Initial Testing of Assembled Miniscopes - Revision history

Myqlavu: /* Testing the Stability of the DAQ Software */

2017-10-11T20:43:14Z

‎Testing the Stability of the DAQ Software

Myqlavu: /* Imaging GFP Slides */

2017-10-11T18:19:47Z

‎Imaging GFP Slides

Jjulian at 13:42, 25 September 2017

2017-09-25T13:42:29Z

Chrisl1026: /* Imaging GFP Slides */

2016-09-21T22:12:29Z

‎Imaging GFP Slides

Wilsona2: /* Testing the Stability of the DAQ Software */

2016-06-20T18:27:54Z

‎Testing the Stability of the DAQ Software

Jesper.vernooij: /* Imaging Your Surroundings */

2016-05-16T22:21:04Z

‎Imaging Your Surroundings

DAharoni: /* Imaging Calibration Slides */

2016-04-21T23:28:10Z

‎Imaging Calibration Slides

DAharoni: /* Imaging Calibration Slides */

2016-04-21T23:27:28Z

‎Imaging Calibration Slides

DAharoni: /* Imaging Calibration Slides */

2016-04-21T23:11:20Z

‎Imaging Calibration Slides

DAharoni: /* Imaging Calibration Slides */

2016-04-21T23:00:20Z

‎Imaging Calibration Slides

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 *#Observe the current frame rate (labeled with a red '2' in the picture to the right). It should be stable within 1 FPS from the expected value. This is just an approximate measure of the frame rate, the frame rate of the recorded video should be extremely stable.
 *#You can also connect to a behavioral camera to add strain on your system. Generally the behavioral camera will have larger fluctuations in the displayed frame rate.
-*'''Write speed of video data:''' You want to make sure your computer is able to write the uncompressed video data to your HDD or SSD as quick (and hopefully much quicker)than the rate at which you are acquiring it. Slow or encryted hard drives can be a source of problems here.
+*'''Write speed of video data:''' You want to make sure your computer is able to write the uncompressed video data to your HDD or SSD as quick (and hopefully much quicker than) as the rate at which you are acquiring it. Slow or encryted hard drives can be a source of problems here.
 *#Connect a scope to the DAQ Box and then the DAQ Box to the computer.
 *#Open up the DAQ software and connect to the Miniscope.

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 {{#ev:youtube|https://youtu.be/lxEUkP-YI8g|640|right}}
 Similar to imaging test slides, you can use your Miniscope to image brain slice slides expressing GFP or slides with other fluorescent sources. A few key points for imaging slides are listed below:
-*Your scope has a green bandpass filter (500nm to 550nm) in front the the CMOS imaging sensor. This means only will only be able to see green fluorescence.
+*Your scope has a green bandpass filter (500nm to 550nm) in front the the CMOS imaging sensor. This means you will only be able to see green fluorescence.
 *Take note of the thickness of your cover slip, usually around 170um thick. You will be imaging through the cover slip to reach the fluorescing sample. If you are using less than a 15mm focal length achromatic lens, you will likely not be able to focus past the bottom of the cover slip. Even a 15mm focal length achromatic lens will generally just reach about 50um below the bottom of the cover slip with the focusing slider pushed to its lowest position.

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 The video to the right is of a TetTag GFP slide being imaged with a 15mm focal length achromatic lens and 0.25 pitch GRIN lens. A few notes:
-*Their are some compression artifacts present due to YouTube's compression of the video.
+*There are some compression artifacts present due to YouTube's compression of the video.
 *The disk of light near the center of the field of view is due to reflection of the fluorescent light between the cover slip and bottom of GRIN lens... you should not see this disk when imaging in vivo.
 *The left and right portions of the field of view are dimmer than what you should expect to see with your own system. The outer dimness was greatly reduced in the newer versions of the Miniscope main body.

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 *Take note of the thickness of your cover slip, usually around 170um thick. You will be imaging through the cover slip to reach the fluorescing sample. If you are using less than a 15mm focal length achromatic lens, you will likely not be able to focus past the bottom of the cover slip. Even a 15mm focal length achromatic lens will generally just reach about 50um below the bottom of the cover slip with the focusing slider pushed to its lowest position.
-The video to the left is of a TetTag GFP slide being imaged with a 15mm focal length achromatic lens and 0.25 pitch GRIN lens. A few notes:
+The video to the right is of a TetTag GFP slide being imaged with a 15mm focal length achromatic lens and 0.25 pitch GRIN lens. A few notes:
 *Their are some compression artifacts present due to YouTube's compression of the video.
 *The disk of light near the center of the field of view is due to reflection of the fluorescent light between the cover slip and bottom of GRIN lens... you should not see this disk when imaging in vivo.
 *The left and right portions of the field of view are dimmer than what you should expect to see with your own system. The outer dimness was greatly reduced in the newer versions of the Miniscope main body.

