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Spot Counting

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Monitoring autophagy modulation by nanoparticles using multispectral smaging		   Technical Information:
Quantitation of gamma-H2AX spots on the ImageStream (09-008)
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Merck:/Freestyle/BI-Bioscience/Cell-Analysis/amnis/spot-counting-100x100.jpgMany applications take advantage of the ImageStream®X's capability of counting the number of spots in an image. DNA repair proteins form foci around double stranded breaks that when stained look like small punctate spots; probes that identify viral DNA tethers will form spots in infected cells; phagocytosed particles appear as distinct spots within cells. These are just a few example applications enabled by the ability of Amnis® imaging flow cytometry to quantify spot counts within cell populations faster and easier than manual microscopy.

Quantitation of γ-H2AX foci in Irradiated Cells Using the ImageStream®X

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Phosphorylated H2AX (γ-H2AX) facilitates recognition and repair of DNA double strand breaks (DSBs) that may occur from exposure to ionizing radiation. Staining irradiated cells for γ-H2AX reveals nuclear foci that are readily observed microscopically in a dose response manner. Irradiated cells were analyzed for the number of spots in the nuclear region using advanced masking techniques that identify the punctate staining. Morphological measurements employed in this analysis including object shape, size, and punctate fluorescence spot counting emphasizing the advantages of quantitative multiparametric image analysis on large numbers of cells provided with the ImageStream®X

Babesia Spot Count Within RBC

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Babesiosis is a malaria-like disease caused by infection of red blood cells by the parasite Babesia. While healthy individuals may be asymptomatic, infection can be fatal in immune-compromised people. Using the ImageStream®X, Babesia were identified within infected RBC by staining parasite DNA with YOYO1. The stage of infection can be inferred by counting the number of infected cells and the number of parasites in each RBC.
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