Displaying publications 1 - 20 of 53 in total

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  1. Wan Mohamad WAF, Buckow R, Augustin M, McNaughton D
    Food Chem, 2017 Oct 15;233:197-203.
    PMID: 28530566 DOI: 10.1016/j.foodchem.2017.04.086
    Confocal Raman microscopy (CRM) was able to quantify the β-carotene concentration in oil droplets and determine the partitioning characteristics of β-carotene within the emulsion system in situ. The results were validated by a conventional method involving solvent extraction of β-carotene separately from the total emulsion as well as the aqueous phase separated by centrifugation, and quantification by absorption spectrophotometry. CRM also enabled the localization of β-carotene in an emulsion. From the Raman image, the β-carotene partitioning between the aqueous and oil phases of palm olein-in-water emulsions stabilized by whey protein isolate (WPI) was observed. Increasing the concentration of β-carotene in an emulsion (from 0.1 to 0.3g/kg emulsion) with a fixed gross composition (10% palm olein:2% WPI) decreased the concentration of β-carotene in the oil droplet. CRM is a powerful tool for in situ analyses of components in heterogeneous systems such as emulsions.
    Matched MeSH terms: Microscopy, Confocal*
  2. Tan YF, Leong CF, Cheong SK
    Malays J Pathol, 2010 Dec;32(2):97-102.
    PMID: 21329180 MyJurnal
    Dendritic cells (DCs) are professional antigen presenting cells of the immune system. They can be generated in vitro from peripheral blood monocytes supplemented with GM-CSF, IL-4 and TNF alpha. During induction, DCs will increase in size and acquire multiple cytoplasmic projections when compared to their precursor cells such as monocytes or haematopoietic stem cells which are usually round or spherical. Morphology of DCs can be visualized by conventional light microscopy after staining or phase-contrast inverted microscopy or confocal laser scanning microscopy. In this report, we described the morphological appearances of DCs captured using the above-mentioned techniques. We found that confocal laser scanning microscopy yielded DCs images with greater details but the operating cost for such a technique is high. On the other hand, the images obtained through light microscopy after appropriate staining or phase contrast microscopy were acceptable for identification purpose. Besides, these equipments are readily available in most laboratories and the cost of operation is affordable. Nevertheless, morphological identification is just one of the methods to characterise DCs. Other methods such as phenotypic expression markers and mixed leukocyte reactions are additional tools used in the characterisation of DCs.
    Matched MeSH terms: Microscopy, Confocal*
  3. Saini R, Azmi AS, Ghani NB, Al-Salihi KA
    Med J Malaysia, 2007 Aug;62(3):238-40.
    PMID: 18246915 MyJurnal
    This study was designed to identify surface and subsurface microscopic changes in different carious lesions by using Confocal Laser Scanning Microscope (CLSM) and Image analyzer (light microscopy). Thirty extracted carious posterior teeth were fixed, embedded and polymerized in plastic fixation medium. The final thin sections (80mm) were stained with H&E and Masson Goldner's Tricome while others were left unstained. Under Confocal, marked differences between control sound enamel and dentin, and carious area of the samples were observed which illustrated that a correlation existed between the zone of autofluoresence, demineralization and carious enamel and dentin. Compared to CLSM, Image Analyzer only produce two-dimensional images but the histopathological changes were better appreciated by using various staining methods.
    Matched MeSH terms: Microscopy, Confocal*
  4. Hassan T, Thiberville L, Hermant C, Lachkar S, Piton N, Guisier F, et al.
    PLoS One, 2017;12(12):e0189846.
    PMID: 29267317 DOI: 10.1371/journal.pone.0189846
    BACKGROUND: Malignant solitary pulmonary nodules (SPN) have become more prevalent, with upper lobes predilection. Probe-based confocal laser endomicroscopy (pCLE) provides in-vivo imaging of SPN. However, the stiffness of the 1mm confocal probe (AlveoFlex) causes difficult accessibility to the upper lobes. A thinner 600μm probe designed for bile duct exploration (CholangioFlex) has the potential to reach the upper lobes.

