• Title/Summary/Keyword: Negative Stain

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Detection and Identification of Mycobacterium Tuberculosis in Patients with Tuberculous Cervical Lymphadenitis by PCR-RFLP (경부 결핵성 임파선염 환자에서 PCR-RELP를 이용한 결핵균의 검출 및 확인)

  • Lee Sang-Sook;Cho Young-Rok;Chun Ji-Min;Choi Yong-Seok;Sohn Eun-Ju;Park Nam-Cho;Park June-Sik
    • Korean Journal of Head & Neck Oncology
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    • v.12 no.2
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    • pp.169-176
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    • 1996
  • Tuberculous cervical lymphadenitis is still an important cause of neck mass in Korea. Tuberculosis is an important differential diagnosis in patients of cervical lymphadenopathy. Rapid and sensitive test for the diagnosis of tuberculosis is essential for the approapiate treatment. Up to now, conventional diagnostic methods for M. tuberculosis were acid-fast bacilli(AFB) stain and culture of M. tuberculosis. The direct microscopic examination of AFB by Ziehl-Neelsen stain is rapid, but often negative. The culture for M. tuberculosis is time-consuming, taking 4 to 8 weeks. Recently various methods to detect Mycobacterial DNA, including PCR and restriction fragment length polymorphism(RFLP) analysis have been reported. Here we represent a simple method for the confirmation of M. tuberculosis and exclusion of the other Mycobacterial species by RFLP analysis and silver staining of polyacrylamide gel electrophoresis after nested PCR for a repetitive DNA sequence(IS986) specific for M. tuberculosis from fresh or paraffin-embedded biopsy specimens. This result leads us to conclude that this method is simple, rapid and possibly applicable to confirm M. tuberculosis and rule out the other Mycobacteria species from the clinical specimens in the clinical laboratories.

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A Case of Abdominal Tuberclosis Mimicking Ovarian Cancer (난소 종양의 임상적 양상을 보인 소아 복막 결핵)

  • Yeom, Jung Sook;Kim, Jae Hui;Cho, Jae Min;Lee, Jeong Hee;Park, Eun Sil;Seo, Ji Hyun;Lim, Jae Young;Park, Chan Hoo;Woo, Hyang Ok;Youn, Hee Shang
    • Pediatric Gastroenterology, Hepatology & Nutrition
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    • v.8 no.2
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    • pp.238-242
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    • 2005
  • We report the case of a girl with peritoneal tuberculosis (Tb) who presented with mimicking features of ovarian cancer including massive ascites, elevation of CA-125 and ovarian masses. These ovarian masses cannot be ruled out malingnacy with image study. Gram stain, acid-fast stain, bacteriological and mycological culture of centrifuged ascites cells were negative. Tumor markers revealed an elevated CA-125 level of 112 U/mL and adenosine deaminase level were significantly high with 118 U/L. Diagnostic laparoscopy was done. The histology of the peritoneum and the ovary showed multiple granulomatous nodules with epithelioid cells and multi-nucleated giant cells. And there were no evidence of malignancy. Furthermore acid-fast bacilli (AFB) was identified in this specimen. Peritoneal Tb was difficult to diagnosis in our patient because it mimicked ovarian cancer. And this is the first case of abdominal Tb mimicking ovarian cancer in children in Korea.

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Induction of Effective Osteogenesis by Mesenchymal Stem Cells from the Human Subchondral Bone (사람 연골하골 중간엽 줄기세포의 효율적인 골형성 유도)

