Epi Image                                                                           TIRF Image.

What Is TIRF?


The facility currently has one Total Internal Reflection Fluorescence (TIRF) Microscope and one TIRF_FLIM system .TIRF is a powerful technique that uses an evanescent wave to excite flurophores that are present at the bottom of the specimen adjacent to the glass water interface. The optical section that is excited is in the range of 100nm-150nm . It is often employed to study cellular membrane activities, the dynamics of actin, cellular adhesion, cell movement, single molecular events, vesicle and protein tracking.

 TIRF is so named because it involves total internal reflection of the excitation light. The excitation light must strike the glass- medium interface at the critical angle (the angle at which no light passes from the glass slide to the medium). Instead the light is reflected and this results in an evanescent wave being produced. One of the advantages of TIRF is that the background and out of focus signals are dramatically reduced which allows you to view very dim fluorescent events.

 

               

Above image courtesy of  Juliana Schwarz R19 showing TIRF optical path.                                    Image confocal optical section verses TIRF

 

 

 

 

TIRF used to study membrane dynamics at the cell basal membrane. It can only be used to visualize the bottom plane of the cell. (Optical section of 150nm-200nm )

 
 

Above comparison of different imaging methods.

 

 


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Copyright 2009 Beatson Advanced Imaging Reaource
Last modified: 02/25/16