    OBJECTIVES: To examine the accessibility of malignant SPNs in all segments of the lungs using either the 0.6mm or 1.4 mm probe and to assess the quality and inter observer interpretation of SPN confocal imaging obtained from either miniprobes.

    METHODS: Radial(r)-EBUS was used to locate and sample the SPN. In-vivo pCLE analysis of the SPN was performed using either CholangioFlex (apical and posterior segments of the upper lobes) or AlveoFlex (other segments) introduced into the guide sheath before sampling. pCLE features were compared between the two probes.

    RESULTS: Fourty-eight patients with malignant SPN were included (NCT01931579). The diagnostic accuracy for lung cancer using r-EBUS coupled with pCLE imaging was 79.2%. All the SPNs were successfully explored with either one of the probes (19 and 29 subjects for CholangioFlex and AlveoFlex, respectively). A specific solid pattern in the SPN was found in 30 pCLE explorations. Comparison between the two probes found no differences in the axial fibers thickness, cell size and specific solid pattern in the nodules. Extra-alveolar microvessel size appeared larger using CholangioFlex suggesting less compression effect. The kappa test for interobserver agreement for the identification of solid pattern was 0.74 (p = 0.001).

    CONCLUSION: This study demonstrates that pCLE imaging of SPNs is achievable in all segments of both lungs using either the 0.6mm or 1.4mm miniprobe.

