Scientifica MDU - Excessive noise during imaging at high detector gain (voltages between -800 V and -1100 V); Variable noise and offset down an image.

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from stray light ingress. No stray light can enter the optics block other than via the
objective port or the laser port.
o Check that any substage transmission-type illumination source is turned off during
imaging even if it is an infrared source (some of which may emit some visible light).
o Stray light is most likely to be coming from visible light which has been scattered off the
sample and into the objective lens. Check that the experimental arrangement provides
fully dark conditions at the sample during imaging.
o It is possible for visible light to enter the optics block from the top (i.e. from the direction
of the scanner). Again, check that there is no stray source of light in that part of the
equipment (look for a red emission indicator light on the laser for example, and cover it).
o If you are using a MOC with your detection module, check that the DIC prism cover is
properly fitted to the MOC top plate .
Excessive noise during imaging at high detector gain (voltages between -800V and -1100V)
 Probable cause: detector noise and dark current or recent detector overload.
o At high gain, some degree of dark current is to be expected: it will often take the form of
randomly-timed short impulses clearly visible on the oscilloscope. These may be more
frequent on red-sensitive PMTs than on blue-green sensitive PMTs and more frequent
on warm days.
o If the noise level is excessive, or significantly higher than usual, and stray light ingress
can be ruled out with certainty, then it is possible that one of the detectors has
accidentally been exposed to an overload or to an overly bright light when operating at
high gain.
o Recovery from such exposure can take several hours so our recommendation is to
leave the detector switched off in the dark for 12 hours before testing again if accidental
exposure to bright light is suspected.
o If, after a period in the dark, the detector noise has not reduced to previous levels, the
detector may need to be replaced. Contact Scientifica for advice.
 Variable noise and offset down an image.
 Probable cause: visible (especially deep red) light entering the system along the laser beam
o Some lasers emit a significant “deep red” component as well as the intended infra-red
beam, and this can enter the system. If the noise increases from the top to the bottom
of the image, then this stray light is probably beginning to reflect off the input dichroic
mirror with variable reflectivity as the beam is scanned vertically. Try adding a long-
pass filter to the system before the scanner optics but after any shutter. Suitable filters
include:
 Semrock LP02780RS or LP02-780RU
 Semrock LP02-664RS or LP02-664RU