Linear unmixing, spectral images

Standard linear unmixing of a spectral image of a sample composed of two fluorophores yields a measure of the concentration of each fluorophore present for each pixel. If FRET is occurring, linear unmixing will produce an apparent donor concentration ( apparent) that underestimates the true donor concentration (d) by a factor of 1 -ED ... 

Linear unmixing is applied to each spectral image to produce four measurements at each pixel, Apparent an[Pg.384]

Linear unmixing of the spectral image acquired at will also yield two observables ... 

Multi-spectral imaging and linear unmixing add a whole new dimension to laser scanning fluorescence microscopy. Biotechniques 31, 1272-8. 

Zimmermann, T. (2005). Spectral imaging and linear unmixing in light microscopy. Adv. Biochem. Eng. Biotechnol. 95, 245-65. 

Thaler, C. and Vogel, S. S. (2006). Quantitative linear unmixing of CFP and YFP from spectral images acquired with two-photon excitation. Cytometry A 69, 904—11. 

LSM 510 META software to process time-lapse fluorescence images, and spectrally resolved images of Lambda Stack using the Linear Unmixing Function of LSM510 META. 

Spectral Imaging and Linear Unmixing in Light Microscopy... 

Keywords Spectral imaging Linear unmixing Fluorescent proteins - FRET... 

Fig. 2 Spectral imaging of fluorescence signals. Contributions of CFP, GFP and YFP to eight successive spectral channels are shown. The distribution of emission signal to the channels is a direct representation of the fluorophore emission spectrum and constitutes a spectral signature. With linear unmixing using these spectral signatures as reference, even combined and mixed signals can be clearly separated into the fluorophores that contribute to the total signal...

Both determined (n channels=n fluorophores) as well as overdetermined systems (n channels>n fluorophores) can be used for spectral imaging and linear unmixing [19,20]. Are there differences in performance ... 

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