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Microscopic findings in key tissues are often critical to determine the cause of death in medical autopsies. The overall quality of histologic sections depends on numerous pre-analytic factors, among which are tissue section size and thickness. We designed a prospective quality improvement study to determine whether a simple intervention of formalin pre-fixation of myocardium, liver, and kidney tissues could improve the ease of cutting and quality of autopsy histologic sections as assessed by histotechnicians and pathologists. Of 46 autopsies included in the study, 21 were randomly assigned to formalin pre-fixation, and 25 underwent routine sectioning without formalin pre-fixation. A significant improvement in overall quality score by histotechnicians was detected in the sections from pre-fixed autopsy tissues compared to the control group (p=0.0327). There was no significant difference in quality score between the two groups as assessed by pathologists. Our autopsy quality improvement study demonstrates that a simple, low-cost intervention of formalin pre-fixation of fresh autopsy tissues for 90 minutes could significantly improve the overall quality of sections submitted for histologic processing.
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Humans , Male , Female , Autopsy/methods , Histological Techniques/methods , Tissue Fixation/methods , Quality ImprovementABSTRACT
Objective@#To investigate the impact of ultrasonic assisted rapid processing technique combined with the environment friendly reagent (which can be utilized in fixing,dehydrating and clearing) on processing tumor biopsy specimens and the subsequent target detection.@*Methods@#Postoperative tissue samples of 56 cases of breast cancer, colorectal cancer, lung cancer, stomach cancer, liver mass, bladder mass, uterus mass were obtained at the National Cancer Center, Cancer Hospital, Chinese Academy of Medical Sciences from February to April, 2017. Three specimens ranging in size from 1 to 3 mm were collected from each sample, and were separated into control group (traditional tissue-processing method); experiment group 1 (3.7% neutral buffered formaldehyde fixation, composite environment friendly reagent and ultrasonic assisted rapid processing) and experimental group 2 (composite environment friendly reagent direct fixation, higher temperature and longer time for tissue processing). Two pathologists blinded to the experimental groups scored totally the nuclear, cytoplasmic, and membrane staining of 43 cases of immunohistochemistry (IHC), four HER2 fluorescence in situ hybridization (FISH), 20 extracted DNA quality and four EGFR gene mutation detection in lung adenocarcinoma; the results were compared with the control group.@*Results@#There was no difference in the IHC staining, HER2 FISH, the DNA quality, and EGFR genetic results between experimental group 1 and control group. For experiment group 2, comparing results of IHC staining, HER2 FISH and the quality of DNA, there was no obvious difference from control group and experiment group 1, but might show an increase in the background of IHC staining. The difference between the treatment temperature and time in the experimental group 2 did not affect the results of the gene mutation detection.@*Conclusions@#Environment freindly reagent and ultrasonic assisted rapid processing equipment could be used for rapid processing and diagnosis for tumor biopsies. Using complex environment-friendly reagents supplement fixation, higher treatment temperature and longer treatment time do not significantly affect the IHC, FISH and molecular detection accuracy.
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Introdução: Mastopexia associada à inclusão de implante é uma situação desafiadora para o cirurgião plástico. O objetivo é descrever a colocação de implante submuscular com descolamento anatômico mais pexia firme do tecido glandular usando pontos de fixação do tecido mamário ao muscular e analisar os resultados estéticos das pacientes operadas. Método: Foram realizadas 23 mastopexias com implantes no período entre abril de 2015 e julho de 2017, pelo mesmo cirurgião, sendo as mamas das pacientes marcadas previamente, na posição sentada. Realizou-se incisão no sulco mamário e descolamento até o polo superior da mama no plano subfascial, fixação da glândula ao músculo peitoral maior com 9 a 12 pontos. A seguir, iniciou-se a dissecção do músculo peitoral maior através de sua origem costal e transição com os músculos reto abdominal e serrátil, liberando amplamente na porção inferior. Introduziu-se o implante e completou-se a mastopexia. Os tamanhos dos implantes variaram de 255ml a 355ml. Fotos das mamas de 12 pacientes foram avaliadas por dois cirurgiões plásticos e dois leigos, nos seguintes parâmetros: resultado estético, simetria das aréolas e grau de ptose mamária. As avaliações podiam ser Ruim, Razoável ou Bom. Resultados: A técnica cirúrgica mostrou-se reprodutível, apenas 1 caso de hematoma unilateral, nenhuma infecção, queixas de dor discretas. Apenas um caso foi considerado, por um único avaliador, como Razoável; as demais avaliações consideradas como Bom. Conclusão: O tratamento de ptoses mamárias com colocação de implante submuscular acrescido de pexia da glândula ao músculo peitoral é uma técnica reprodutível e com bons resultados estéticos.
