• Title/Summary/Keyword: Negative Stain

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Immunocytochemical Characteristics of the Short-term Cultured Mesothelial Cells (단기배양한 중피세포의 면역세포화학적 연구)

  • Jeon, Ho-Jong;Lee, Mi-Ja;Lee, Mi-Sook;Jeong, Yu-Kyung;Lee, Young-Mi;Choi, Hyung-Ho
    • The Korean Journal of Cytopathology
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    • v.6 no.2
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    • pp.106-115
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    • 1995
  • Reactive humsn mesothelial cells were examined by immunocytochemical stain with intermediate filaments (cytokeratin [CK1, CK7, CK8, CK18, CD19), vimentin, desmin, actin), epithelial membrane antigen, carcinoembryonic antigen (CEA), MHC class II antigen (HLA-DR), LeuM-1 (CD15), $\alpha1-antitrypsin$(ACT), $\alpha1-antichymotrypsin$ (ACHT), CD68(KP-1) and FcyRIII(CD16). The mesothelial cells were isolated from patients with liver cirrhosis and pleural effusion, and short-term cultured in RPMI 1640 media containing 10% heat inactivated fetal calf serum and 1% identical supernatant fluid of the patients' transudates. The results obtained are as follows 1. The cultured-reactive mesothelial cells were positive for the protein of cytoskeleton such as cytokeratin and vimentin, but negative for desmin and actin. The resting mesothelial cells showed positive reactions for cylokeratin, but negative for vimentin, desmin and actin. 2. The primary antibodies to the cytokeratin were strongly reactive for CK1, CK8 and CK18 but negative for CK7 and CK19 in both reactive and resting mesothelial cells. 3. Resting mesothelial cells showed negative reactions for CEA, but strong positive reactions in cultured-reactive mesothelial cells. 4. The markers for the monocytes/histiocytes(CD11b, CD14, CD16, CD68, Iysozyme and $\alpha1-antitrypsin$ and $\alpha1-antichymotrypsin$) were nonreactive in resting mesothelial cells, but lysozyme and $\alpha1-antitrypsin$ were weakly reactive in reactive and proliferative mesothelial cells. 5. MHC Class II molecule(HLA-DR antigen) was negative in both resting and reactive mesothelial cells. These results suggest that the short-term cultured, reactive mesothelial cells show a newly aberrant expression of the vimentin and calcine-embryonic antigen. The reason of the aberrant expression of the intermediate filament and oncofetal antigen in reactive and proliferative mesothelial cells should be further evaluated.

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Modified Toluidine Blue: an Alternative Stain for Helicobacter pylori Detection in Routine Diagnostic Use and Post-eradication Confirmation for Gastric Cancer Prevention

  • Sakonlaya, Dussadee;Apisarnthanarak, Anucha;Yamada, Nobutaka;Tomtitchong, Prakitpunthu
    • Asian Pacific Journal of Cancer Prevention
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    • v.15 no.16
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    • pp.6983-6987
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    • 2014
  • Background: Modified toluidine blue staining (MTBs) is a simple, inexpensive and time saving method to detect H. pylori in gastric biopsy specimens. As a metachromatic stain, it simultaneously highlights intestinal metaplasia, a gastric cancer precancerous lesion. The aim of this study was to assess the reliability of MTBs compared with hematoxylin-eosin (H&E) for H. pylori detection using immunoperoxidase staining as the gold standard. This technique would be beneficial for a routine diagnosis and confirmation of H. pylori eradication in developing countries where endoscopic-based approaches are dominant. Materials and Methods: Esophagogastroduodenoscopy with triple site gastric biopsies was undertaken in 207 dyspeptic patients at Thammasat University Hospital, Thailand between 1997 and 1999. H&E, MTBs and immunoperoxidase staining were applied to each specimen. The presence or absence of H. pylori with each stain was interpreted separately and the sensitivity, specificity, positive and negative predictive values of H&E and MTBs were calculated. Results: A total of 282 specimens from 207 patients were evaluated. Using immunoperoxidase staining, organisms were positive in 117 specimens (41%). MTBs proved almost equally sensitive as immunoperoxidase (99%) and significantly more sensitive than H&E (85%). It has comparable specificity (96% vs 96%), PPV (95% vs 94%), and NPV (99% vs 90%) to H&E, using immunoperoxidase staining as gold standard. MTBs compared with immunoperoxidase staining, is cheaper (2 USD vs 12 USD) and faster (20 min vs 16 hrs) compared to immunoperoxidase staining. Conclusions: MTBs is effective, economical and easy to use in daily practice for the detection of H. pylori in gastric biopsy specimens. In addition to saving time in evaluating H. pylori associated gastritis, with a high sensitivity and ability to demonstrate intestinal metaplasia, the technique may have a role in confirmation of H. pylori eradication for gastric cancer prevention in a developing country setting.

