管法的有效性探讨

目的探讨吞咽石蜡油置胃管法的临床效果。方法将102例患者随机分为观察组（52例）和对照组（50例）,观察组置管之前吞咽10～15ml石蜡油后迅速将胃管插入胃内,对照组采用常规置管法。观察两组患者一次置管成功率,置管过程中恶心呕吐发生率,留置胃管后喉部干痛症状发生率。结果观察组和对照组一次置管成功率分别为92．30％和74．00％．置管过程中恶心呕吐发生率分别为19．23％和68．00％,留置胃管后喉部干痛发生率分别为11．54％和56．00％。两种方法观察指标比较采用χ2检验,P值均小于0．05,差异有统计学意义。结论吞咽石蜡油置胃管法能提高一次置管成功率,减轻患者痛苦,是一种较理想的置管方法。     

乳腺病变冰冻切片误诊及延迟诊断分析

目的 分析乳腺病变冰冻误诊及延迟诊断原因,提高冰冻诊断准确性.方法 收集2007年1月至2010年1月乳腺术中冰冻切片,把乳腺病变分为良性病变、交界性病变和恶性病变,将冰冻切片与石蜡切片对比分析.结果 乳腺冰冻诊断假阴性(误诊)2例,分别为神经内分泌癌和乳头状癌,其主要原因为对疾病认识不足和取材不到位;延迟诊断6例,分...     

乳腺癌冰冻的病理诊断分析

目的:提高乳腺癌冰冻切片的分析能力,减少乳腺癌的漏诊. 方法:回顾性对比观察243例乳腺冰冻切片和石蜡切片. 结果:冰冻切片诊断为良性215例,恶性肿瘤25例,推迟诊断3例.石蜡切片中良性218侧,恶性肿瘤26倒,其中冰冻漏诊3例,假阴性诊断率为1.8%.结论:提高切片质量,掌握正确的取材方法,提高对乳腺癌的识别和鉴别...     

Immunohistochemical Determination of Hepatic Cytochrome P-4502E1 in Formalin-Fixed, Paraffin-Embedded Sections

Cytochrome P-4502E1 (2E1) is inducible by chronic ethanol consumption that results in enhanced activation of anesthetics and commonly used drugs (such as acetaminophen) to hepatotoxins. Therefore, assessment of hepatic 2E1 is needed in prescribing these drugs for the management of alcoholic patients. Currently, measurement of 2E1 requires either immunohistochemistry on frozen sections or Western blot (WB) analysis of homogenized tissue in excess of that needed for pathology. To obtain a more widely applicable method, we developed a procedure to detect 2E1 by immunohistochemistry in formalin-fixed, paraffin-embedded liver biopsies obtained routinely for diagnosis. Data were collected from rats fed ethanol-containing or control liquid diets for 3 weeks. lmmunostaining was performed using anti-human rabbit 2E1 antibody as the primary antibody, and the immunoreaction was detected by the avidin-biotin immunoperoxidase method after treating sections with target unmasking fluid, an antigen retrieval buffer that enhanced the staining of 2E1. In control rats, 2E1 staining was weak and perivenular. After ethanol feeding, it showed a lobular gradient, strongest perivenular and weakest periportal, similar to that seen in frozen sections. The staining intensity was scored as: 0 (no staining) to 3 (strong staining). The zonal staining was scored as follows: 1 = perivenular zonal staining, 2 = midzonal, and 3 = panlobular. With the product of the two scores, a significant difference was found between alcohol-fed and control rats (5.1 ± 0.3 vs. 0.8 ± 0.2, p < 0.001). 2E1 assessments by WB were also significantly different for these rat pairs (68.5 ± 2.1 vs. 7.9 ± 0.8 arbitrary units/mg protein, p < 0.001), with a parallel increase of immunostaining scores and WB measurement of 2E1 content. This immunohistochemical method was then validated in 14 paraffin-embedded percutaneous human liver biopsy samples. In livers of nonalcoholics, 2E1 staining was seen in the perivenular zone only, whereas in samples of alcoholics, the staining was perivenular to midzonal and sometimes periportal. A significant correlation between the zonal staining scores (r_s= 0.67, p < 0.005) or intensity ± zonal staining scores (r_s= 0.79, p < 0.001) and WB analysis was found. The immunohistochemical assessments of 2E1 expression in formalin-fixed, paraffin-embedded sections from livers of alcoholics was found to correlate with WB analysis, and lobular distribution was consistent with that seen in frozen sections. The proposed method should therefore be useful for the assessment of 2E1 content in paraffin-embedded liver samples, thereby aiding in the management of heavy drinkers.     

