Messages

1

DAQ Software/Firmware / Re: Custom frame rates

« on: January 20, 2019, 05:33:34 AM »

Hi mathewrynes,
Yes you can modify the source code to get custom frame rates. In order to do this you will need to add a 45FPS option in the frame rate drop down list as well as edit the Miniscope DAQ firmware running on the Miniscope DAQ PCB. Just search for 'FPS', 'framerate' and 'frame rate' to locate relevant portions of the code. You will also need to read the CMOS imaging sensor data sheet to calculate the correct register values to achieve 45FPS.

When running at 60FPS, where do you see the large variations in frame rate? Is it in the numbers displayed in the Miniscope DAQ software or is in the timestamps in timestamps.dat? The CPU timer we use to calculate instantaneous frame rate and inter frame interval has a couple millisecond variability which could cause the calculated frame rate to bounce around a bit. Also try grabbing the newest version of the Miniscope DAQ software which might help.

Lastly, if you are seeing a significant increase in dropped frames then there is likely no non-hardware related solution.

2

Electronics / Re: Excitation LED got too weak after I replaced a new coax cable

« on: November 16, 2018, 04:03:32 AM »

Hi qdong8,
Can you tell me the specs of the new cable or post a link to it? If it is extremely thin then there is a chance it could decrease the total amount of current the LED can draw.

3

Behavior / Re: Sync Behavior - More TTLs sent than frames recorded

« on: October 30, 2018, 09:42:27 PM »

Hi hahnj,
This is due to the computer's USB hardware not being able to keep up with the data being sent out of the Miniscope DAQ. There are a couple ways to correct for this and luckily David Tank's group just published a paper with a detailed supplement on how to deal with this. Here is a link to that info: https://www.cell.com/neuron/fulltext/S0896-6273(18)30852-3. Section B-2 in the supplemental materials.

Hope this helps.

4

Optics / Re: Non Uniform illumination or vignetting in miniscope?

« on: October 19, 2018, 07:10:47 PM »

Hi kyrollosyanny,
Proper positioning of the excitation LED and half-ball lens will decrease vignetting but it is true that you can't get rid of it completely. During offline analysis you can normalize the pixel intensity across all columns which does a decent job of making the image uniform but with the downside of increasing noise around the edge of your image.

5

Surgery and Baseplating / Re: baseplate making tool

« on: October 03, 2018, 09:03:04 PM »

Hi Richardson,
This is a great idea. If you would like to share this picture, a short description, and your gcode on the miniscope.org site shoot me an email at dbaharoni_at_gmail.com. It would be a great addition!

6

Surgery and Baseplating / Re: nothing but gray

« on: August 10, 2018, 02:40:43 AM »

Hi hsiao,
You need to always use the objective GRIN lens in conjunction with the relay lens. If you try to image with just the relay lens you will never see anything in focus.

1. You remove the corpus callosum with aspiration, never forceps. I think we should have videos in the surgery power point presentation on miniscope.org under Workshop Resources.
2. You should be doing all your aspiration under a steroscope. You will visually see the fibers turn direction as you aspirate deeper.
3. We generally do not see much during implantation. It takes a could weeks for the imaging field to clear up to see a decent image.

7

Miniscope.org Suggestions / Re: Member pages broken

« on: August 09, 2018, 02:00:58 AM »

Thanks. I'll try and look into this soon.

8

DAQ Software/Firmware / Re: Camera not recognized

« on: July 26, 2018, 07:09:12 AM »

Hi amankapadia,
Once the EEPROM memory on the Miniscope DAQ has been programmed it will no longer show up in the Cypress Control Center but instead show up as a MINISCOPE under cameras in Device Manager. So that part seems to be working fine.

Try clicking the "Connect" button in the Miniscope software a few times in a row spaced out by a couple of seconds. If that doesn't connect your computer to the Miniscope video stream try using a different computer to see if it connects at all. There are a few other things to check..

* Make sure you are plugged into a USB3.0 port
* Make sure you have a decent quality USB3.0 cable
* Is there a chance you shorted the coax cable line in the recent past? This can cause a power surge through the L13 inductor resulting in all 3 green status LEDs lighting up but not being able to connect to the Miniscope video stream.