  • Huh, Jeong-Eun;Cho, Yoon-Je;Yoo, Myung-Chul;Baek, Yong-Hyeon;Lee, Jae-Dong;Choi, Do-Young;Park, Dong-Suk
    • Journal of Acupuncture Research
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    • v.23 no.5
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    • pp.69-77
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    • 2006
  • Background : Mesenchymal stem cells (MSCs) are present in most of the tissue matrix, taking part in their regeneration when injury or damage occurs. The aim of this study was to investigate the presence of cells with pluripotential characteristics in human subchondral bone and the capacity of these cells to differentiate to osteoblast. Methods : Human subchondral bone were digested with collagenase. Isolated cells were cultured with a-MEM, 15% FBS, 10-8M dexamethasone and 50 ng/mL ascoric acid. Cells from 0 day(isolated cells), 7 day (first subculture) and 14 days (third subculture) were used to carry out phenotypic characterization experiments flowcytometry analysis with 11 monoclonal antibodies) and osteogenic differentiation experiments. Osteogenic differentiation of cells was assessment by quantification of bone extracellular matrix components by following analysis: alkaline phosphatase(ALP) stains to detect ALP activity, RT-PCR and western blot to detect osteocalcin (OCN), osteopontin (OPN) and type I collagen(Col I), and Alizarin red stains to detect calcium deposition. Results : Flowcytometry analyses showed that in our population more than 98% of cells were positive for MSC markers: SH-2(CD105, 99%), CD29 (95%), CD73 (95%). Cells were negative for hematopoietic markers (CD11b, CD34, and CD45). Furthermore, cells showed positive stain to multipotent markers such as CDl17 (c-kit) (15.1%), and CD166 (74.9%), and cell adhesion molecules such as CD54 (78.1%) and CD106 (63.5%). The osteogenic specific marker analyses showed that the culture of these cells for 7 and 14 days stimulates ALP, OCN, OPN and Col I synthesis by RT-PCR and Western blot analysis. Also, after 14 days in the culture of MSCs induces mineralization by Arizarin red stain. Conclusion : In this work, we demonstrated a new and efficient method for osteoblastic differentiation of human subchondral bone stem cells. As MSCs takes part in reparative processes of adult tissues, these cells could play an important role in osteogenesis.

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Usefulness of Automated PCR Test for the Detection of Mycobacterium Tuberculosis in Fresh Biopsy Tissues (신선조직 검체에서 결핵균 검출을 위한 자동화 중합효소연쇄반응 검사의 유용성)

  • Choi, Woo Soon;Shin, So Young;Kim, Jong Ok;Kim, Myung Sook;Lee, Hye Kyung
    • Tuberculosis and Respiratory Diseases
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    • v.61 no.1
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    • pp.54-59
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    • 2006
  • Background: Although there have been several studies regarding the clinical value of an automated TB-PCR study using sputum, bronchial washing, and other body fluid samples for the detection of pulmonary tuberculosis, there are only a few reports on the use of fresh tissue samples. Materials and methods: The acid-fast bacilli stain(AFB), tuberculosis culture, automated TB-PCR study, and histopathology examination were performed in 42 fresh tissue samples. Results: Among the 42 cases, 18 cases were diagnosed with tuberculosis based on the clinical findings. Sixteen of the 18 cases were TB-PCR positive and of these 16 cases, only 2 cases were positive in the AFB stain or culture study. However, all 18 cases showed the histopathology findings of chronic granulomatous inflammation that was compatible with tuberculosis. Based on the clinical findings, the sensitivity, specificity, positive predictability, and negative predictability of the automated TB-PCR study were 88.9%, 100%, 100%, and 92.3% respectively. Conclusion: An automated TB-PCR assay is an important diagnostic tool for diagnosing tuberculosis in fresh tissue samples.

Can the C-14 Urea Breath Test Reflect the Extent and Degree of Ongoing Helicobacter pylori Infection? (C-14 요소호기검사의 정량치가 Helicobacter pylori 감염 정도를 반영할 수 있을까?)

