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Monitoring lysosomal fusion in electrofused hybridoma cells

Monitoring lysosomal fusion in electrofused hybridoma cells,10.1016/j.bbamem.2007.10.013,Biochimica Et Biophysica Acta-biomembranes,Mateja Gabrijel,Ma

Monitoring lysosomal fusion in electrofused hybridoma cells   (Citations: 3)
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Dendritic and tumor cells are fused to produce hybridoma cells, which are considered to be used as cellular vaccines to treat cancer. Previous strategies for hybridoma cell production were based on the quantification of the electrofusion yield by labeling the cytoplasm of both parental cell types. However, a better physiological strategy would be to label subcellular structures related directly to the antigen presentation process. Therefore, we here electrofused the same amount of CHO cells stained with red and green fluorescent dextrans and have monitored the yield of hybridoma cell formation by measuring the fusion of red and green late endocytic organelles that are involved in antigen presentation. By using confocal microscopy, the level of fused, fluorescently labelled late endocytic compartments in a single hybridoma cell was determined. The results demonstrate that organellar fusion occurs in hybridomas, which is time- and temperature-dependent. This approach therefore provides a new method for the hybridoma cell vaccine evaluation, which is based on the intracellular physiological mechanism of antigen presentation.
Journal: Biochimica Et Biophysica Acta-biomembranes - BBA-BIOMEMBRANES , vol. 1778, no. 2, pp. 483-490, 2008
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    • ...A review of the published literature showed that different authors use different electrofusion protocols, detection methods and quantifications of fused cells (Gabrijel et al. 2008; Jaroszeski et al. 1994; Ramos et al. 2002; Scott-Taylor et al. 2000; Wang and Lu 2006); therefore, the results of different studies cannot be directly compared...

    Marko Ušajet al. Cell–Cell Electrofusion: Optimization of Electric Field Amplitude and ...

    • ...A line profile of fluorescent intensities is frequently examined (Kreft et al. 2006 ;S tenovec et al.2007a, b; Gabrijel et al. 2008)...
    • ...Line profiles of pixel intensities are used to measure distances in image and to compare the colocalization of markers in the images (Diéguez-Uribeondo et al. 2003; Kreft et al. 2006; Gabrijel et al. 2008)...
    • ...Line profiles were also used to determine the appearance of a late endocytic compartment in several optical planes of hybridoma cells, where dendritic and tumour cells are fused (Gabrijel et al. 2008)...

    Marko Kreftet al. Analysis of confocal images using variable-width line profiles

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