Introduction: Mastopexy associated with implant placement is challenging for plastic surgeons. The objective is to describe the placement of a submuscular implant with anatomical detachment in combination with stable fixation of the breast tissue to the pectoralis muscle and analyze the aesthetic results. Method: Twenty-three mastopexy procedures with implants were performed from April 2015 to July 2017 by the same surgeon, and surgical markings were made in the breasts of the patients in a seated position. An incision was made in the inframammary fold, and the breast tissue was elevated to the upper pole in the subfascial plane and attached to the pectoralis major muscle using 9-12 stitches. Subsequently, the inferior margin of the pectoralis major muscle and the transition from the rectus abdominis muscle to the serratus muscle were dissected to expose the muscle. The implant was introduced and mastopexy was completed. Implant size ranged from 255 mL to 355 mL. Photographs of the breasts of 12 patients were evaluated by two plastic surgeons and two non-medical subjects, who considered the aesthetic results, symmetry of the nipple-areola complex, and degree of breast ptosis. The results were scored as unsatisfactory, satisfactory, or good. Results: The surgical technique was reproducible; there was only one case of unilateral hematoma, no implant infections, and only complaints of mild pain. Only one case was scored as satisfactory by one evaluator, whereas the results of the other cases were considered good. Conclusion: The treatment of breast ptosis with the placement of a submuscular implant in combination with fixation of the breast to the pectoralis major muscle is reproducible and yields good aesthetic results.
Subject(s)
Humans , Female , Adult , Breast/surgery , Tissue Fixation/methods , Mammaplasty/adverse effects , Mammaplasty/methods , Breast Implants/adverse effects , Plastic Surgery Procedures/adverse effects , Plastic Surgery Procedures/methods , Breast , Tissue Fixation , Mammaplasty , Breast Implants , Plastic Surgery ProceduresABSTRACT
Objective@#To compare the performance of Miseq and Ion Torrent PGM platforms and library construction method for next-generation sequencing (NGS) technology for formalin-fixed and paraffin-embedded (FFPE) samples.@*Methods@#A total of 204 FFPE cancer samples including 100 non-small cell lung cancers at the First Affiliated Hospital of Zhejiang University, and 104 colorectal cancers at West China Hospital of Sichuan University were retrospectively selected from January 2013 to December 2016. By using the same samples, DNA was extracted, and the same amount of DNA was used for library construction with the same kit, and sequenced on Miseq and Ion Torrent PGM respectively, after passing the quality control. Any discordant mutations between two platforms were validated by amplified refractory mutation system-polymerase chain reaction (ARMS-PCR) method and Sanger sequencing.@*Results@#A total of 204 FFPE samples were included and 197 samples were successfully analyzed by both platforms. The number of reads generated by the samples on Miseq platform sequencing was higher than PGM platform (median 391 634 vs. 298 030, P<0.01). Alignment with human reference genome showed that the mapping rate of Miseq platform was higher than PGM platform (median 100.0% vs. 99.7%, P<0.01). The median sequence depth of samples on Miseq was higher than PGM platform (median 853× vs. 698×, P<0.01). A total of 236 mutations were detected by two platforms, of which 221 were detected on both platforms, with a 93.6% concordance. Miseq platform detected 11 mutations not detected on PGM platform, while PGM platform detected 4 more mutations not detected on Miseq platform. With validation by ARMS-PCR and Sanger sequencing, Miseq platform was more reliable for low-frequency mutations. The main reasons for the discordant mutations between two platforms were that mutation frequency on undetected platform was lower than mutation reporting range (5%) and FFPE samples were stored for a long time.@*Conclusions@#Compared with PGM, Miseq platform shows higher sequencing quality in terms of the number of reads, alignment results and coverage depth, and the test results are more reliable. In clinical practice, the appropriate platform should be chosen based on sample size and actual throughput requirements to aid in the molecular characterization of tumors.
