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NeuroCCD-SM Cooled-CCD Imaging System
Start-up Procedures


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  NeuroPlex

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  FastCMOS-128X

 

  NeuroCMOS-DW

 

 

 

 

 

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  MacroScope-II & -IIa

 

 

 

 

 

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Camera Gain Settings  |
Optimal use of the CMOS Camera 
 

Initial Start-up Procedures

1.

Turn on computer

 

2.

Turn on camera (switch on power supply)

 

3.

Start the NeuroPlex software (click on NeuroPlex icon)

 

4.

>> Acquire

 

5.

Set the offsets

 

>> CMOS Camera

The process of calculating and recording the offsets for a specific camera is done by us before shipping it. Unlike some 8 bit A/D cameras, once these offsets were set for the NeuroCMOS-DW, there is no need to change them during the experiments. Moreover, using 14 bit digitization one can resolve the dark noise in most gains and rates. Thus changing the offsets is not very useful.

In some rare situations, an experiment could be improved by reducing the dynamic range (increase bit resolution) of the camera. If you believe this is the case, and would like to change the offsets, please refer to the help manual.

 

6.

Test the response to light using the LED

 

We have already tested the camera with LED for synchronization. All the test files are stored on the system computer in C:\Data\Camera_Test\ .

 

8.

Setting Camera Amplifier Gain:

 

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It is recommended that the following focusing procedures (8a and 8b) be done under low light levels to minimize possible photo-bleaching or photo-damage to the preparation.

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8.a.

Open the shutter

 

 

8.b.

>> Acquire Continuously

 

 

...and adjust the microscope focus on the preparation.

 

 

8.c.

Close the shutter and increase light level to maximum

 

 

8.d.

>> Acquire One Frame

 

 

Normally using >> Subtraction (auto shutter control) and >> Auto (min/max) to check that the light levels are neither too low or too high.

 

 

8.e.

>> Histogram

 

 

If the values are < 0.5 volts, then you may be able to increase the signal-to-noise ratio if you change the acquisition settings in a direction to increase the values (i.e. use a lower frame rate and/or high gain). Repeat step 8d then go to step 9.

If the values are > 8.5 volts and appear piled up at a high-voltage boundary, it is likely that many pixels are saturated. (Saturated pixels will appear as flat traces when you take data.) Look for many values between 9 and 10 volts. If it appears that there are a substantial number of saturated pixels, change the acquisition settings in a direction to decrease the values (i.e. use a faster frame rate and/or lower gain). Repeat step 8d. If the values still appear piled up at a boundary at 2000 fps and a gain of 1, then the only way to avoid saturation is to lower the incident light intensity. (You can use a faster frame rate than you need for your signal frequency and still not save excessive data by using Temporal Bin after Acquisition.)

 

 

9.

Select the acquisition parameters

 

Most functions are controlled in the left panel; the parameters in the center panel are used for signal averaging. Additional information about the acquisition functions can be found in the on-line manual.

 

10.

>> Take Data

 

 

Optimal Use of the CMOS camera

 

 

1.  

CMOS imagers have fixed pattern in their dark offsets, so it's recommended to use dark frame subtraction in order to obtain a better image display.

2.

In order to achieve optimum signal-to-noise ratio, it is generally bestto operate in the shot-noise limited condition.
In the 1Me- well size setting, the read noise is below 100e- and the camera is shot-noise limited at light levels down to less than 5% of saturation for either 5X or 2X for camera gain.
In the 12Me well size setting, the camera won't be shot noise limited unless the light level is over 10% of saturation for 2X and 20% of saturation for 5X. So we recommend that the reading of resting light intensity (RLI) of your region of interest be brighter than 3V** (after dark frame subtraction) at 5X and 1.5V at 2X. An increased RLI can be achieved by increasing the incident light level or reducing the frame rate. If you cannot get enough light to get an optimal S/N ratio at the 12Me well size setting, it may be best to reduce the incident light intensity or to speed up the frame rate so you can use the 2X - 1Me well size setting. In the circumstance where you increased the frame rate, temporal binning after acquisition can be used to return to the desired frame rate with no noise penalty.
The same principle applies to even higher light intensities when you have to switch to the 100Me well size setting, e.g. for absorption experiments. At this setting, the shot noise is smaller in proportion to the total light intensity and is smaller than the read noise at almost all intensities except for near saturation of 100Me-. However, 0.1% changes in signal should be easily detected if you have above 2-3V resting light intensity (with dark frame subtraction). Adjust the incident light level, frame rates and gain settings to increase the RLI. Or adjust in the other direction so that you can use the 12Me well size setting.
            In general, 5X has just a slightly lower read noise and 2X uses the full well for the chip setting.
           
** 10V is the full range for each setting.