    Matched MeSH terms: Microscopy, Confocal/instrumentation; Microscopy, Confocal/methods*
  5. Kapitonova MY, Kuznetsov SL, Salim N, Othman S, Kamauzaman TM, Ali AM, et al.
    Bull. Exp. Biol. Med., 2014 Jan;156(3):393-8.
    PMID: 24771384 DOI: 10.1007/s10517-014-2357-8
    Morphological and phenotypical signs of cultured readaptation osteoblasts were studied after a short-term space mission. The ultrastructure and phenotype of human osteoblasts after Soyuz TMA-11 space flight (2007) were evaluated by scanning electron microscopy, laser confocal microscopy, and ELISA. The morphofunctional changes in cell cultures persisted after 12 passages. Osteoblasts retained the drastic changes in their shape and size, contour deformation, disorganization of the microtubular network, redistribution of organelles and specialized structures of the plasmalemma in comparison with the ground control cells. On the other hand, the expression of osteoprotegerin and osteocalcin (bone metabolism markers) increased; the expression of bone resorption markers ICAM-1 and IL-6 also increased, while the expression of VCAM-1 decreased. Hence, space flight led to the development of persistent shifts in cultured osteoblasts indicating injuries to the cytoskeleton and the phenotype changes, indicating modulation of bone metabolism biomarkers.
    Matched MeSH terms: Microscopy, Confocal
  6. Fatimah IS, Iswadi IM, Khairul O, Nurhazilah M, Fadzilah MS, Padzil AR, et al.
    Clin Ter, 2010;161(2):125-8.
    PMID: 20499025
    There is an association between reactive oxygen species (ROS) and DNA damage to sperm. Researchers believe that ROS is always present at the sperm's head. The variation of ROS concentration within the area has an impact on the integrity of the DNA.
    Matched MeSH terms: Microscopy, Confocal
  7. Thomas J, Idris NA, Collings DA
    J Microsc, 2017 10;268(1):13-27.
    PMID: 28654160 DOI: 10.1111/jmi.12582
    Pontamine fast scarlet 4B is a red paper and textiles dye that has recently been introduced as a fluorescent probe for plant cell walls. Pontamine exhibits bifluorescence, or fluorescence dependent on the polarization of the excitation light: Because cellulose is aligned within the cell wall, pontamine-labelled cell walls exhibit variable fluorescence as the excitation polarization is modulated. Thus, bifluorescence measurements require polarized excitation that can be directly or indirectly modulated. In our confocal microscopy observations of various cellulose samples labelled with pontamine, we modulated excitation polarization either through sample rotation or by the confocal's scanfield rotation function. This variably rotated laser polarizations on Leica confocal microscopes, but not those from other makers. Beginning with samples with directly observable microfibril orientations, such as purified bacterial cellulose, the velamen of orchid roots and the inner S2 layer of radiata pine compression wood, we demonstrate that modelling the variations in pontamine fluorescence with a sine curve can be used to measure the known microfibril angles. We then measured average local microfibril angles in radiata pine samples, and showed similar microfibril angles in compression and normal (opposite) wood. Significantly, bifluorescence measurements might also be used to understand the degree of local cellulose alignment within the cell wall, as opposed to variations in the overall cellulose angle.
    Matched MeSH terms: Microscopy, Confocal
  8. Gunasegar S, Himratul-Aznita WH
    FEMS Yeast Res., 2019 Mar 01;19(2).
    PMID: 30476044 DOI: 10.1093/femsyr/foy123
    Candida albicans ATCC 14053 and Candida parapsilosis ATCC 22019 hyphal-wall protein 1 (HWP1) are involved in hyphae formation and pathogenesis. The transcriptional agglutinin-like sequence 3 (ALS3) genes in both species are responsible for the development of biofilm and colonization on tooth surfaces. Therefore, we investigated the expression of HWP1 and ALS3 quantitatively in C. albicans and C. parapsilosis and examined the biofilm structure upon exposure to various nicotine concentrations. In vitro, biofilms of Candida species were developed directly on slides using the Lab-Tek Chamber Slide System and visualized by confocal laser scanning microscopy. Quantitative real-time polymerase chain reaction was used to measure HWP1 and ALS3 expression in C. albicans ATCC 14053 and C. parapsilosis ATCC 22019. The results indicated that nicotine multiplied the number of yeast cells and increased the extracellular polysaccharides of Candida species. We also found that 1-2 mg/mL nicotine could enhance the formation of biofilm. The findings also revealed that the expression of HWP1 and ALS3 in Candida species were increased as the nicotine concentration increased. Therefore, nicotine influences the biofilm development of oral-associated C. albicans ATCC 14053 and C. parapsilosis ATCC 22019.
    Matched MeSH terms: Microscopy, Confocal
  9. Chai WL, Moharamzadeh K, Brook IM, Van Noort R
