b'Chemiluminescence and fluorescence detection data captureWith chemiluminescence-based detection methods, theAlthough X-ray film is inexpensive, consider the expenses chemical reaction (HRP- or AP-driven) produces light thatof maintaining a darkroom and the associated developing can be captured with X-ray film or a capable western blotequipment. Modern western blot imaging instruments may imaging instrument. When capturing chemiluminescentcost more up-front than the typical expense of X-ray film, signal with X-ray film, it is necessary to expose severalbut these instruments can often capture signal for multiple pieces of film for different time periods in an attempt toapplications (such as colorimetric protein gel stains) in obtain the proper balance between signal and background.addition to chemiluminescent western blot signals, which The goal is to time the exposure so that the desiredadds value to the overall investment.signal is clearly visible while the background remains low. This is difficult to accomplish since the process cannotSeveral manufacturers offer fluorescence western blot be observed and stopped when the desired endpointimaging systems capable of capturing fluorescence is reached. signals, most of which use either filter-based or laser-based technologies to deliver the appropriate excitation If the film is underexposed, the signal will not be visible.wavelength and then record the emission light output. If the film is overexposed, the signal may be saturated,The majority of the available instrumentation is CCD lost in the background, or once-distinguishable bandscamerabased, but scanning systems with photoavalanche may become blurred together. All situations make post- diodebased sensors are also available. Captured images signal capture data analysis challenging. Although X-rayare saved digitally. The number of commercially available film provides qualitative and semi-quantitative data andfluorescently labeled antibodies with different excitation and is useful to confirm the presence of target proteins,emission spectra continues to increase, which provides purpose-built western blot imaging instruments canmultiple options to the end user based on preference or the offer higher sensitivity and broader dynamic range thanselected instruments imaging capability.film, which makes it possible to detect more subtle differences between samples. Furthermore, you dont have to spend time in the darkroom exposing your blot to film and developing the film thereafter. With western blot imaging instruments, the data are captured digitally, which negates the need to scan X-ray film to perform post-capture analysis. Furthermore, many of these systems offer auto-exposure algorithms to help best optimize signal-to-noise ratio.62'