A report of 38 unrecorded bacterial species in Korea within the classes Bacilli and Deinococci isolated from various sources

  • Kang, Heeyoung;Kim, Haneul;Bae, Jin-Woo;Lee, Soon Dong;Kim, Wonyong;Kim, Myung Kyum;Cha, Chang-Jun;Yi, Hana;Im, Wan-Taek;Kim, Seung Bum;Seong, Chi Nam;Joh, Kiseong
    • Journal of Species Research
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    • v.8 no.2
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    • pp.176-190
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    • 2019
  • A total of 38 bacterial strains within the classes Bacilli and Deinococci were isolated from various sources in Korea. Samples were collected from animal intestine, urine, soil, tidal flat mud, and kimchi. In the sequence comparison and phylogenetic analysis of 16S rRNA sequences, the 38 isolates were assigned to the classes Bacilli and Deinococci with sequence similarities more than 98.7%. Twenty-four strains and 13 strains were classified the order Bacillales and Lactobacillales in the class Bacilli, respectively. In the order Bacillales, there were nine species in the genus Bacillus, seven species in the genus Paenibacillus, and the remaining eight species in the genera Domibacillus, Halobacillus, Virgibacillus, Lysinibacillus, Paenisporosarcina, Planococcus, Savagea, and Staphylococcus. In the order Lactobacillales, there were four species in the genus Lactobacillus, three species in the genus Leuconostoc, three species in the genus Lactococcus, and the remaining three species in the genera Aerococcus, Enterococcus, and Streptococcus. One species was related to the genus Deinococcus of the order Deinococcales. Most of the isolated strains were Gram-stain-positive, but some were Gram-stain-variable or Gram-stain-negative. Cells were rod or cocci-shaped. Based on the results of 16S rRNA analysis, we report 38 strains as previously unrecorded species to Korea, and the basic characteristics of strains are described herein.

Comparison of Pre-Stain Suspension Liquids in the Contrasting Ability of Neutralized Potassium Phosphotungstate for Negative Staining of Bacteria

  • Kim, Ki-Wooh;Jung, Woo-Kyung;Park, Yong-Ho
    • Journal of Microbiology and Biotechnology
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    • v.18 no.11
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    • pp.1762-1767
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    • 2008
  • Image contrast of whole bacteria was compared in Staphylococcus aureus and Escherichia coli depending on pre-stain suspension liquids by energy-filtering transmission electron microscopy. The two bacterial strains were suspended in three most commonly used liquids for negative staining (triple distilled water [DW], phosphate-buffered saline [PBS], and nutrient broth [NB]) and directly observed without staining or stained with neutralized potassium phosphotungstate (PTA), respectively. Even though in low contrast, unstained bacteria were observed owing to their inherent electron density and cell shape in zero-loss (elastic scattering) images. After being suspended in PBS, unstained bacteria appeared to have higher contrast and more refined periphery than DW-suspended ones, and extracellular appendage structures such as fimbriae and flagella could be discerned. The unstained bacteria appeared to be invariably surrounded with electron-lucent precipitates, possibly from PBS. As far as delineation of the structures, the combination of DW or PBS suspension with subsequent staining provided the most satisfactory results, as evidenced by the high contrast of bacterial morphology and appendage structures. However, after being suspended in NB and stained with PTA, bacteria often had too high contrast or poor staining, with electron-dense aggregates around the bacteria. These results suggest that suspension with concentrated organic aliquots including broth media before PTA staining could deteriorate image contrast, and should be used only in dilute form for visualizing bacterial morphology and appendage structures. Moreover the contrast enhancement of unstained bacteria by salt granules would be advantageous in demonstrating bacterial sorption of environmental particles like heavy metals, maintaining minimal contrast for cell imaging.