  • Lim, Seok-Tae;Sohn, Myung-Hee;Lee, Seung-Ok;Lee, Soo-Teik;Jeong, Myoung-Ja
    • The Korean Journal of Nuclear Medicine
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    • v.35 no.1
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    • pp.61-68
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    • 2001
  • Purpose: The C-14 urea breath test (C-14 UBT) is the most specific noninvasive method to detect Helicobacter (H) pylori infection. We investigated if the C-14 UBT can reflect the presence and degree of H. pylori detected by gastroduodenoscopic biopsies (GBx). Materials and methods: One hundred fifty patients (M:F=83:67, age $48.6{\pm}11.2$ yrs) underwent C-14 UBT, rapid urease test (CLO test) and GBx on the same day. For the C-14 UBT, a single breath sample was collected at 10 minutes after ingestion of C-14 urea (137 KBq) capsule and counting was done in a liquid scintillation counter for 1 minute, and the results were classified as positive (${\geq}200dpm$), Intermediate ($50{\sim}199dpm$) or negative (<50 dpm). The results of CLO tests were classified as positive or negative according to color change. The results of GBx on giemsa stain were graded 0 (normal) to 4 (diffuse) according to the distribution of H. pylori by the Wyatt method. We compared C-14 UBT results with GBx grade as a gold standard. Results: In the assessment of the presence of H. pylori infection, the C-14 UBT global performance yielded sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV) and accuracy of 92.5%, 88.4%, 97.1%, 88.4% and 91.3%, respectively. However, the CLO test had sensitivity, specificity, PPV, NPV and accuracy of 83.2%, 81.4%, 91.8%, 81.4% and 82.7%, respectively. The quantitative values of the C-14 UBT were $45{\pm}27$ dpm in grade 0, $707{\pm}584dpm$ in grade 1, $1558{\pm}584dpm$ in grade 2, $1851{\pm}604dpm$ in grade 3, and $2719{\pm}892dpm$ in grade 4. A significant correlation (r=0.848, p<0.01) was found between C-14 UBT and the grade of distribution of H. pylori infection on GBx with giemsa stain. Conclusion: We conclude that the C-14 UBT is a highly accurate, simple and noninvasive method for the diagnosis of ongoing H. pylori infection and reflects the degree of bacterial distribution.

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Comparison of the Quantitative Values of C-14 and C-13 UBT to Reflect the Presence and Degree of Ongoing Helicobacter pylori Infection (Helicobacter pylori 감염 유무와 정도 반영에 대한 C-14와 C-13 요소호기검사 정량치 비교)

  • Lim, Seok-Tae;Kim, Dong-Wook;Jeong, Hwan-Jeong;Sohn, Myung-Hee
    • Nuclear Medicine and Molecular Imaging
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    • v.42 no.3
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    • pp.229-234
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    • 2008
  • Purpose: A urea breath test (UBT) using C-14 or C-13 has been developed for identifying Helicobacter (H) pylori infection on the basis of urease production with release of labeled $CO_2$. We investigated if the C-14 and C-13 UBT have the difference to reflect the presence and degree of H. pylori infection detected by gastro-duodenoscopic biopsies (CBx) in the same patients. Materials and methods: Thirty eight patients (M:F = 28:10, age $53.4{\pm}13.0$ yrs) with upper gastrointestinal symptoms such as indigestion, gastric fullness or pain consecutively underwent C-14 UBT, GBx and C-13 UBT within one week before medications. For the C-14 UBT, a single breath sample was collected at 10 minutes after ingestion of C-14 urea (37 KBq) capsule and counting was done in a liquid scintillation counter for 1 minute, and the results were classified as positive (${\ge}$ 200 dpm), intermediate (50-199 dpm) or negative (50 dpm). For the C-13 UBT, the results were classified as positive (${\ge}2.5\%_{\circ}$) or negative ($<2.5\%_{\circ}$). The results of GBx with Giemsa stain were graded 0 (normal) to 4 (diffuse) according to the distribution of H. pylori by the Wyatt method. We compared C-14 UBT and C-13 UBT results with GBx grade as a gold standard. Results: The prevalence of H. pylori infection by GBx with Giemsa stain was 25/38 (65.8%). In the assessment of the presence of H. pylori infection, the C-14 UBT global performance yielded sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV) and accuracy of 92.0%, 92.3%, 95.8%, 91.7% and 92.1%, respectively. However, the C-13 UBT had sensitivity, specificity, PPV, NPV and accuracy of 96.0%, 84.6%, 92.3%, 91.7% and 92.1%, respectively. The more significant correlation in C-14 than C-13 UBT (r=0.948 vs r=0.819, p <0.001) was found between the value of UBT and the grade of distribution of H. pylori infection. Conclusion: We conclude that the diagnostic performance between C-14 and C-13 UBT to detect H. pylori infection is not significantly different, but the value of C-14 UBT more significantly reflects the degree of bacterial distribution.

