Olympus FV1000 manual Diffusion Measurement Package, RICS-Raster Imaging Correlation Spectroscopy

1.5

1

130

0.5

125 Pixels

0

105

125105

130

Pixels

Diffusion Measurement Package

This optional software module enables data acquisition and analysis to investigate the molecular interaction and concentrations by calculating the diffusion coefficients of molecules within the cell.

Diverse analysis methods (RICS/ccRICS, point FCS/point FCCS and FRAP) cover a wide range of molecular sizes and speeds.

RICS—Raster Imaging Correlation Spectroscopy

Raster image correlation spectroscopy (RICS) is a new method for analyzing the diffusion and binding dynamics of molecules in an entire, single image. RICS uses a spatial correlation algorithm to calculate diffusion coefficients and the number of molecules in specified regions.

Cross correlation RICS (ccRICS) characterizes molecular interactions using fluorescent- labeled molecules in two colors.

point FCS—Point scan Fluorescence Correlation Spectroscopy

point scan fluorescence correlation spectroscopy (point FCS) analyzes intensity fluctuations caused by diffusion or binding/unbinding interactions of a protein complex. point FCS uses an auto correlation function to carry out operations on fluorescence signals obtained by continuous scanning of a single pixel on the screen.

point scan fluorescence cross-correlation spectroscopy (point FCCS) analyzes the fluctuation of fluorescent-labeled molecules in two colors. The coincidence of fluctuations occurring in two detection channels shows that the two proteins are part of the same complex.

point FCS and point FCCS can now be performed with a standard detector, eliminating the need for a special high-sensitivity detector.

FRAP Analysis

The Axelrod analytical algorithm is installed as a FRAP analysis method. The algorithm is used to calculate diffusion coefficients and the proportions of diffusing molecules.

Analytical methods according to molecule diffusion speeds

point FCS

Capable range

of measurement RICS

FRAP

Small molecules in solution

> 100

Proteins in solution

~ 100

Diffusion of proteins in cell

1 ~ 100

Lateral diffusion in cell membrane

< 0.1

Protein trafficking

< 0.01

Molecular complex

formation, aggregation

<< 0.001

15