Chlorophyll a fluorometry
The success of chlorophyll a fluorometry is due, in large part, to two important practical developments; the 'light addition' method (Bradbury and Baker, 1984; Quick and Horton, 1984), which uses a multiple-turnover pulse to achieve the Fm and Fm' fluorescence levels, and the application of either locked-in amplification (Quick and Horton, 1984) or Pulse Amplitude Modulation (PAM) (Schreiber et al., 1989), both of which generate a signal that is proportional to fluorescence yield, with a high signal to noise ratio. In combination, these developments have made it possible to quantify the relative contributions of photochemical and non-photochemical quenching processes to changes in fluorescence yield, with minimal impact on the photosynthetic apparatus.
A number of theoretical advances and empirical observations have also played critical roles in the development of chlorophyll a fluorometry. Perhaps the most useful, and certainly the most widely appreciated, is the demonstration by Genty et al. (1989), that Fq'/Fm' frequently correlates with the quantum yield of CO2 assimilation in the absence of photorespiration. As noted by Lavergne and Trissl (1995), this relationship is most easily explained if the process of downregulation (which results in an increase in non-photochemical quenching of chlorophyll a fluorescence) is assumed to operate in the same way as a Stern-Volmer quencher; a point that has important implications for the interpretation of chlorophyll a fluorescence data.
There are two other factors that make chlorophyll a fluorometry a viable technique. Firstly, the yield of chlorophyll a fluorescence (greater than 1% of the photons absorbed by the photosynthetic apparatus as a whole). Secondly, the broad action spectra for photosynthesis within oxygenic organisms (which typically extend from 400 nm to 700 nm) make it easy to avoid overlap between the measuring light and fluorescence signal (which has a peak between 682 and 685 nm and a shoulder that extends to 740 nm).
Links and references
Fo' calculation method
The value of Fo' is required in the calculation of Fv'/Fm' and Fq'/Fv'. Fo' has frequently been measured during exposure of the sample to weak far-red light in the absence of actinic light (van Kooten and Snel, 1990; Maxwell and Johnson, 2000). It is assumed that, under these conditions, QA is maximally oxidized through preferential excitement of PS I (relative to PS II) during the exposure to far-red light. There are at least two potential sources of error with this method. Firstly, it is very likely that non-photochemical quenching will partly reverse during the far-red light treatment. Secondly, and more importantly, this method relies on the near-complete oxidation of QA (which, in turn, relies upon oxidation of the plastoquinone pool) within a relatively short time (typically ca. 3 s) in the presence of a pmf across the thylakoid membrane. It is almost certainly the case that, under these conditions, the oxidation of plastoquinone at the QO site of the cytochrome b6f complex imposes a much greater limitation to non-cyclic electron flow than does the rate of excitation of PS I (Bendall, 1982). Consequently, it is unlikely that the preferential excitation of PS I by far-red light treatment will have the desired effect and that the level of Fo' will be overestimated, particularly at high PPFD. An alternative method for estimating the value of Fo' has been developed, originally for use with a high-resolution, chlorophyll fluorescence imaging systems (Oxborough and Baker, 1997). With this method, Fo' is calculated using the value of Fm' at the point of measurement and dark-adapted values of Fo and Fm measured before or (more usually) after the measurement of Fm':
Links and references
Oxborough (2004)
Oxborough & Baker (1997)
How a laser works
Links and references
There are lots of explanations available on the web. The links below are excellent starting points.
Wikepedia - Lasers
Laser optics UK - Laser
tutorial
How LIBS works
Links and references
Aguilera et al. (2003) Spatial characterization of laser-induced plasmas by deconvolution of spatially resolved spectra
Bulajic et al. A new self-calibrated method for precise quantitative analysis by laser induced breakdown spectroscopy
Boyain-Goitia et al. (2003) Single pollen analysis by laser-induced breakdown spectroscopy and Raman microscopy
Carranza et al. (2003) Comparison of non intensified and intensified CCD detectors for laser-induced breakdown spectroscopy
Carranza et al. (2003B) Conditional data processing for single-shot spectral analysis by use of laser-induced breakdown spectroscopy
Corsi et al. (2003) Application of laser-induced breakdown spectroscopy technique to hair tissue mineral analysis
Detalle et al. (2003) Influence of Er:YAG and Nd:YAG wavelength on laser-induced breakdown spectroscopy measurements under air or helium atmosphere
Hahn et al. (2003) Laser-induced breakdown spectroscopy: an introduction to the feature issue
Morel et al. (2003) Detection of bacteria by time-resolved, laser-induced breakdown spectroscopy
Martin et al. (2003) Laser-induced breakdown spectroscopy for the environmental determination of total carbon and nitrogen in soils
Portnov et al. (2003) Emission following laser-induced breakdown spectroscopy of organic compounds in ambient air
Sabsabi et al. (2003) Comparative study of two new commercial echelle spectrometers equipped with intensified CCD for analysis of laser induced breakdown spectroscopy
Samuels et al. (2003) Laser induced breakdown spectroscopy of bacterial spores, molds, pollens, and protein: initial studies of discrimination potential
Sturm & Noll (2003) Laser-induced breakdown spectroscopy of gas mixtures of air, CO2, N2 and C3H8 for simultaneous C, H, O, and N measurement
Cremers et al. (2001) Measuring total soil carbon with Laser-Induced Breakdown Spectroscopy (LIBS)
Hahn & Lunden (2000) Detection and analysis of aerosol particles by laser-induced breakdown spectroscopy