    Biotech Histochem, 2011 Aug;86(4):242-54.
    PMID: 20392135 DOI: 10.3109/10520291003707916
    The success of dental implant treatment depends on the healing of both hard and soft tissues. While osseointegration provides initial success, the biological seal of the peri-implant soft tissue is crucial for maintaining the long term success of implants. Most studies of the biological seal of peri-implant tissues are based on animal or monolayer cell culture models. To understand the mechanisms of soft tissue attachment and the factors affecting the integrity of the soft tissue around the implants, it is essential to obtain good quality histological sections for microscopic examination. The nature of the specimens, however, which consist of both metal implant and soft peri-implant tissues, poses difficulties in preparing the specimens for histomorphometric analysis of the implant-soft tissue interface. We review various methods that have been used for the implant-tissue interface investigation with particular focus on the soft tissue. The different methods are classified and the advantages and limitations of the different techniques are highlighted.
    Matched MeSH terms: Microscopy, Confocal/methods*
  10. Syakila RN, Lim SM, Agatonovic-Kustrin S, Lim FT, Ramasamy K
    Anal Bioanal Chem, 2019 Feb;411(6):1181-1192.
    PMID: 30680424 DOI: 10.1007/s00216-018-1544-2
    The cholesterol-lowering properties of 12 lactic acid bacteria (LAB) in the absence or presence of 0.3% bile salts were assessed and compared quantitatively and qualitatively in vitro. A new, more sensitive and cost-effective high-performance thin-layer chromatography method combined with digital image evaluation of derivatised chromatographic plates was developed and validated to quantify cholesterol in LAB culture media. The performance of the method was compared with that of the o-phthalaldehyde method. For qualitative assessment, assimilated fluorescently tagged cholesterol was visualised by confocal microscopy. All LAB strains exhibited a cholesterol-lowering effect of various degrees (19-59% in the absence and 14-69% in the presence of bile salts). Lactobacillus plantarum LAB12 and Pentosaceus pentosaceus LAB6 were the two best strains of lactobacilli and pediococci. They lowered cholesterol levels by 59% and 54%, respectively, in the absence and by 69% and 58%, respectively, in the presence of bile salts. Confocal microscopy showed that cholesterol was localised at the outermost cell membranes of LAB12 and LAB6. The present findings warrant in-depth in vivo study. Graphical abstract (A) 3D plots based on scan at 525 nm of (B) derivatized HPTLC plate of separated cholesterol and (C) confocal microscopic image showing the localisation of NBD-cholesterol assimilated by LAB.
    Matched MeSH terms: Microscopy, Confocal/methods
  11. Osahor AN, Ng AWR, Narayanan K
    Methods Mol Biol, 2021;2211:29-40.
    PMID: 33336268 DOI: 10.1007/978-1-0716-0943-9_3
    Visual analysis of the gene delivery process when using invasive bacteria as a vector has been conventionally performed using standard light and fluorescence microscopy. These microscopes can provide basic information on the invasiveness of the bacterial vector including the ability of the vector to successfully adhere to the cell membrane. Standard microscopy techniques however fall short when finer details including membrane attachment as well as internalization into the cytoplasm are desired. High-resolution visual analysis of bacteria-mediated gene delivery can allow accurate measurement of the adherence and internalization capabilities of engineered vectors. Here, we describe the use of scanning electron microscopy (SEM) to directly quantify vectors when they are external to the cell wall, and confocal microscopy to evaluate the vectors when they have internalized into the cytoplasm. By performing the invasion procedure on microscope coverslips, cells can be easily prepared for analysis using electron or confocal microscopes. Imaging the invasion complexes in high resolution can provide important insights into the behavior of bacterial vectors including E. coli, Listeria, and Salmonella when invading their target cells to deliver DNA and other molecules.
    Matched MeSH terms: Microscopy, Confocal*
  12. Hagiwara Y, Koh JEW, Tan JH, Bhandary SV, Laude A, Ciaccio EJ, et al.
    Comput Methods Programs Biomed, 2018 Oct;165:1-12.
    PMID: 30337064 DOI: 10.1016/j.cmpb.2018.07.012
    BACKGROUND AND OBJECTIVES: Glaucoma is an eye condition which leads to permanent blindness when the disease progresses to an advanced stage. It occurs due to inappropriate intraocular pressure within the eye, resulting in damage to the optic nerve. Glaucoma does not exhibit any symptoms in its nascent stage and thus, it is important to diagnose early to prevent blindness. Fundus photography is widely used by ophthalmologists to assist in diagnosis of glaucoma and is cost-effective.

