LMCB SP5 Hybrid Detectors

Owned by Andrew Vaughan
Last updated: Jul 04, 2014Version comment

Using the Hybrid Detectors

On 3rd July 2014 three of the SP5's four photomultiplier tubes (PMTs) were replaced by new HyD hybrid detectors. The HyD detectors are in positions 1, 3 and 4. Position 2 is still occupied by a PMT because of a design limitation of the scan head.

The HyD controls are a little different to those for PMTs. If you hover the mouse over the detector in the Beam Path view a set of three radio buttons will appear (see Figure 1, right). Of these Standard mode is most similar to normal PMT operation and is likely to be most popular. The gain in standard mode is adjustable between 10% and 500% (see Figure 2, right). BrightR is a high dynamic range mode that allows you to image bright and dim areas in the same image without experiencing saturation but it should not be used if quantification is required since it changes the intensities in a non-linear manner. The most quantitative mode is Photon Counting, where each intensity value represents a photo-electron. You may find the image appears very dim since there are actually very few photo-electrons detected in confocal microscopy because of the low fluorescent signals and optical sectioning. There is also no gain, since the detector is actually counting the photo-electron signal and not amplifying it. You can use the auto-contrast feature in the left image tool bar (labelled A) to auto range the image, but this only refreshes after a complete image is acquired.

How to avoid saturating the detectors (or what to do if you see nothing when you scan)

The HyD detectors are extremely sensitive, which means that it is very possible that illuminating specimens with the laser powers you are used to using with the PMTs will saturate them, causing them to shut down so they aren't damaged. If you regularly load saved settings from the instrument parameters or from saved data these will be suitable for an all PMT system but may well saturate the HyDs. You should try the following steps before starting your scan to avoid inadvertently saturating the detectors:

  1. Set the laser powers to approximately 10 - 30% of what you would normally use
  2. Set the HyD gain to its lowest value of 10% (make sure it genuinely is 10% - occasionally the slider jumped back up to about 100% when I was doing this)
  3. Adjust the laser power and gain settings carefully to optimise the brightness
  4. This does not count for the detector in position 2, which is still a PMT and works in exactly the same way as before

If a HyD does shut down you won't see anything on the screen except if you have the Glow Over/Under look-up table applied. In the latter case the image will be totally green because every value will be zero. To reactivate the detector you can simply uncheck and check the Active check box in the detector controls (Figure 1)

Interference - switch mobile phones off

You must switch off mobile phones, put them into flight mode or otherwise make them incapable of sending or receiving signals when in room 1.20B with the SP5. This is because the HyD detectors pick up mobile phone signals as interference lines on the scan. There is a sticker on the door to remind you of this.