Course work: confocal microscopy

Confocal microscope Confocal microscope allows to obtain three dimensional images of non-biological and biological specimens. The key point of this technique depend on the elimination of out of focus glare and increase the clarity, contrast and detection sensitivity using spatial filtering , point source of light for excitation and a pinhole confocal with excitation pinhole in front of the detector(2). Laser scanning confocal microscope (LSC M) used to analyse the structural details of thick specimens and providing 3-D volume renderings of living cells over time (singh A 1998). LSCM provides better understanding of structural details of cellular processes . this include (i) Cellular components and its location in the cell such as organelles, cytoskeletal elements and macromlecules (e.g. proteins, RNA and DNA). (ii) (iii) Tracing specific cells through a tissue Producing optical sections for stereo image production and three dimension reconstruction (iv) (v) Imaging in four dimension Ion imaging and fluorescence recovery after photobaeaching 33

LSCM used in the reflectance mode by removing or reducing in out of focus glare form nonfluorescent labels products or specimens formed in cytochemial detecting in situ hybridization 34 However, there are different application of

Application of the LSCM with different techniques such as Confocal reflected light microscopy used in imaging unstained 36 Confocal florescence microscopy used in FISH in thick sample 35. Also, in chromosomal and cytogenetic analysis 50, 3-D reconstruction of live tissue, cell- cell communication 70 and FRAP 69 Interactive laser cytometer used in membrane potential in single cells, Diffusion of membrane components and cytoplasmic structure 75, multi well screening for fluorescence quantitation in cell cultures.

Course work: confocal microscopy Confocal microscopy In microbiology Wall in 2010 evaluated the utility confocal microscopy in the identification of fungal and acanthmoeba infection. Confocal scans performed at OSU from 2002 trhough 2009 of 64 cases determined to be positive or negateive for infection based on clinical impression, outcome and culture which ware compared to confocal readings. Results showed that out of 101 studies and charts reviewed, the inclusion criteria were met by 64 confocal microscopy studies which had specificity for dectecing of infectious keratitis and had sufficient follow up to determine the efficacy of treatment . confocal microscopy identified fungal or acanthamoeba keratitis with 66% sensitivity and 71 specificity. Conclustion Confocal microscopy has many application in different kind of science . In microbiology, confocal microscoby is helpful in diagnosis of many microbial disease such fungal disease and protozoic infection and also mechanism of infection and immunity response mechanisms. Reference Singh A. and Gopinathan K.P (1998) confocal microscopy: A powerful technique for biological re J Cell Sci. 1994 May;107 ( Pt 5):1309-19. 33 Brian Storrie1,*, Rainer Pepperkok1,, Ernst H. K. Stelzer1,2 and Thomas E. Kreis1,, The intracellular mobility of a viral membrane glycoprotein measured by confocal microscope fluorescence recovery after photobleaching Journal of Cell Science 107, 1309-1319 (1994) 34 J Histochem Cytochem. 1989 Dec;37(12):1761-5.

Detection of diaminobenzidine reactions using scanning laser confocal reflectance microscopy.

Robinson JM, Batten BE Pp

35 Biotechniques. 1991 Oct;11(4):486-93.

Improved detection of in situ hybridization by laser scanning confocal microscopy.

Course work: confocal microscopy

Paddock S, Mahoney S, Minshall M, Smith L, Duvic M, Lewis D Okabe t teshima r furuno t t tungoe sawada 1996 69 J Cell Biol. 1991 Oct;115(1):245-55.

Influence of receptor lateral mobility on adhesion strengthening between membranes containing LFA-3 and CD2.

Chan PY, Lawrence MB, Dustin ML, Ferguson LM, Golan DE, Springer TA. 70 Carcinogenesis. 1991 Nov;12(11):1993-9.

Cell-to-cell communication: a differential response to TGF-beta in normal and transformed (BEAS-2B) human bronchial epithelial cells.

Albright CD, Grimley PM, Jones RT, Fontana JA, Keenan KP, Resau JH.

75 Yeh, g c lopaczynska j poore c m and phang j m 1992