Wild Raccoons (Procyon lotor) with Ancylostomiasis in Kora (한국 야생 너구리에서 발생한 구충증)

  • 조경오;박남용;강성귀;김용환;배성열;김영대;김계엽;이근우
    • Journal of Veterinary Clinics
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    • v.18 no.2
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    • pp.178-181
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    • 2001
  • Four dead wild racoons found in Chonnam province were examined parasitologically, pathologically and bacteriologically. Using standard flotation technique with feces many number of ancylostoma eggs which were oval in shape and $60-70{\times}35-45 {\mu}m$ in size were detected. Grossly, severe anemia, multiple congestion and consolidation of lung and catarrhal exudate in small intestine were observed. One of racoons had multifocal white to yellow nodules in the liver. Histopathologically, ancylostoma larvas were found in intestinal lumen. Some of them penetrated and attached to intestinal mucosa. Lungs revealed multiple severe fibrinopurulent pneumonia in which bacterial colonies were scattered. Liver had multiple microabscesses containing peripheral bacterial colony. By Gram stain using paraffin sections of liver and lung, bacterial colony was Gram negative. Isolated bacteria from lung and liver was Gram negative rod. This bacteria was identical with Escherichia coli by biochemical tests. From these results, ancylostomiasis caused severe problems in Korean wild racoons. This is the first report about ancylostomiasis in wild racoons in Korea.

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Flavonoids as a Possible Preventive of Dental Plaque

  • Ammar-Nagwa;El, Diwany-Ahmed;Nagwa-Osman;Soheir-Gaafar;Nagwa-Amin
    • Archives of Pharmacal Research
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    • v.13 no.2
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    • pp.211-213
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    • 1990
  • To test flavonoids for antibacterial activity against oral micraorganisms, flavonoids, quercetrin and naringenin, were incorporated into two pharmaceutical preparations in the form of tooth paste. Samplees of dental plaque, the msot accused dental deposit which initiates the gingival and periodental diseases, were collected from the teeth surface of ten dental students at one week interval before and after using placebo, followed by two formulae of tooth paste containing 0.1% of quercetrin and naringenin (formulas I and II, respectively). The amount of dental plaque was assessed by the quigley and Hens index. Then plaque samples were subjected to bacteriological examination of Gram stain and plate counts of microorganisms. The amount of dental plaque was assessed by the Quigley and Hens index. Then plaque samples were subjected to bacteriological examination of Gram stain and plate counts of microorganisms. The results revealed that most of Gram negative cocci and bacilli were highly affected by the two formulae : the number of actinomycetes were decreased after using formula I and disappeared completely by the sue of formula II, while the number of Gram positive streptococci was highly decreased after the treatment with the two formulae. These results indicate a possible use of flavonoids to inhibit dental plaque formation.

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Mycobacterium abscessus Skin Infection Associated with Shaving Activity in a 75-year-old Man

  • Choi, Hoon;Kim, Yong Il;Na, Chan Ho;Kim, Min Sung;Shin, Bong Seok
    • Annals of Geriatric Medicine and Research
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    • v.22 no.4
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    • pp.204-207
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    • 2018
  • Mycobacterium abscessus comprises rapidly growing mycobacteria, and the clinical manifestations of M. abscessus skin infection include papule, nodule, ulcer, scar and mixed form. The cutaneous infections have been reported due to minor trauma, cosmetic therapy, acupuncture and disseminated infection. A 75-year-old man presented with pruritic diffuse various sized erythematous papuloplaques and pustules on the neck and chest for 2 months. The cutaneous lesions were spread around the wound of the shaving on the neck. The histopathologic findings were consistent with abscess showing infiltrations of neutrophils and lymphocytes in the dermis and negative findings were observed on immunohistochemical stain including acid-fast bacilli stain. One month later, mycobacterial culture result showed positive findings, and the pathogen was identified by reversetranscriptase polymerase chain reaction with hybridization. The patient was treated with combination of clarithromycin and ethambutol for 5 months and there is no evidence of recurrence after 6 months of follow-up. Herein, we report a case of M. abscessus cutaneous infection through minor trauma caused by shaving in the elderly.