9

Surgery and Baseplating / Re: nothing but gray

« on: July 22, 2018, 04:56:29 PM »

Hi kjlawlor,
There are a few labs I know of successfully imaging in rats. We work most closely with Tad Blair's Lab at UCLA and they are getting really nice data sets. Surgeries are a bit trickier in rats since their brains are larger than mice's but you still need to get the implanted less withing a couple 100um of the cells you want to image. Also, I know getting the correct GCaMP virus was very important. I think they had the most luck using AAV9 but you should double check with them.

Bastijn van den Boom at NIN has also had success imaging in rats.

10

Surgery and Baseplating / Re: nothing but gray

« on: July 18, 2018, 05:17:53 PM »

Hi,

The lens was above dCA1 cell body over ~500um. I should lower it further.

Yea, you should really try to keep this distance to under 300um. Even 300um is a bit large.

But here comes other questions.
1. The working distance of #64-519 is 0, so I should implant the lens as close to the cell bodies of dCA1 as possible (at least < 250 um)? If I choose a lens with a 0.2 mm WD, then it gives me more space so that I can implant the lens like 0.2mm or more above the cell bodies?

You should always implant the lens as close as possible to the cells you want to image. The lenses from Edmund have a design wavelength different than what you will be using. This wavelength affects the actual pitch and working distance of the lens. So the quoted 0 or 0.2mm WD willbe different when imaging green fluorescence. But in general you want the effective pitch of your objective lens to be slightly under 0.25.

2. The medial hippocampus is higher than the lateral. Will you tilt the lens a little bit to fit the hippocampus?

I'm not sure. A little tilt might help with imaging.

3. In the dual lens system, the wiki page uses the title "Imaging With Thin GRIN Lenses". My question is that is it necessary to use a relay lens that is thinner than the objective lens? Will there be any problems if I use a relay lens with the same diameter with the objective lens?

No, it can be the same diameter.

You might want to contact Walter Boyles at GoFoton, walter.boyles_at_gofoton(dot)com, and request a custom 2mm diameter GRIN lens made by them. If you give him the working distance you want (something like 200um on both ends of the GRIN lens) and the overall length you need he can maybe make you a single lens that will work for you. This would result in a single GRIN lens instead of stacking multiple lenses.

11

Surgery and Baseplating / Re: nothing but gray

« on: July 16, 2018, 05:43:29 PM »

Thanks for the schematics. This clarifies things a bit. So you actually just want to use 3 of those GRIN lenses stacked end to end with no gaps in between them.  This will effectively make a single GRIN lens with a 0.75 pitch which will work for imaging with a short working distance of ~50um to 250um.

12

Surgery and Baseplating / Re: nothing but gray

« on: July 14, 2018, 06:33:22 AM »

Hi hsiao,
Can you clarify what you mean by stacking 2 objective lenses? Do you mean you put two of the #64-519 Edmund lenses end to end? If so this will create a ~0.5pitch GRIN lens which has very different optical properties than what the Miniscope is expecting as an objective lens. In other words, you have to keep the pitch of the objective lens to ~0.25 + 0.5*N pitch where N can be 0, 1, 2, ... We have imaged in rats by stacking 3 objective lenses together but using only 2 won't work.

 I am guessing the reason you saw clear signals outside the brain with a stack of 2 objective lenses is because you were imaging with a rather large working distance, a working distance much larger than what would work in the brain.

13

DAQ Software/Firmware / Re: "Screen Shot" and "Record" Functions not Working Properly

« on: July 12, 2018, 05:13:05 PM »

Hi Chancey,
I'm not sure exactly what would be causing this. Make sure you are running the .exe file on a local hard drive and not over a network. Maybe try it out on another computer to make sure you can get it functioning properly somewhere.

14

Optics / Re: Calibration

« on: July 10, 2018, 08:59:59 PM »

Hi Ryan,
The relevant GRIN lens information you need can be found here http://miniscope.org/index.php/GRIN_Lens_Information and here http://miniscope.org/index.php/Imaging_With_Thin_GRIN_Lenses.

15

Optics / Re: Calibration

« on: July 10, 2018, 07:11:04 AM »

Hi Ryan,
Just to clarify, are you trying to imaging with only these thin (1mm diameter or less) GRIN lenses or are you imaging in conjunction with a larger (1.8 or 2mm diameter) objective GRIN lens. You need the objective GRIN lens in place to see anything.