How Reliable is Sputum PCR Test in the Diagnosis of Pulmonary Tuberculosis When Sputum Smear is Negative? (객담 결핵균 도말검사가 음성일때 중합효소연쇄반응검사와 진단적 신뢰도에 관한 연구)

  • Baek, Seung-Hoon;Lee, Jae-Myung;Kang, Min-Jong;Son, Jee-Woong;Lee, Seung-Joon;Kim, Dong-Gyu;Lee, Myung-Goo;Hyun, In-Gyu;Jung, Ki-Suck;Lee, Kyung-Wha;Joe, Hyun-Chan
    • Tuberculosis and Respiratory Diseases
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    • v.50 no.2
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    • pp.222-228
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    • 2001
  • Backgrounds : Recent technological developments have introduced a new method to identifying M. tuberculosis complex DNA in clinical samples directly. The direct amplification test (DAT) is approved for identifying M. tuberculosis complex in respiratory specimens that are smear-positive for acid-fast bacilli (AFB). When there is a discrepancy between the AFB smear and DAT, no information on their clinical utility is currently available. In this study, the diagnostic reliability of DAT was investigated in suspected pulmonary tuberculosis patients whose sputum AFB smear was negative. Methods : From June 1, 1998 through May 30, 1999, 909 patients with presumed active pulmonary tuberculosis were enrolled. A sputum AFB stain, culture, DAT and/or biopsy were performed. Using the criteria of clinical tuberculosis or confirmed tuberculosis, the positive predictive value of DAT in diagnosing pulmonary tuberculosis was investigated. Results : The positive predictive value of DAT was 82.1% by the clinically active tuberculosis criteria. However, it decreased to 61.5% when diagnosis was restricted to only to culture positive or biopsy proven cases. The false positive rate of DAT was 18.0%. Conclusion : The DAT is a valuable diagnostic method in suspected patients whose sputum AFB is was negative.

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Evaluation for detection of Cryptosporidium oocysts in diarrheal feces of calves (야외 송아지 설사변에서 작은와포자충 검출에 대한 평가)

  • Wi, Seong-Hwan;Ju, Hu-Don;Gang, Yeong-Bae
    • Parasites, Hosts and Diseases
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    • v.34 no.2
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    • pp.121-126
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    • 1996
  • For the detection of Cwptospori,mum oocysts, fecal samples were collected from 201 calves which showed diarrhea. Among the 201 samples, 29 samples (14.4%) were positive for Cwptosporinium spry. by the DMSO-modified acid-fast stain (MAFS) , 23 samples (11.4%) were positive by commercial kit (Meridian Diagnostics, Cincinnati, Ohiol and 23 by the indirect immunofluorescence antibody (IFA )assay employing the monoclonal antibody (mAb C6). When tested by both IFA and MAFS, 20 fecal samples were positive for Cwptosporinium oocysts whereas 169 fecal samples were negative. If the MAFS is considered a standard method for oocyst detection, the IFA showed 69% of sensitivity and 98% of specificity. When tested by both IFA and commercial kit, 22 fecal samples were positive for Cwptospori,mum oocysts while 177 samples were negative. One sample tested by IFA was found to be false negative, when compared with the results by commercial kit. The sensitivity of IFA was calculated as high as 96%; the specificity as 99% and the predictive value was also 99%. In the present study, IFA employing the nAb C6 revealed that 23 samples (11.4%) were positive among the 201 calves showing diarrhea. Of 23 IFA positive samples, 4 samples (5%) showed cryptosporidial oocysts more than 105 OPG Therefore. it is concluded that the calves showing cryptosporidial oocysts more than 105 OPG in the feces were highly associated with clinical cryptosporidiosis.