A Comparative Analysis on The Efficiency of Various Clinical Methods for Diagnosis of Tuberculosis (결핵 진단을 위한 검사 방법간의 효율성에 관한 비교 분석)

  • 최석철;정천환;성희경;김태운;이원재
    • Biomedical Science Letters
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    • v.5 no.2
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    • pp.191-200
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    • 1999
  • In recent years continuously increasing number of tuberculosis (TB) cases due to the emergence of strains with multidrug resistance and AIDS is a significant global health problem. Therefore, more rapid and reliable diagnosis of TB may be one of the most urgent needs in efforts to eradicate the disease. The present study was designed to compare and assess the diagnostic values and efficiencies between the conventional methods (X-ray, AFB stain and culture) and PCR for pulmonary TB on 171 cases. Chest X-ray finding and clinical features revealed that 39 (22.8%) of 171 sputum specimens were pulmonary TB cases. The statistical data were taken on the basis of the definitive diagnosis: In X-ray, overall sensitivity, specificity, efficiency and false positive and false negative incidence was respectively 69.2%, 87.1%, 83.0%, 12.9%, and 30.8%; 79.5%, 95.5%, 91.8%,4.6% and 20.5% in AFB-stain; 56.4%, 99.2%,89.5%, 0.8% and 43.6% in culture; 82.1%, 96.2%, 93.0%, 3.8% and 17.9% in PCR. PCR got a highest sensitivity and efficiency as well as a lowest false negative incidence. Culture had a highest specificity with a lowest false positive incidence. These results imply that PCR assay is fast, sensitive and efficient method for diagnosis of pulmonary TB. However, combined use of the conventional methods with thorough quality control may offer more opportunities for detecting Mycobacterium tuberculosis and diagnosting TB although they have some limits.

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A Study on the Somatic Cells in Half Milk Samples of Dairy Goats (젖염소 분방유즙에서 체세포에 대한 연구)

  • Kim, Hye-Ra;Lee, Jeong-Chi;Jeong, Ji-Young;Lee, Youn-Kyung;Shin, Sung-Shik;Lee, Chai-Yong
    • Journal of Veterinary Clinics
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    • v.24 no.2
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    • pp.177-181
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    • 2007
  • In the present study, a total of 547 half milk samples were collected from 274 dairy goats to perform somatic cell counts (SCC) and California mastitis test (CMT). Milk smear was stained with Pyronin Y-Methyl Green stain were classified into either epithelial or blood cells, etc. Of the 547 halves the percentage of CMT negative milk samples were 86%. Among these, 58.2% were CMT negative with SCC<500,000/ml, while 27.8% were CMT negative with SCC>500,000 ml. As expected, CMT score increased with the increase of SCC. The number of epithelial cells decreased with the increasing number of somatic cells, while the opposite was observed with the number of blood cells. These results indicate that the critical point in milk quality & CMT should be considered on the false (pseudo-SC) SCC in dairy goat.

Dynamic lipopolysaccharide transfer cascade to TLR4/MD2 complex via LBP and CD14

  • Kim, Soo Jin;Kim, Ho Min
    • BMB Reports
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    • v.50 no.2
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    • pp.55-57
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    • 2017
  • Toll-like receptor 4 (TLR4) together with MD2, one of the key pattern recognition receptors for a pathogen-associated molecular pattern, activates innate immunity by recognizing lipopolysaccharide (LPS) of Gram-negative bacteria. Although LBP and CD14 catalyze LPS transfer to the TLR4/MD2 complex, the detail mechanisms underlying this dynamic LPS transfer remain elusive. Using negative-stain electron microscopy, we visualized the dynamic intermediate complexes during LPS transfer-LBP/LPS micelles and ternary CD14/LBP/LPS micelle complexes. We also reconstituted the entire cascade of LPS transfer to TLR4/MD2 in a total internal reflection fluorescence (TIRF) microscope for a single molecule fluorescence analysis. These analyses reveal longitudinal LBP binding to the surface of LPS micelles and multi-round binding/unbinding of CD14 to single LBP/LPS micelles via key charged residues on LBP and CD14. Finally, we reveal that a single LPS molecule bound to CD14 is transferred to TLR4/MD2 in a TLR4-dependent manner. These discoveries, which clarify the molecular mechanism of dynamic LPS transfer to TLR4/MD2 via LBP and CD14, provide novel insights into the initiation of innate immune responses.