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Abstract Most Departments of Pathology around the world have a considerable archive of formalin-fixed paraffin-embedded (FFPE) tissue suitable for molecular assessment. This article points out the potential DNA damage that may occur if basic steps are not followed during processing and storage of these samples. Furthermore, it hopes to establish parameters to optimize quality and quantity of DNA extracted from FFPE tissues.
Resumo A maioria dos Departamentos de Patologia em todo o mundo têm um considerável acervo de tecidos embebidos em parafina e fixados em formalina, que são passíveis para análises moleculares. Este artigo apresenta os danos ao DNA que podem ocorrer se passos básicos não forem seguidos durante o processamento e armazenamento destas amostras. Além disso, procura estabelecer parâmetros para otimizar a qualidade e quantidade do DNA extraído de tecidos FFPE.
Subject(s)
Humans , Tissue Fixation , Paraffin EmbeddingABSTRACT
Objective:To observe the difference of the human telomeres RNA component (hTERC) genes’amplification in the cervical tissue by applying the environment-friendly fixative poly hydroxy acrylic acid and the transparent dewaxing solution Van-clear separately or jointly to replace the traditional fixative 4% (volume fraction)neutral buffered formalin and the conventional transparent dewaxing solu-tion xylene in the use of fluorescence in situ hybridization (FISH)for detection.Methods:In the study, 255 cases of cervical tissue specimens submitted by the Department of Gynecology in Zhongshan Boai Ho-sipital were collected from Mar.2013 to Apr.2015.Four samples were taken from the same lesion site. All the cases were divided into 4 groups and named group A,B,C,and D.Group A used 4% neutral buffered formalin fixed and xylene dewaxing to make slices.Group B used poly hydroxy acrylic fixed and xylene dewaxing to make slices.Group C used 4% neutral buffered formalin fixed and Van-clear trans-parent to make slices.Group D used poly hydroxy acrylic fixed and Van-clear transparent dewaxing to make slices.The amplification of hTERC genes in the four groups of cervical specimens was also detected by FISH technique.Results:When the hTERC genes were detected by FISH method under the fluore-scence microscope,it was obvious that the tissue profile and the background of group A,B,C and D were all clear.The probe was fixed in the accurate position so that the bright red or green fluorescence signals were easily found in these four groups.Compared with the positive rate of group A,there was no statistical significance in that of group B,C and D (P>0.05).At the same time,the coincidence rate of the FISH results was high,which showed that the new environment-friendly reagent had no significant difference in the detection of cervical hTERC genes by FISH technique.Conclusion:It is possible for the environment-friendly reagent poly hydroxy acrylic acid and Van-clear to replace 4%neutral buffered for-malin and xylene separately or jointly to detect the cervical hTERC genes by adopting FISH technique.
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Experimental pathology is an important part of life science research associated with animal experiment. Acquisition and fixation of optimum specimen and subsequent section of paraffin embedded tissue and dyeing are key factors playing important role in reliability, authenticity of pathological diagnosis.This paper summarizes the problems encountered in pathological section making of animal experiment and it correspond solutions.
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The purpose of our study was to compare the clinical results between arthroscopic bone fixation on intertubercular groove using suture anchor and soft tissue fixation at the rotaor interval for biceps tenodesis when partial tear or instability of biceps tendon accompanied with rotator cuff tear. From January 2010 to January 2012, 34 cases who were performed biceps tenodesis for partial tear or instability were enrolled in our study. Mean follow-up period was 30.2 months. Bone fixation using suture anchor was performed in 18 cases, and soft tissue fixation was performed in 16 cases. Clinical result was evaluated by pain visual analogue scale (VAS), Speed test, Yergason test, muscle strength, and Constant score. Pain VAS of cases with soft tissue fixation was significantly higher than that of cases with bone fixation at 6 months and final follow-up. Positive results for the final follow-up Speed and Yergason test were checked in 4 cases (25%) with soft tissue fixation and 1 (5.6%) with bone fixation. The Popeye deformity was seen in 4 cases (25%) with soft tissue fixation and 2 (11%) with bone fixation. Constant score was improved 47 to 78 in cases with soft tissue fixation and 48 to 86 in cases with bone fixation. In patient with partial tear or instability of biceps tendon accompanied with rotator cuff tear, biceps tenodesis using soft tissue fixation showed worse result compared with bone fixation because of long duration of the pain. Therefore, when performing the biceps tenodesis, bone fixation will be recommended.
