The combiner module consists of two identical circuits, each of
which can combine up to four inputs to produce a single (out)put. The signals
to be combined will normally (but not necessarily!) be from two or more signals
of the same wavelength in the rotor, and they will normally come directly from
the outputs of the output module. The combination can be done in two different
ways, to give either improved time resolution or improved signal-to-noise
ratio. For the majority of applications the combiner is likely to be used to
improve the signal-to-noise ratio, but the possibility of improving the time
resolution may also be useful, and in any event an appreciation of this mode of
operation will be of assistance in setting up the module.
As with our other modules, the operation of the combiner is programmed via jumper links. This module is the current record holder for the largest number of such links on any of our boards, but once its basic operation is understood, the setting up of the jumper links is relatively straightforward.
The combiner works by averaging together up to four signals in each circuit, but the individual signals do not NECESSARILY contribute to the average all the time. Instead, each signal can contribute to the output for a fraction of the total revolution period of the rotor, with the fraction size being determined by the total number of filter positions in the rotor (i.e. normally six, but up to eight). This allows the combined output to be programmed so that it carries the MOST RECENTLY ACQUIRED signal for each filter wavelength.
For example, consider a situation in which filter positions 1 3 and 5 are occupied by filters of centre wavelength A, and filter positions 2 4 and 6 are occupied by filters of centre wavelength B. The combiner can be programmed so that the output of one circuit carries the signal from filter 1 as soon as it is acquired (i.e. as soon as its associated sample-and-hold amplifier in the output module has sampled the filter signal), with none of the other filter signals contributing to the output. As soon as the signal from filter position 3 has been acquired, the signal from filter 1 is turned off and the signal from filter 3 is turned on. Similarly, as soon as the signal from filter position 5 has been acquired, the signal from filter 3 is turned off and the signal from filter 5 is turned on. The other combiner circuit can be programmed to perform the corresponding operation for filters 2 4 and 6. Further details of the required jumper settings are given subsequently.
When the combiner is used in this way, it is important that the filters to be combined give exactly equal signals, otherwise the switching action will cause unwanted changes in the output. Since it is unlikely that the signals will be exactly equal in practice, they must be made so by an electrical adjustment. For this purpose, the combiner has a multiturn preset potentiometer for each of the signals to be combined. Presets P1-P4 provide variable attenuation of signals 1-4 for combiner circuit 1, and P5-P8 serve the corresponding functions for combiner circuit 2. The presets are numbered starting from the top of the module. There is no need to alter these presets if you do not require this facility, and the combiner is supplied with all the presets at their maximum gain settings. Please note that the combiner has a unity gain, and the presets serve only to attenuate the individual signals, so they should be used (if required) to reduce the gains of the larger signals to match the smallest of the signals to be combined. If you wish to AMPLIFY signals from individual filters, then you should use the input gain balancer module (which operates before the output module) for this purpose. When that module is in use, it may be more convenient in any case to balance the signals there rather than using the combiner module presets. PLEASE NOTE THAT THERE IS NO NEED TO ADJUST THE INDIVIDUAL SIGNAL AMPLITUDES WHEN THE COMBINER IS USED AS A CONTINUOUS AVERAGER AS DESCRIBED BELOW.
Using the combiner as described above allows a signal sampling rate that is several times faster than the rotor speed, which may be useful in some applications. However, there is inevitably a tradeoff between bandwidth and noise, and for most fluorescence applications reduction of noise rather than increase in bandwidth is desirable. In this case, the signal from each filter of the same wavelength contributes to the output continuously. The output is the average (not the sum) of the individual inputs.
The filter positions for which each signal is sent to the combiner is set by the jumper banks along the top of the board. Switching of signals 1-4 for combiner 1 is by the four banks nearer the front of the board, and control of signals 1-4 for combiner 2 is by the four banks nearer the back. Each bank has eight jumpers, i.e. one for each of the up to eight filter positions, with filter 1 at the bottom. To switch on the combiner for the appropriate signal and filter position, the appropriate jumper should be in the right-hand position, i.e. connecting the pair of pins which are closer to the back of the board, as marked on the board itself. When signals are continuously averaged, each bank should have ALL its jumpers in either the off or on positions as appropriate. Jumpers for filter positions which are not present in the rotor (i.e. positions 7 and 8 in a six-filter rotor) have no effect, but for clarity of operation it is recommended that they are also set as appropriate.
The filter signals which correspond to the four input signals for each combiner are determined by the jumper banks which are nearer the bottom of the board. Each bank is labelled 1-4 to correspond with the switch selector banks above them. The individual jumper positions are all labelled, but to summarise, each signal can be any of the following; either the A or B outputs for filter positions 1-8 (i.e. the signals normally supplied by the output amplifiers), the output from either of two ratio amplifiers (again A and B) which are labelled as "OUT" on the board, or the direct outputs from any of the four input amplifiers A-D, which are labelled on the board as "IN" for A (left) and B (right) and C/D for C (left) and D (right). Normally only the A and B positions for filters 1-8 would be used as inputs. To select an input, one jumper is used to connect the appropriate signal pin to one of the central row of pins.
The output from each combiner circuit is available on a BNC socket on the front panel, and it can also be sent to one or more other system modules via unused signal lines on the system backplane. The signal line to be used is set by the jumper bank labelled "OUT", which is to the right of each of the two sets of signal selection jumper banks. The choice of signal lines is exactly as described above for them, but it is important that no other module uses the chosen line as an output. Since the standard rotor in our system has six filter positions, the signal lines for positions 7 and 8 are normally spare. We therefore suggest that the outputs from combiner circuits 1 and 2 are connected to signal lines 7A and 8A respectively. However, you should check that the sample-and-hold amplifiers for positions 7 and 8 in the output module have their outputs disconnected from the backplane, i.e. REMOVE the output jumpers for these filter positions.
When using the combiner as a continuous averager, there is no need to fit the filters in the rotor in any particular order. However, it will probably be most convenient to have filters of the same wavelength in adjacent positions. Conversely, when using the combiner to increase the effective sampling rate, it will of course be necessary to distribute the filters evenly around the rotor, as in the example described above.
When selecting the switching, please note that the combiner operates so as to make this as straightforward as possible. Thus, to get the latest signal value for filter 1, you should simply arrange for the corresponding combiner input to be switched on beginning from filter position 1. For the alternating wavelength A/wavelength B filter sequence in the previous example, then in order to derive a combiner output which consists of the most recently acquired wavelength A signal (where filters of wavelength A are on positions 1, 3 and 5), the following configuration is required for one of the combiner circuits. The output from filter 1 should be selected for filter positions 1 and 2, the output from filter 3 should be selected for filter positions 3 and 4, and the output from filter 5 should be selected for filter positions 5 and 6. The other circuit can be configured in an analogous fashion for wavelength B.
For fluorescence applications, a particular advantage of the combiner is that it allows the RELATIVE numbers of filters at the various excitation wavelengths to be optimised. For ratiometric measurements, the signal-to-noise ratio will be determined primarily by the weaker (i.e. noisier) of the two signals, so it is desirable to balance the optical excitation so that they are comparable. Since in many circumstances one of the two excitation wavelengths may give significantly weaker fluorescence than the other, it is desirable for best overall signal-to-noise ratio to illuminate at this wavelength for a relatively longer period. Use of a continuously-spinning rotor with differing numbers of the two excitation filter wavelengths (e.g. 4:2) in conjunction with the combiner is a very elegant way of achieving this, as it gives a much higher sampling speed and much less mechanical vibration than the alternative solution of a discontinuous filter changer with different dwell times in the two positions.
