LeClair Lab Fluorender Protocol 2014

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Jonas Mitchell
5 years ago
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3. Using Flurender This prtcl reviews a few f the basic tls fr Flurender visualizatin, including: 1. Psitining the Image Rendered 2. Chsing the Render View mde 3. Adjusting Each Layer 4.

1 3. Using Flurender This prtcl reviews a few f the basic tls fr Flurender visualizatin, including: 1. Psitining the Image Rendered 2. Chsing the Render View mde 3. Adjusting Each Layer 4. Clipping the Vlume 5. Adding A Scale Bar 6. Saving Yur Wrk 7. Making Mvies. Opening the Image Open Flurender frm desktp. Select Open Vlume t pen yur prject. File shuld pen rendering a 3D image f all three channels. (It may take a few minutes) 3.1 Psitining the Image Rendered T rtate the image left click and drag the muse acrss the screen. This mves the bject within the 3 dimensinal vlume. Exact rtatin is pssible by assigning X, Y, and Z plane values and typing them int the prgram r by scrlling thrugh these parameters. T center the view n different parts f the image, click the middle scrll buttn and drag the muse acrss the screen. 1

2 T zm the view, click and drag with the right muse buttn. This can als be cntrlled with the zm scrlling tl fund n the right: T save changes, made click n save t default lcated under the manual zm bar. By hitting Reset t 0 the screen will return t the default (scanned) XYZ view. 3.2 Chsing Render View Mde Three views are available t visualize the rendered 3D image: layer, depth, and cmpsite. 2

Layer view- Layers vxels in a channel rder.

n the left. Right clicking and dragging a layer t mve it up r dwn.

Depth View- Renders all vxels accrding t their depth values.

3 Layer view- Layers vxels in a channel rder. The rder f the channels can be selected by changing the rder they appear in the wrkspace area n the left. Right clicking and dragging a layer t mve it up r dwn. will mve them and change the rder. Depth View- Renders all vxels accrding t their depth values. Regardless f channel, shallw vxels appear n tp; while deeper vxels disappear. Mst spatially true t the cnfcal data. Cmpsite View- Each channel is rendered individually and the layers are blended in a semi-transparent mde. Layer; Red n tp Depth Cmpsite 3.3 Adjusting Each Layer The gamma, brightness and equalizatin f each layer can be changed independently EXCEPT when in Depth mde. Yu must select each clr befre adjusting the layer prperties: 3

Scrlling dwn increases brightness. Equalizatin- impacts hw much filling each clr accmplishes. T designate ne set f parameter fr all channels, select Link t link all channels. 3.

4 Gamma- impacts hw much surface area is illuminated in the image. Scrlling dwn increases surface area illuminatin. A gamma value greater than 1 highlights signals f lw intensity A gamma value less than 1 highlights signals f high intensity Luminescence- impacts hw bright each layer appears in the image. Scrlling dwn increases brightness. Equalizatin- impacts hw much filling each clr accmplishes. T designate ne set f parameter fr all channels, select Link t link all channels. 3.4 Clipping the Vlume The clipping feature allws yu t subtract ne r mre channels in selected areas f yur image. Yur first three grups in yur wrkspace will be blue, green, and red. (Yur first grup is blue, even thugh it has a red, green and blue light bulb in it.) Adjusts blue channel! Adjusts green channel! Adjusts red channel! The current selectin (blue light bulb in Grup 1) wuld allw yu t adjust the slider f the blue channel in Clippings. T clip the green channel, yu wuld select the Grup 2 green light bulb. Similarly, t clip the red channel, select the grup three red light bulb selectin. Select the channel that yu wish yu mdify. Then, lk at the Clippings Planes Bx: 4

The results will lk like this: Yu can adjust the level f the clip s the

5 If yu mve the X1 slider, Flurender will clip away the selected channel in the X directin. The results will lk like this: Yu can adjust the level f the clip s the desired amunt f that channel remains. The Y1 tggle is similar but perates alng the Y-axis. The results will lk like this: 5

Finally, the Z1 tggle will clip visibility f the selected channel alng the z-axis. This allws yu t eliminate superficial structures in that channel and bserve the deep nes.

6 Finally, the Z1 tggle will clip visibility f the selected channel alng the z-axis. This allws yu t eliminate superficial structures in that channel and bserve the deep nes. The results will lk like this: The X2, Y2, and Z2 sliders all perfrm the same functin, but in the ppsite directin. Yu can als adjust the angle f the clip t cut n diagnals. 3.5 Adding a Scale Bar Flurender easily adds an accurate scale bar t yur image, using the riginal cnfcal data. At the bttm f the screen there are tggles fr Scale Bar and Text. Here yu can set the visibility f the scale and the assciated units. Yu must make the scale bar visible BEFORE exprting files r mvies r it will NOT appear 6

3.6 Saving Yur Wrk There are several ptins fr saving yur wrk: 1. Yu may save yur entire prject and cntinue wrking n it later by selecting the Save Prject buttn in the tp left crner.

Yu may als take a screensht f yur 3D image by clicking Capture alng the tp f the viewing windw. This will allw yu t save the render view as a simple 2D image. 3. Yu may save a mvie file that yu create.

7 3.6 Saving Yur Wrk There are several ptins fr saving yur wrk: 1. Yu may save yur entire prject and cntinue wrking n it later by selecting the Save Prject buttn in the tp left crner. When yu wish yu retrieve this prject later, pen Flurender and select the Open Prject buttn in the tp left crner. 2. Yu may als take a screensht f yur 3D image by clicking Capture alng the tp f the viewing windw. This will allw yu t save the render view as a simple 2D image. 3. Yu may save a mvie file that yu create. This is cvered in the next sectin (3.6). 3.6 Making mvies T make mvies f the 3D image rtatins select 3D Rt in the bttm left crner Select the plane (X r Y) that the specimen will be rtating n Fr Rtatin Deg: set hw many degrees the image will rtate between each sht. Preview can be used t view a sample f the vide. T save it select Save then select a destinatin flder. Yur mvie will save as a series f TIFF files. In rder t cnvert the series f TIFFs int a mvie, yu will have t use Phtshp and VirtualDub, which is cvered in the next mdule. 7

Scroll down to New and another menu will appear. Select Folder and a new

Scroll down to New and another menu will appear. Select Folder and a new Creating a New Flder Befre we begin with Micrsft Wrd, create a flder n yur Desktp named Summer PD. T d this, right click anywhere n yur Desktp and a menu will appear. Scrll dwn t New and anther menu will