Subject(s)
Humans , Congenital Abnormalities , Follow-Up Studies , Muscle Strength , Rotator Cuff , Shoulder , Suture Anchors , Tendons , Tenodesis , Tissue FixationABSTRACT
PURPOSE: The use of graft tissue fixation using bioabsorbable interference screws (BISs) in anterior cruciate ligament (ACL) reconstruction offers various advantages, but limited pullout strength. Therefore, additional tibial fixation is essential for aggressive rehabilitation. We hypothesized that additional graft tissue fixation using bioabsorbable suture anchors (BSA) would provide sufficient pull-out strength. MATERIALS AND METHODS: Twenty four fresh frozen porcine distal femur and patellar tendon preparations were used. All specimens were divided into three groups based on additional fixation methods: A, isolated BIS; B, BIS and BSA; and C, BIS and post cortical screw. Tensile testing was carried out under an axial load. Ultimate failure load and ultimate failure load after cyclic loading were recorded. RESULTS: The ultimate failure loads after load to failure testing were 166.8 N in group A, 536.4 N in group B, and 438 N in group C; meanwhile, the ultimate failure loads after load to failure testing with cyclic loading were 140 N in group A, 466.5 N in group B, and 400 N in group C. Stiffness after load to failure testing was 16.5 N/mm in group A, 33.5 N/mm in group B, and 40 N/mm in group C. An additional BSA fixation resulted in a significantly higher ultimate failure load and stiffness than isolated BIS fixation, similar to post screw fixation. CONCLUSION: Additional fixation using a BSA provided sufficient pullout strength for ACL reconstruction. The ultimate failure load of the BSA technique was similar to that of post cortical screws.
Subject(s)
Animals , Anterior Cruciate Ligament Reconstruction/methods , Bone Screws , Suture Anchors , SwineABSTRACT
Objective To solve the problem that the rat retina paraffin sections are easily exfoliated from the slides and each layer are easily separation and fracture,we need to find a way to improve the retina paraffin section method and evaluate the tissue fixation. Methods We used 4 fixation liquids including 10%paraformaldehyde,4%paraformaldehyde, 4% paraformaldehyde and 95% alcohol and glacial acetic acid mixed liquid (FAA fixatiue solution ) combined with paraformaldehyde to fix the retinal tissue, and observed the fixation efficacy under microscope after HE staining. Results The effects of 10%paraformaldehyde and 4%paraformaldehyde fixed samples showed moderate separation and fracture of retina,but the HE staining retinal slices pre-treated by the FAA fixafive solution had bright and uniform color,although occasionally some parts of the retina were exfoliated from the slide, but it was not easy to take off,and had complete structure without separation and rupture. Conclusion The retina paraffin section fixed by FAA ixafive solution with 4% paraformaldehyde is superior to pure paraformaldehyde, and the paraformaldehyde concentration has no obviously influence on HE staining results.
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Objective To qualitatively and quantitatively compare the effect of tracheal inflation method, vascular perfusion method and combined trachea-vascular perfusion method on alveolar structure and cell antigen preservation. Methods Fifteen healthy Sprague Dawley rats were randomly divided into three groups (5 each): tracheal inflation group, vascular perfusion group and combined trachea-vascular perfusion group. After lungs were respectively fixed with the methods mentioned above, tissue blocks were paraffin embedded and sectioned. HE staining and immunofluorescence staining were then performed for qualitative observation and stereological analysis of the fixation effect. Results Qualitative study showed that the fixation effect of cytoplasm and nuclear antigen was better in vascular perfusion group and combined trachea-vascular perfusion group than in tracheal inflation group. Stereological analysis showed that the alveolar linear intercept in combined trachea-vascular perfusion group was longer than that in tracheal inflation and vascular perfusion group (P<0.05). The lung parenchyma volume density and alveolar septum thickness of combined trachea-vascular perfusion group were smallest among all the three groups, followed by tracheal inflation group and vascular perfusion group in turn (P<0.05). Conclusions The combined trachea-vascular perfusion method could not only preserve better alveolar structure but also display perfect cellular antigen. Therefore, it is suitable for the morphometry-based researches on lung pathological and clinical diagnosis.
