Differences
This shows you the differences between two versions of the page.
Both sides previous revision Previous revision Next revision | Previous revision | ||
software:labview [2015/06/17 12:34] Jon Daniels |
software:labview [2016/02/14 13:41] (current) Abhishek Kumar |
||
---|---|---|---|
Line 1: | Line 1: | ||
====== Labview | ====== Labview | ||
- | The page has been created, you can go ahead and edit it. | + | LabView implementations have been created |
- | Start by just entering your text here. | + | In general LabView is recommended only for groups doing significant custom software development within an existing LabView framework. |
- | Before saving your changes it is a good idea to use the **Preview** button. | ||
- | FIXME Go ahead and delete all this text except the top line (the header) and add your content! | + | |
+ | This tutorial provides an outline of the front panel controls of two LabVIEW (LV) programs developed by the Shroff lab at the NIH to control new functionality in diSPIM1,2: stage-scanning | ||
+ | We previously described a similar program for controlling diSPIM in ‘conventional’ image acquisition mode (synchronously sweeping the light sheet and detection focal plane through the sample)2; the LV programs described below use the same hardware. | ||
+ | |||
+ | **A. Schematic of example waveforms provided to the diSPIM for hardware control:** Waveforms are generated by the DAQ and provided to the various diSPIM hardware components. See the MBL Biological Bulletin paper for more information. | ||
+ | |||
+ | {{: | ||
+ | |||
+ | The ‘Y-Galvo Amplitude’, | ||
+ | |||
+ | **B. Stage-scan LV front panel details:** | ||
+ | |||
+ | {{: | ||
+ | |||
+ | 1. **Control Mode Selection: | ||
+ | **DiSPIM Module:** Used to move the diSPIM module housing the two diSPIM objectives. User can define the distance (in mm) and LV provides a serial command to the TIGER Controller, moving the LS-50 motorized stage to the specified position. | ||
+ | **GFP Image Acquisition: | ||
+ | **mCherry Image Acquisition: | ||
+ | **Dual Color Image Acquisition: | ||
+ | **Parameter Adjustment: | ||
+ | |||
+ | 2. **Time Delay:** Extra, ‘pre-acquisition’ time delay is provided to the X-stage for acceleration. This way, when image acquisition starts, the stage is moving with a constant speed. A typical value is 10 um. | ||
+ | |||
+ | 3. **Various Imaging Parameters: | ||
+ | |||
+ | **Time/ | ||
+ | **Step size:** in um | ||
+ | **# of Planes:** i.e. the number of planes per volume | ||
+ | **Lightsheet width:** In um, and depends on the camera ROI. | ||
+ | **Speed:** Defines the speed of motorized stage and is dependent on the timing and step size. We typically use 0.1 mm/s for single color, 1 um z’ step size (equivalent to 0.7 um z step size). | ||
+ | **Number of time points:** The number of volumetric time points in the image acquisition. | ||
+ | **Delay b/w time points:** In seconds | ||
+ | **Start Position:** The X-coordinate of the sample being imaged. Once the sample is positioned using the bottom camera, the X-coordinate of the stage can be determined. | ||
+ | **Laser Intensity (488 nm):** arbitrary units | ||
+ | **Laser Intensity (561 nm):** arbitrary units | ||
+ | |||
+ | 4. **Camera ROI Settings:** Set ROI depending on sample size being imaged. | ||
+ | |||
+ | 5. **File Path and File Name:** The LV Program saves the acquired images in HCIMAGE format (Hamamatsu proprietary format) and then converts the data into TIFF format. The file path for both formats needs to be provided. LV creates sub directories named SPIMA and SPIMB in the parent directory. TIFF conversion can be performed during acquisition or after, using the TIFF conversion sub VI. The file name needs to be changed for each run. For time series, the file name is appended with the time point number. | ||
+ | |||
+ | 6. **Galvo and Piezo Offsets:** | ||
+ | **SPIMA & B Z-galvo:** Used to position the lightsheet so that it is well synchronized with the detection piezo. Voltage can be set between 0-4 V. | ||
+ | **SPIMA & B Y-galvo:** Defines the laser scan start position on the camera ROI (range 0-4 V). | ||
+ | **SPIMA & B Piezo:** Used to position the imaging objective and excitation beam at the appropriate position (range 0-10 V). | ||
+ | |||
+ | **C. Stage-scan with Slit-scan LV front panel details: | ||
+ | All controls in this mode are the same as described above, | ||
+ | |||
+ | {{: | ||
+ | |||
+ | 1. **Light Sheet Width:** Set to zero in this mode. | ||
+ | |||
+ | 2. **Exposure Time (us):** A user defined parameter for slit scan. Please look at the Hamamatsu Flash 4.0 V2 camera manual for more details about lightsheet mode, ROI, readout direction and timing information. | ||
+ | **Readout Time (us):** The readout time of a column of pixels for the Flash 4.0 Hamamatsu camera. This can be set in multiples of 10 us. | ||
+ | Slit width is defined as the exposure time divided by the readout time. | ||
+ | |||
+ | 3. **SPIMA and B Amplitude for Single and Dual Color:** This control is given to set the amplitude of each arm individually. This is a critical parameter for proper synchronization of rolling shutter with the laser scan (y-scan). **SPIMA and B Camera Delay:** This sets the camera trigger delay with respect to the Y-galvo scan. This is in % of total time per plane for two views. Typical values are ~2.4-3.2 ms delay assuming 8 ms/plane for each view. This is ~15-20 % delay of the total time/plane (16 ms) for two views. | ||
+ | |||
+ | **D. Examples showing the effect of Y-offset, Y-Amplitude and Camera Trigger Delay in slit scan mode.** | ||
+ | |||
+ | See also the waveform schematic in section A. Parameters are optimized using dye solution (here, fluorescein). The bright | ||
+ | |||
+ | {{: | ||