Abstract

In applications where random multi-photon events must be distinguishable from the background, detection of the signals must be based on either analog current measurement or photon counting and multi-level discrimination of single and multi-photon events. In this paper a novel method for optimizing photomultiplier (PMT) pulse discrimination levels in single-and multi-photon counting is demonstrated. This calibration method is based on detection of photon events in coincidence to short laser pulses. The procedure takes advantage of Poisson statistics of single- and multi-photon signals and it is applicable to automatic calibration of photon counting devices on production line. Results obtained with a channel photomultiplier (CPM) are shown. By use of three parallel discriminators and setting the discriminator levels according to the described method resulted in a linear response over wide range of random single- and multi-photon signals.

© 2004 Optical Society of America

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References

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  1. R. J. Ellis and A. G. Wright: �??Optimal use of photomultipliers for chemiluminescence and bioluminescence applications,�?? Luminescence 14, 11-18 (1999).
    [CrossRef] [PubMed]
  2. J. T. Soini, J. M. Soukka, E. Soini, and P. E. Hänninen, �??Two-photon excitation microfluorometer for multiplexed single-step bioaffinity assays,�?? Rev. Sci. Instrum. 73, 2680-2685 (2002).
    [CrossRef]
  3. P. Hänninen, A. Soini, N. Meltola, J. Soini, J. Soukka, and E. Soini, �??A new microvolume technique for bioaffinity assays using two-photon excitation,�?? Nature Biotechnol. 18, 548-550 (2000).
    [CrossRef]
  4. J. T. Soini, J. M. Soukka, N. J. Meltola, A. E. Soini, E. Soini, and P. E. Hänninen, �??Ultra Sensitive Bioaffinity Assay for Micro Volumes,�?? Single Molecules 1, 203-206 (2000).
    [CrossRef]
  5. PerkinElmer Optoelectronics data sheet, �??Channel Photomultipliers, Overview and Specifications�?? (PerkinElmer Optoelectronics, 2001), <a href="http://optoelectronics.perkinelmer.com/content/RelatedLinks/cpm_brochure.pdf">http://optoelectronics.perkinelmer.com/content/RelatedLinks/cpm_brochure.pdf</a>.

Luminescence (1)

R. J. Ellis and A. G. Wright: �??Optimal use of photomultipliers for chemiluminescence and bioluminescence applications,�?? Luminescence 14, 11-18 (1999).
[CrossRef] [PubMed]

Nature Biotechnol. (1)

P. Hänninen, A. Soini, N. Meltola, J. Soini, J. Soukka, and E. Soini, �??A new microvolume technique for bioaffinity assays using two-photon excitation,�?? Nature Biotechnol. 18, 548-550 (2000).
[CrossRef]

Rev. Sci. Instrum. (1)

J. T. Soini, J. M. Soukka, E. Soini, and P. E. Hänninen, �??Two-photon excitation microfluorometer for multiplexed single-step bioaffinity assays,�?? Rev. Sci. Instrum. 73, 2680-2685 (2002).
[CrossRef]

Single Molecules (1)

J. T. Soini, J. M. Soukka, N. J. Meltola, A. E. Soini, E. Soini, and P. E. Hänninen, �??Ultra Sensitive Bioaffinity Assay for Micro Volumes,�?? Single Molecules 1, 203-206 (2000).
[CrossRef]

Other (1)

PerkinElmer Optoelectronics data sheet, �??Channel Photomultipliers, Overview and Specifications�?? (PerkinElmer Optoelectronics, 2001), <a href="http://optoelectronics.perkinelmer.com/content/RelatedLinks/cpm_brochure.pdf">http://optoelectronics.perkinelmer.com/content/RelatedLinks/cpm_brochure.pdf</a>.

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Figures (4)

Fig. 1.
Fig. 1.

Block diagram of the measurement set-up optical components.

Fig. 2.
Fig. 2.

Block diagram of the measurement electronics set-up.

Fig. 3.
Fig. 3.

Measured photoelectron distribution from different concentrations of Rhodamine B solution.

Fig. 4.
Fig. 4.

Measured fluorescence photon count rates (CPS) from Rhodamine B solutions. The signal has been measured using a single comparator (sphere) and three comparators (square).

Equations (8)

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P ( n ) = μ n e μ n !
P ( 0 ) = 1 N C 1 N L
μ = ln ( 1 N C 1 N L )
N ̂ C 2 = N L [ P ( 2 ) + P ( 3 ) + ] = N L [ 1 P ( 0 ) P ( 1 ) ] = N L [ N C 1 N L P ( 1 ) ] = N C 1 N L P ( 1 )
N ̂ C 3 = N L [ P ( 3 ) + P ( 4 ) + ] = N L [ 1 P ( 0 ) P ( 1 ) P ( 2 ) ] = N C 1 N L [ P ( 1 ) + P ( 2 ) ]
N ̂ CM = N C 1 N L i = 1 M 1 P ( i )
N tot = N C 1 + N C 2 + N C 3 N L μ

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