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BACKGROUND: The preservation of optimized DNA and its extraction from formalin-fixed, paraffin-embedded (FFPE) tissues are important issues. There has been some doubt over whether 10% neutral-buffered formalin is an ideal fixation solution for DNA preservation over non-buffered formalin, as conventionally recommended. In this study, the correlation between the efficiency of DNA extraction from FFPE tissues and buffered formalin was evaluated. METHODS: Several tissues with same conditions except fixatives were fixed in four different formalin solution groups and were routinely processed as paraffin-embedding protocols. DNAs were extracted from four different FFPE tissues that were stored for over 3 months and over 9 months. The quantity and quality of the DNAs were assessed with a NanoDrop ND-1000 spectrophotometer, and the polymerase chain reaction (PCR) amplification and degradation were analyzed via microchip electrophoresis. KRAS mutation analysis and microsatellite instability (BAT25) PCR were performed with each sample. RESULTS: The results showed no remarkable difference in the four groups. CONCLUSIONS: The study findings demonstrate that DNA preservation is fairly unaffected by a neutral buffer where there is short formalin manufacture period and an adequate formalin fixation time before embedding in paraffin.
Subject(s)
DNA , Electrophoresis, Microchip , Fixatives , Formaldehyde , Microsatellite Instability , Paraffin , Pathology, Molecular , Polymerase Chain Reaction , Tissue Fixation , Tissue PreservationABSTRACT
Introdução: Para o adequado estudo anatomopatológico dos tecidos obtidos dos pacientes é crucial a boa técnica para a melhor evidenciação das células pesquisadas. É imprescindível que não só a técnica ideal de coloração/marcação seja utilizada, mas também a correta técnica de fixação da peça cirúrgica. Em trabalho prévio pesquisando miofibroblastos e contração na pele restaurada de áreas doadoras de enxertos de pele de espessura parcial, verificamos baixa densidade de miofibroblastos, ao contrário dos estudos em tecido de granulação. Investigando as causas disso, e baseando-se em artigo específico sobre a eficácia da evidenciação imuno-histoquímica com diferentes métodos de fixação tecidual, comparamos a eficácia dos diferentes fixadores, formol a 10% e álcool a 70% para a evidenciação de miofibroblastos em tecido de granulação de úlceras crônicas de 21 pacientes. Método: Estudamos 42 amostras por meio de técnica imuno-histoquímica, utilizando-se o anticorpo anti-Smooth Muscle Actin, nas quais contamos os miofibrobastos em 10 campos com aumento de 400 X. Resultados: Demonstraram-se melhores resultados qualitativo e quantitativo no material fixado em álcool, quando comparado ao fixado em formol, com densidades médias de miofibroblastos de 86,5 e 48 células/mm2, respectivamente, o que nos leva a recomendar o álcool como fixador de tecidos para estas análises imuno-histoquímicas.
Background: The use of a good visualization technique is crucial for an adequate pathological anatomy study of tissues obtained from patients. Optimal staining and fixation techniques of the surgical piece are essential. In a previous study investigating myofibroblasts and contraction in split skingraft donor sites, a low density of myofibroblasts was observed, as opposed to studies in granulation studies. To investigate the causes for this finding and based on a specific manuscript on the efficacy of immunohistochemical visualization using different tissue fixation methods, we compared the efficacy of fixation in 10% formaldehyde and 70% alcohol for the visualization of myofibroblasts in granulation tissues of chronic ulcers in 21 patients. Method: Forty-two samples were studied by immunohistochemistry, using the anti-Smooth Muscle Actin antibody, and myofibroblasts were counted in 10 fields with a magnification of 400X. Results: Better qualitative and quantitative results were observed in alcohol-fixed materials, when compared to those fixed in formaldehyde, with mean myofibroblasts densities of 86.5 and 48 cells/mm2, respectively, which encourages us to recommend the use of alcohol for tissue fixation in immunohistochemical studies.