    METHODS: The morphological features of the disc that is characteristic of glaucoma are clearly seen in the fundus images. However, manual inspection of the acquired fundus images may be prone to inter-observer variation. Therefore, a computer-aided detection (CAD) system is proposed to make an accurate, reliable and fast diagnosis of glaucoma based on the optic nerve features of fundus imaging. In this paper, we reviewed existing techniques to automatically diagnose glaucoma.

    RESULTS: The use of CAD is very effective in the diagnosis of glaucoma and can assist the clinicians to alleviate their workload significantly. We have also discussed the advantages of employing state-of-art techniques, including deep learning (DL), when developing the automated system. The DL methods are effective in glaucoma diagnosis.

    CONCLUSIONS: Novel DL algorithms with big data availability are required to develop a reliable CAD system. Such techniques can be employed to diagnose other eye diseases accurately.

    Matched MeSH terms: Microscopy, Confocal/methods
  13. Yang X, Wang S, King TL, Kerr CJ, Blanchet C, Svergun D, et al.
    Faraday Discuss, 2016 Jul 18.
    PMID: 27430046
    We have developed a new class of lanthanide nano-clusters that self-assemble using flexible Schiff base ligands. Cd-Ln and Ni-Ln clusters, [Ln8Cd24(L(1))12(OAc)39Cl7(OH)2] (Ln = Nd, Eu), [Eu8Cd24(L(1))12(OAc)44], [Ln8Cd24(L(2))12(OAc)44] (Ln = Nd, Yb, Sm) and [Nd2Ni4(L(3))2(acac)6(NO3)2(OH)2], were constructed using different types of flexible Schiff base ligands. These molecular nano-clusters exhibit anisotropic architectures that differ considerably depending upon the presence of Cd (nano-drum) or Ni (square-like nano-cluster). Structural characterization of the self-assembled particles has been undertaken using crystallography, transmission electron microscopy and small-angle X-ray scattering. Comparison of the metric dimensions of the nano-drums shows a consistency of size using these techniques, suggesting that these molecules may share similar structural features in both solid and solution states. Photophysical properties were studied by excitation of the ligand-centered absorption bands in the solid state and in solution, and using confocal microscopy of microspheres loaded with the compounds. The emissive properties of these compounds vary depending upon the combination of lanthanide and Cd or Ni present in these clusters. The results provide new insights into the construction of novel high-nuclearity nano-clusters and offer a promising foundation for the development of new functional nanomaterials.
    Matched MeSH terms: Microscopy, Confocal
  14. Marzuki, A.F., Masudi, S.M.
    MyJurnal
    Dentin morphology and the lesion found in dental caries have been studied for many years. It was first observed under optical microscopy, and later using electron microscopy. Confocal laser scanning microscopy (CLSM) applied with several fluorescent dyes such as alizarin red to see normal dentinal tubules. However, as far as authors aware, the CLSM studies of dentinal tubules in human caries using alizarin red is rare. The aim of this study is to examine histopathological and morphological changes in dentinal tubules of dentin caries stained with alizarin red using CLSM. Fifteen extracted carious teeth (premolar or molar) was collected and fixed in neutral formalin solution buffered with phosphate buffer, rinsed and stored in calcium free phosphate buffer saline (PBS) at 4°C. The specimens were dehydrated and embedded in resin. Longitudinal or cross sections were cut and polished and then stained with alizarin red S (100 μg/ml) in 0.5 M HCl solution for 24-48 hour at 37°C. After dehydration specimens were mounted on glass slide and examined under CLSM using epi-flourescent mode or transmission light mode with wave length of 512 nm. The images of dentinal tubules were taken serially and optimum images of three-dimensional structures were reconstructed using software of CLSM. Histopathological changes of dentinal tubules in human caries showed area of demineralized dentin, translucent zone, and normal area. The dentinal tubules were thin and had numerous branches. In conclusion, confocal microscopy revealed Study shows that confocal microscopy revealed histopathological changes in dentinal tubules affected by carious lesions.
    Matched MeSH terms: Microscopy, Confocal
  15. Kumarn S, Churinthorn N, Nimpaiboon A, Sriring M, Ho CC, Takahara A, et al.
    Langmuir, 2018 10 30;34(43):12730-12738.
    PMID: 30335388 DOI: 10.1021/acs.langmuir.8b02321
    The stabilization mechanism of natural rubber (NR) latex from Hevea brasiliensis was studied to investigate the components involved in base-catalyzed ester hydrolysis, namely, hydrolyzable lipids, ammonia, and the products responsible for the desired phenomenon observed in ammonia-preserved NR latex. Latex stability is generally thought to come from a rubber particle (RP) dispersion in the serum, which is encouraged by negatively charged species distributed on the RP surface. The mechanical stability time (MST) and zeta potential were measured to monitor field latices preserved in high (FNR-HA) and low ammonia (FNR-LA) contents as well as that with the ester-containing components removed (saponified NR) at different storage times. Amounts of carboxylates of free fatty acids (FFAs), which were released by the transformation and also hypothesized to be responsible for the like-charge repulsion of RPs, were measured as the higher fatty acid (HFA) number and corroborated by confocal laser scanning microscopy (CLSM) both qualitatively and quantitatively. The lipids and their FFA products interact differently with Nile red, which is a lipid-selective and polarity-sensitive fluorophore, and consequently re-emit characteristically. The results were confirmed by conventional ester content determination utilizing different solvent extraction systems to reveal that the lipids hydrolyzed to provide negatively charged fatty acid species were mainly the polar lipids (glycolipids and phospholipids) at the RP membrane but not those directly linked to the rubber molecule and, to a certain extent, those suspended in the serum. From new findings disclosed herein together with those already reported, a new model for the Hevea rubber particle in the latex form is proposed.
    Matched MeSH terms: Microscopy, Confocal
  16. Thomas AR, Mani R, Reddy TV, Ravichandran A, Sivakumar M, Krishnakumar S
    J Contemp Dent Pract, 2019 Sep 01;20(9):1090-1094.
    PMID: 31797835
    AIM: The aim of the study was to assess the antibacterial efficiency of a combination of 1% alexidine (ALX) and 5.25% sodium hypochlorite (NaOCl) against E. faecalis biofilm using a confocal scanning electron microscopy.

