SPOT 4.1 Basic and Advanced Software Manual for SPOT Insight ...

Ch. 5 – Preparing to Take a PictureMinimum Exposure of 25 ms ensures that the camera’s exposure time is greater than the pulse interval ofthe light source and that any error is minimized. The greater the value above 25 ms, the more the error isreduced. Alternatively, you can enter any multiple of 8.333 ms. To enter a minimum exposure, check theMinimum box, and enter a value in milliseconds.White BalanceThe exposure window also indicates the white balance settings currently associated with the open imageset-up. These values from Red, Green and Blue can be modified by using the increase/decrease arrow keysnext to the value or by directly typing in the new value.White balance is the ratio of red, green, and blue necessary to achieve the proper color rendition for animage. Many factors impact the coloration of an image capture, including lamp voltage, the coloration ofthe glass used in the slide, and the coloration of the glass used in the lenses of the microscope’s objectives.To minimize the impact of these factors, use the Compute White Balance feature. When you perform awhite balance calculation, SPOT samples the color of the light from your light source after it has passedthrough the slide and the objective, and then calculates the exposure values needed to produce imageswhere the white areas are white and the color areas are true.You should perform a new white balance calculation when:• Starting a new image capture session.• Changing the lamp voltage.• Switching to a different objective on the microscope.When you open an image setup, the Image Setup screen displays the white balance values that were lastsaved to the Image Setup. If you capture an image without computing a new white balance, SPOT usesthese default values.To compute new white balance values for an image setup, follow these steps:1. Set the illumination and the objective that you will use for the image capture.2. Show the camera a sample of white light. This step varies according to microscopy technique andsample type, as shown in the table below:Image TypeBrightfield-transmittedlightDarkfield-transmittedlightFluorescenceNomarski DIC/HoffmanPhase contrastPolarized lightBrightfield-reflected lightWhite Balance TechniquePosition the slide so that the specimen is not in the field of view, but light is going throughthe slide near the specimen.Perform the white balance using the brightfield mode on your microscope. See Brightfield–transmitted light above.Use the Match Color feature. Refer to Ch. 8 – Viewing, Editing, and Annotating Images formore information.For Nomarski DIC: Adjust the Nomarski prism until the image is in either the “brightfield” orthe “gray scale” view.For Hoffman: Perform the white balance directly on the Hoffman image.Position the slide so that the specimen is not in the field of view, but light is going throughthe slide near the specimen.Pull all polarizers and compensators out of the light path and then position the slide so thatthe specimen is not in the field of view, but light is going through the slide near thespecimen.Replace your sample with a mirror.Darkfield-reflected light Perform the white balance using the brightfield mode on your microscope. See Brightfield –reflected light above.Stereo/MacroUsing an EPI-illuminator (shining a light on the top of your specimen from off to one side),replace your sample with a white sheet of paper.98 User Guide to the SPOT Insight Camera