石蜡油滴鼻减轻鼻饲患者鼻咽部黏膜损伤作用的研究

目的探讨减轻鼻饲患者鼻咽部黏膜损伤的作用。方法选取如东县人民医院2015年5月至2017年1月105例留置16Fr复尔凯鼻胃管鼻饲患者为研究对象,使用抛硬币的方法随机分为对照组和试验组。对照组采用生理盐水口腔护理、雾化吸入、棉签蘸温开水清洁双侧鼻腔等常规护理,试验组在此基础上给予医用石蜡油滴鼻;观察两组患者置管后3天、1周、2周及拔管后鼻黏膜出血、血痂、溃疡以及咽喉部红肿、脓苔等情况。结果试验组患者置管1周、2周及拔管后鼻咽部黏膜损伤发生率显著低于对照组（均P<005）。结论石蜡油滴鼻能有效减轻鼻饲患者鼻咽部黏膜的损伤,值得临床推广应用。     

Evaluation of five DNA extraction methods for detection of H. pylori in formalin-fixed paraffin-embedded (FFPE) liver tissue from patients with hepatocellular carcinoma

Since Helicobacter spp. DNA was identified in liver tissue resected from patients with hepatocelullar carcinoma (HCC), researchers have suggested a role of this bacterium in hepatic carcinogenesis. Archives of formalin-fixed, paraffin-embedded (FFPE) tissues represent an extraordinary source for clinical studies providing many advantages. However, DNA extraction from FFPE tissues is laborious, time-consuming and still remains a challenge. The aim of this study was to evaluate five protocols for DNA extraction from FFPE liver obtained from patients with HCC in order to detect Helicobacter pylori DNA. These methods were: (1) QIAamp FFPE Tissue Kit, (2) QIAamp DNA Mini Kit, (3) Wizard SV Genomic DNA Purification System, (4) RealiaPrep FFPE gDNA Miniprep System and (5) phenol–chloroform. H. pylori detection was performed using 16S rRNA gene amplification by PCR. The highest total amount of DNA was obtained using the phenol–chloroform method. Analyses of 16S rRNA gene amplification did not show statistically significant differences among the methods (p = 0.466), although the highest percentage of positive cases (70%) was found in samples extracted with phenol-chloroform. We suggest that of the five methods evaluated, phenol/chloroform is the most suitable for detection of H. pylori in FFPE liver from patients with HCC.     

子宫肿瘤病理学临床研究

目的：探讨子宫肿瘤患者的冰冻切片的病理学诊断方法的可靠性与局限性。方法：分析我院收入的80例子宫肿瘤病人的完整病理切片资料，比较冰冻切片与石蜡切片病理学诊断的相符合情况、确诊率、未确诊率及误诊率。结果：冰冻切片组与石蜡切片组确诊率、未确诊率及误诊率之间的差异有统计学意义（P〈0．05），冰冻切片组子宫肿瘤确诊率明显高于石蜡切片组。结论：冰冻切片对子宫肿瘤的确诊率高，结果准确，但对于子宫内膜早期肿瘤存在局限性，临床医生要结合患者病史做出合理诊断，以防仅凭冰冻切片结果而误诊。     

Tissue expansion of lung bronchi due to tissue processing for histology – A comparative analysis of paraffin versus frozen sections in a pig model

AimTissue shrinking due to fixation and processing is well known. However, the degree of shrinking varies significantly with the tissue type as well as the processing method and is not well studied in various tissues. In daily pathological routine workflow, histological specimens from frozen and paraffin sections are performed from the same tissue. In the present study we compared the thickness of bronchus walls obtained from paraffin and frozen sections.MethodsPig lungs were frozen in ventilated condition in liquid nitrogen and 36 bronchi were isolated after dissection. Frozen sections of 5 μm thickness were performed and the remaining tissue was fixed and embedded in paraffin after fixation in 4% formalin. Frozen and paraffin sections from the same cutting edge were analysed after haematoxylin and eosin staining by measuring the wall thickness of the bronchi using high power fields of 400-fold magnification. In each bronchus 40 measurements were implemented at different wall positions distributed over the entire wall area. Summed up, in each group 1440 wall measurements were performed in total. Statistical analysis was conducted using the Wilcoxon test and t-test as well as Pearson’s correlation coefficient with a significance level at P < 0.05.ResultsThe bronchial wall thickness was significantly (p < 0.001) smaller in frozen sections (median: 0.50 mm; min: 0.37 mm; max: 0.97 mm) compared to paraffin sections (median: 0.58 mm; min: 0.35 mm; max: 1.06 mm). The median difference between paraffin and frozen sections was 0.05 mm (min: -0.11 mm; max: 0.22 mm). The wall thickness ratio of both groups was as follows: frozen/paraffin section = 0.8609, thus yielding a difference between paraffin and frozen of 13.91%. High correlation was found between wall thickness measurements on paraffin and frozen sections (R = 0.87, p < 0.001).ConclusionsThe bronchus wall thickness in the frozen section was 14% reduced compared to the paraffin section. In routine pathology as well as in scientific studies these results are of relevance, as airway wall thickness represents a relevant marker for pathological interpretation, especially using CT image techniques.