    MATERIALS AND METHODS: An estimated 120 human root dentin disks were prepared, sterilized, and inoculated with E. faecalis strain (ATCC 29212) to develop a 3-weeks-old biofilm. The dentin discs were exposed to group I-control group: 5.25% sodium hypochlorite (NaOCl) (n = 20); group II-1% ALX + 5.25% NaOCl (n = 40); group III-1% alexidine (ALX) (n = 40) (Sigma-Aldrich, Mumbai, India); group IV-negative control: saline (n = 20). After exposure, the dentin disks were stained with the fluorescent live/dead dye and evaluated with a confocal scanning electron microscope to calculate the proportion of dead cells. Statistical analysis was done using the Kruskal-Wallis and Mann-Whitney U test (p < 0.05).

    RESULTS: The maximum proportion of dead cells were seen in the groups treated with the combination of 1% ALX + 5.25% NaOCl (94.89%) and in the control group 5.25% NaOCl (93.14%). The proportion of dead cells presented in the 1% ALX group (51.79%) and negative control group saline (15.10%) were comparatively less.

    CONCLUSION: The antibacterial efficiency of a combination of 1% ALX and 5.25% NaOCl was more effective when compared with 1% ALX alone.

    CLINICAL SIGNIFICANCE: Alexidine at 1% could be used as an alternative endodontic irrigant to chlorhexidine, as alexidine does not form any toxic precipitates with sodium hypochlorite. The disinfection regimen comprising a combination of 1% ALX and 5.25% NaOCl is effective in eliminating E. faecalis biofilms.

    Matched MeSH terms: Microscopy, Confocal
  17. Chew ST, Eshak Z, Al-Haddad A
    Microsc Res Tech, 2023 Jul;86(7):754-761.
    PMID: 37078493 DOI: 10.1002/jemt.24323
    To assess the interfacial adaptation and penetration depth of three different bioceramic-based sealers (CeraSeal, EndoSeal MTA, Nishika Canal Sealer BG) compared to an epoxy resin-based sealer (AH Plus) in oval root canals. Fourty extracted single-rooted mandibular premolar with oval canal were prepared and randomly allocated according to the obturation into; CeraSeal, EndoSeal MTA, Nishika Canal Sealer BG and AH Plus. The roots were sectioned at 3, 6 and 9 mm from the apex. The sealer adaptation and the penetration depth were evaluated under confocal laser scanning microscope. One-way ANOVA and Repeated measure ANOVA were used to statistically analyze the data. Nishika Canal Sealer BG showed significantly higher sealer adaptation than EndoSeal MTA (P 
    Matched MeSH terms: Microscopy, Confocal
  18. Ghosh S, Mutalib HA, Kaur S, Ghoshal R, Retnasabapathy S
    Malays J Med Sci, 2017 Mar;24(2):44-54.
    PMID: 28894403 MyJurnal DOI: 10.21315/mjms2017.24.2.6
    PURPOSE: To evaluate corneal cell morphology in patients with keratoconus using an in vivo slit scanning confocal microscope.

