Exposure Control and Camera Sensitivity
Digital cameras require some level of exposure time adjustment in order to control the levels of brightness, color saturation, and contrast in the final image. Three factors defining correct exposure in a charged coupled device (CCD) camera are the visible light spectrum wavelength-dependent sensitivity, the intensity of illumination, and the length of time the CCD is exposed to light.
One of the most important variables in digital photomicrography is to assure the correct exposure to match the dynamic range and sensitivity of the camera system. The DXM 1200 control software (ACT-1) is equipped with an exposure time control panel (Figure 2) that allows the microscopist to produce exact and reproducible exposures from images observed in the microscope. The ACT-1 software can produce exposures at any time interval between one twelve thousandth of a second (1/12000) up to 170 seconds.
A bracketed series of exposures captured using the DXM 1200 and a Nikon Eclipse E600 microscope operating in brightfield mode are presented in Figure 1. The specimen is a stained thin section of fern tissue (Psilotum). Each image in the series was exposed for a time period that was 50 percent less than the preceding image. For example, image (a) in Figure 1 was exposed for one-half (1/2) second, while image (b) was exposed for one-quarter (1/4) second, and image (c) was exposed for one-eighth (1/8) second. The last three images (d-f) were exposed for one-sixteenth (1/16), one-thirtieth (1/30), and one-sixtyfourth (1/64) second, respectively. The image having the best overall exposure, color contrast, and saturation is image (d), which is positioned near the center of the bracket.
Interactive Java Tutorial
Live Settings Panel
Explore the various features available in the DXM 1200 ACT-1 control software Live settings panel, including exposure time, focus, and color levels.
The ACT-1 exposure control panel is presented in Figure 2. The upper portion of the figure illustrates how the control panel appears when exposure times range between 1/12000 and 1 second (Figure 2(a)). The slider is used to adjust the exposure time in discrete increments by dragging with the cursor. As the exposure time is adjusted by translating the slider, the Exposure Time window changes to indicate the reciprocal of the selected time. For example, the number "15" appearing as a simulated LED readout in the control panel window (Figure 2(a)) indicates an exposure time of 1/15 second. If the exposure were 1/12000 second (the fastest exposure), the numerals 12000 would appear in the control panel window.
For longer exposures (exceeding one minute), the slider should be dragged to the rightmost position and a specific time entered into the text box (Figure 2(b)). When the slider reaches the right-hand position, the text box becomes active and the time (in minutes) is displayed in the Exposure Time window. The user can specify an exposure time either by typing a time value in the text box or by clicking on the up/down arrow buttons. The buttons increase or decrease exposure time in discrete increments, but any time value between 2 and 170 seconds (integers only) can be specified by typing that value into the text box.
DXM 1200 Digital Camera Fine Mode
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Table 1
Also appearing in the control panel is the Exposure Mode indicator, which can be toggled between Fine (high resolution images) and Quick (low resolution images) modes. The Fine mode is utilized to capture images up to 12 million pixels in size, while the Quick mode is used for images ranging from 0.3 to 1.3 million pixels (see Tables 1 and 2).
When the DXM 1200 is in Fine mode, the inter-pixel stepping mechanism is activated and the CCD is stepped horizontally and vertically in extremely small increments to capture an image at each position. The final image is a composite of the images gathered at each step and tiled together. Alternatively, in the Quick mode, inter-pixel stepping is disabled and the image is captured in a single step by the CCD, resulting in smaller images having lower resolution. It should be noted that operating the DXM 1200 in Fine mode takes much longer to integrate an image than when the camera is utilized in the Quick mode. It may also be more difficult to image moving specimens or capture rapid events in the higher resolution mode.
DXM 1200 Digital Camera Quick Mode
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Table 2
Images produced by the DXM 1200 in Fine mode can have file sizes exceeding 35 megabytes (Mb), and can quickly fill up a computer hard drive. In cases where capturing images quickly has a higher priority than image quality, the user should select Quick mode. In this mode, the camera operates approximately nine times faster to capture an image than in Fine mode.
Image size options (Tables 1 and 2) are available through a pull-down menu in the Photo control panel (see Figure 3). To access the menu, click on the Photo panel tab in the Settings panel window. This panel allows the user to choose between Fine and Quick photomicrography modes and to simultaneously select the final image resolution size in pixels. In order to select a final image size, first choose the desired photomicrography mode, then click on the down-arrow to activate the pull-down menu (displaying the palette of image sizes) and choose the appropriate pixel size. Another option accessable through this control panel is the Auto Save feature and a file format option for saving image data.
The live image window (Figure 4) contains an exposure frame that outlines the area to be captured in the final digital image. The size of this frame corresponds to the selected image size (chosen in the Photo panel). When an image size is chosen that is smaller than full screen size, only the portion of the screen bounded by the exposure frame is included in the image. The exposure frame can be translated within the boundaries of the live image window by clicking anywhere within the frame and dragging with the mouse. In this manner, users can select a specific portion of the window for photomicrography.
The center of the live image window also contains a digital Focus Mark that denotes the area selected for focus (Figure 4). A set of radio buttons is utilized to toggle the Focus Mark icon on and off and to choose whether it is Docked (positioned and locked into the center of the window and focus frame), or mobile. Unchecking the Docked radio button will allow independent movement of the Focus Mark, a feature discussed more thoroughly in the section describing Focus Control with the DXM 1200 camera system.
Camera sensitivity (gain) can be adjusted through a set of radio buttons located in the Photo control panel (Figure 5). There are three options in the sensitivity window, Normal, High, and Max, which can be utilized to control the exposure time. Increasing the sensitivity by checking the High or Max radio buttons will shorten exposure time at the expense of increased noise. Avoid exceeding long exposure times (several tens of seconds), which can cause the image to be degraded by excess camera noise. Instead, adjust the microscope illumination intensity so that shorter exposure times can be comfortably utilized.