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The effect of polyphosphate on exophytic bone formation (무기인산염이 외방성 수직골 형성에 미치는 영향)

  • Lee, Jean;Park, Joon-Bong;Herr, Yeek;Chung, Jong-Hyuk;Kwon, Young-Hyuk
    • Journal of Periodontal and Implant Science
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    • v.38 no.1
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    • pp.59-66
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    • 2008
  • Purpose: It has been shown that the inorganic polyphosphate is effective for the regeneration of bones through the preliminary animal test of rabbits. The most effective concentration of the polyphosphate, however, is not known yet. Moreover, the effectiveness of carriers inside human body is not confirmed.. Materials and Methods: In this study, we examined the effect of the concentration of the inorganic polyphosphate on the process of the bone regeneration using the 6 weeks old rabbits with the weight of 2.0 kg in average. We performed the experiment using TR-ePTFE membrane(membrane) filled with collagen immersed in 4%, 8% of inorganic polyphosphate, respectively, following removal of the proper sized cortical bones from the rabbit calvaria. The experimental results were compared with the one of the following four groups: The negative control group for membrane only, the positive control group for membrane filled with collagen, the first experimental group for membrane filled with collagen immersed in 4% of inorganic polyphosphate, and the second experimental group for membrane filled with collagen immerse in 8% of inorganic polyphosphate. The fragments of the tissue with membrane obtained from each group of the sacrificed rabbits for 8 or 16 weeks sustained after surgery were then prestained by the Hematoxylin-Eosin stain and coated by resin to form non-decalcified specimens for the histologic examination and analysis. New bone formation was assessed by histomorphometric and statistical analysis. Results: 1. All groups have shown better bone regeneration at 16weeks than 8weeks. 2. Negative control group has shown more bone regeneration relative to the other groups at 8 and 16 weeks. 3. All experimental groups have shown better bone regeneration relative to positive control group. 4. At 16 weeks, the first experimental group has shown more bone regeneration compared to the second experimental group. Exophytic bone formation is not good at the first and the second experimental groups compared with negative control group. But, the use of 4% inorganic polyphosphate was more effective to bone formation than the use of 8% inorganic polyphosphate. Conclusion: With above results, it is suggested the use of inorganic polyphosphate with vehicle under TR-ePTFE membrane.

The Protective Effects of Statins towards Vessel Wall Injury Caused by a Stent Retrieving Mechanical Thrombectomy Device : A Histological Analysis of the Rabbit Carotid Artery Model

  • Lee, Seung Hwan;Shin, Hee Sup;Oh, Inho
    • Journal of Korean Neurosurgical Society
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    • v.64 no.5
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    • pp.693-704
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    • 2021
  • Objective : Endovascular mechanical thrombectomy (MT) has been regarded as one of the standard treatments for acute ischemic stroke caused by large vessel occlusion. Despite the wide use of stent retrievers for MT, arterial intimal damage caused when deployed stent is pulled has been a certain disadvantage. We hypothesized that statin could protect and stabilize vessel damage after endovascular MT using a stent retriever. In this animal study, we observed the protective effects of the statins towards MT-induced vessel wall injury. Methods : Twenty-eight carotid arteries of fourteen rabbits were used in the experiments with MT using stent retriever. We divided the rabbits into four groups as follows : group 1, negative control; group 2, positive control; group 3, statin before MT; and group 4, statin after MT. After MT procedures, we harvested the carotid arteries and performed histomorphological and immunohistochemical analyses. Results : In histomorphological analysis with hematoxylin and eosin and Masson's trichrome stain, significant intimal thickening (p<0.05) was observed in the positive control (group 2), compared to in the negative control (group 1). Intimal thickening was improved in the statin-administered groups (groups 3 and 4 vs. group 2, p<0.05). We also observed that statin administration after MT (group 4) resulted in a more effective decrease in intimal thickness than statin administration before MT (group 3) (p<0.05). We performed immunohistochemical analysis with the antibodies for tumor necrosis factor-alpha (TNF-α), cluster of differentiation (CD)11b, and CD163. In contrast to the negative control (group 1), the stained percentage areas of all immunological markers were markedly increased in the positive control (group 2) (p<0.05). Based on statin administration, the percentage area of TNF-α staining was significantly reduced (p<0.05) in group 3, compared to the positive control group (group 2). However, significant differences were not observed for CD11b and CD163 staining. In group 4, no significant differences were observed for TNF-α, CD11b, and CD163 staining (p≥0.05). The differences in the percentage areas of the different markers between the statin-administered groups (groups 3 and 4) were also not revealed. Conclusion : We presented that statin administration before and after MT exerted protective effects towards vessel wall injury. The efficacy of statins was greater post-administration than pre-administration. Thus, statin administration in routine prescriptions in the peri-procedural period is strongly advised.