    METHODS: A cross-sectional study was conducted to evaluate the corneal cell morphology of 47 keratoconus patients and 32 healthy eyes without any ocular disease. New keratoconus patients with different disease severities and without any other ocular co-morbidity were recruited from the ophthalmology department of a public hospital in Malaysia from June 2013 to May 2014. Corneal cell morphology was evaluated using an in vivo slit-scanning confocal microscope. Qualitative and quantitative data were analysed using a grading scale and the Nidek Advanced Visual Information System software, respectively.

    RESULTS: The corneal cell morphology of patients with keratoconus was significantly different from that of healthy eyes except in endothelial cell density (P = 0.072). In the keratoconus group, increased level of stromal haze, alterations such as the elongation of keratocyte nuclei and clustering of cells at the anterior stroma, and dark bands in the posterior stroma were observed with increased severity of the disease. The mean anterior and posterior stromal keratocyte densities and cell areas among the different stages of keratoconus were significantly different (P < 0.001 and P = 0.044, respectively). However, the changes observed in the endothelium were not significantly different (P > 0.05) among the three stages of keratoconus.

    CONCLUSION: Confocal microscopy observation showed significant changes in corneal cell morphology in keratoconic cornea from normal healthy cornea. Analysis also showed significant changes in different severities of keratoconus. Understanding the corneal cell morphology changes in keratoconus may help in the long-term monitoring and management of keratoconus.

    Matched MeSH terms: Microscopy, Confocal
  19. A Talip B, Snelling WJ, Sleator RD, Lowery C, Dooley JSG
    BMC Microbiol, 2018 11 26;18(1):196.
    PMID: 30477427 DOI: 10.1186/s12866-018-1335-0
    BACKGROUND: The field of diagnostics continues to advance rapidly with a variety of novel approaches, mainly dependent upon high technology platforms. Nonetheless much diagnosis, particularly in developing countries, still relies upon traditional methods such as microscopy. Biological material, particularly nucleic acids, on archived glass slides is a potential source of useful information both for diagnostic and epidemiological purposes. There are significant challenges faced when examining archived samples in order that an adequate amount of amplifiable DNA can be obtained. Herein, we describe a model system to detect low numbers of bacterial cells isolated from glass slides using (laser capture microscopy) LCM coupled with PCR amplification of a suitable target.

    RESULTS: Mycobacterium smegmatis was used as a model organism to provide a proof of principle for a method to recover bacteria from a stained sample on a glass slide using a laser capture system. Ziehl-Neelsen (ZN) stained cells were excised and catapulted into tubes. Recovered cells were subjected to DNA extraction and pre-amplified with multiple displacement amplification (MDA). This system allowed a minimum of 30 catapulted cells to be detected following a nested real-time PCR assay, using rpoB specific primers. The combination of MDA and nested real-time PCR resulted in a 30-fold increase in sensitivity for the detection of low numbers of cells isolated using LCM.

    CONCLUSIONS: This study highlights the potential of LCM coupled with MDA as a tool to improve the recovery of amplifiable nucleic acids from archived glass slides. The inclusion of the MDA step was essential to enable downstream amplification. This platform should be broadly applicable to a variety of diagnostic applications and we have used it as a proof of principle with a Mycobacterium sp. model system.

    Matched MeSH terms: Microscopy, Confocal/methods*
  20. Torey A, Sasidharan S
    Eur Rev Med Pharmacol Sci, 2011 Aug;15(8):875-82.
    PMID: 21845797
    Candida (C.) albicans infection in its biofilm mode of growth has taken centre point with the increasing recognition of its role in human infections due to the development of resistance to the commonly used antibiotic or phenotypic adaptation within the biofilm. Hence, in this study the inhibitory effect of methanol extract of Cassia (C.) spectabilis leaves was evaluated against biofilm forming C. albicans.
    Matched MeSH terms: Microscopy, Confocal/methods
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