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 *#If the video stream is still present (not Red Screen of Doom) your system should not have any driver or OS issues.
 *'''Streaming video frame rate:''' The default frame rate of your Miniscope system is 30FPS but can be adjusted using the drop down 'Frame Rate' box in our DAQ software (labeled with a red '1' in the picture to the right).
-*#Connect a cope to the DAQ Box and then the DAQ Box to the computer.
+*#Connect a scope to the DAQ Box and then the DAQ Box to the computer.
 *#Open up the DAQ software and connect to the Miniscope.
 *#Observe the current frame rate (labeled with a red '2' in the picture to the right). It should be stable within 1 FPS from the expected value. This is just an approximate measure of the frame rate, the frame rate of the recorded video should be extremely stable.
 *#You can also connect to a behavioral camera to add strain on your system. Generally the behavioral camera will have larger fluctuations in the displayed frame rate.
 *'''Write speed of video data:''' You want to make sure your computer is able to write the uncompressed video data to your HDD or SSD as quick (and hopefully much quicker)than the rate at which you are acquiring it. Slow or encryted hard drives can be a source of problems here.
-*#Connect a cope to the DAQ Box and then the DAQ Box to the computer.
+*#Connect a scope to the DAQ Box and then the DAQ Box to the computer.
 *#Open up the DAQ software and connect to the Miniscope.
 *#Click 'record' and observe the 'Write Speed (fps)' (labled with a red '3' in the picture to the right). This box displays the current write speed of your data. The number displayed will fluctuate but should stay above the acquisition frame rate of your Miniscope. If the write speed falls below the acquisition frame rate, video frames will be written into a circular buffer which is 256 frames long. If the write speed stays low for too long the software will begin to overwrite frames in the circular buffer. The 'timestamps.dat' file created during recording keeps track of the buffer size and is an addition place you can look when evaluating the write speed of your computer.

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 == Imaging Your Surroundings ==
 [[File:ImagingWithoutGRINLens.png|right|500px]]
-An assembled scope can image your surrounds when a GRIN lens is '''not placed''' into the hole in the base of the scope. Connect the scope to your computer and then point the base of the scope toward objects that are illuminated with room or sun light. A few notes
+An assembled scope can image your surrounds when a GRIN lens is '''not placed''' into the hole in the base of the scope. Connect the scope to your computer and then point the base of the scope toward objects that are illuminated with room or sun light.
 *The scope has a green bandpass filter sitting before the CMOS imaging sensor. This means you will only be collecting light between 500nm and 550nm.
 *Most objects in the environment will not fluoresce under the blue excitation LED. This means most object will not show up unless being illuminated by a light source that contains green wavelengths such as room light or sunlight.

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 *The middle picture shows a simple GRIN lens holder we had made. This holder is a block of Delrin plastic with different diameter holes drilled into it. a GRIN lens can be placed into the holder and then set on top of the modified test slide. A Miniscope can then be set onto of the holder to image the slide. The holder does a nice job keeping the surface of the GRIN lens co-planer to the surface of the test slide.
 *The right picture should an example configuration to image a test slide. This picture is of a scope that has been modified, [Imaging With Thin GRIN Lenses], to hold a GRIN lens in its base. While the picture shows the scope being held by hand, one could easily mount the scope in a clamp for more stable imaging. If using a GRIN lens holder as described above, a similar configuration would be used except the GRIN lens and holder would be in place of the bare GRIN lens.
 <br clear=all>

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 This is probably the most important initial test you can do with your Miniscope system. Not only can it uncover issues in your system but will also give you a good sense of how the optics, imaging, and focusing slider work. The figure to the right shows a modified test/calibration slide (left), optional GRIN lens holder (middle), and example configuration for imaging a test slide (right).
 *The modified test/calibration slide is a [http://www.thorlabs.us/navigation.cfm?guide_id=2332 resolution test slide] with [http://www.amazon.com/JVCC-Stage-Set-Spike-Tape-Fluorescent/dp/B000QDVNH0/ref=sr_1_12?s=industrial&srs=2529683011&ie=UTF8&qid=1461180753&sr=1-12 green fluorescing tape] attached to the underside of the glass. It is important that the printed surface of the test slide be the side that is closest to the GRIN lens. Different thicknesses of cover glass can be used between the test slide and GRIN lens to get a feeling for imaging at different depths.
+*The middle picture shows a simple GRIN lens holder we had made. This holder is a block of Delrin plastic with different diameter holes drilled into it. a GRIN lens can be placed into the holder and then set on top of the modified test slide. A Miniscope can then be set onto of the holder to image the slide. The holder does a nice job keeping the surface of the GRIN lens co-planer to the surface of the test slide.
 <br clear=all>

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 == Imaging Calibration Slides ==
 [[File:ImagingTestSlide.png|right|500px]]
 [[File:MiniscopeCalibrationSlide.png|thumb|right|500px|Image of a calibration slide with 9.8μm line spacing (superimposed red boxes are 10px x 10px)]]
-[http://www.amazon.com/JVCC-Stage-Set-Spike-Tape-Fluorescent/dp/B000QDVNH0/ref=sr_1_12?s=industrial&srs=2529683011&ie=UTF8&qid=1461180753&sr=1-12 Green Fluorescing Tape]
+This is probably the most important initial test you can do with your Miniscope system. Not only can it uncover issues in your system but will also give you a good sense of how the optics, imaging, and focusing slider work. The figure to the right shows a modified test/calibration slide (left), optional GRIN lens holder (middle), and example configuration for imaging a test slide (right).
 <br clear=all>

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 == Imaging Calibration Slides ==
 This is probably the most important initial test you can do with your Miniscope system. Not only can it uncover issues in your system but will also give you a good sense of how the optics, imaging, and focus slider work.
-[[File:MiniscopeCalibrationSlide.png|thumb|right|500px|Image of a calibration slide with 9.8μm line spacing (superimposed red boxes are 10px x 10px)]]
 [http://www.thorlabs.us/navigation.cfm?guide_id=2332 Test Slides]