20 elements in biological samples, the most common being calcium, magnesium and
manganese. Absorption spectrophotometers are usually more sensitive than emission
instruments and can detect less than 1 p.p.m. of each of the common elements with
the exception of alkali metals. The relative precision is about 1% in a working range of
20–200 times the detection limit of an element.
AES and AAS have been widely used in analytical chemistry, such as environ-
mental and clinical laboratories. Nowadays, the technique has been superseded largely
by the use of ion-selective electrodes (see Section 16.2.2).
Atomic fluorescence spectrophotometry
Despite being limited to only a few metals, the main importance ofatomic fluores-
cence spectrophotometry(AFS) lies in the extreme sensitivity. For example, zinc and
cadmium can be detected at levels as low as 1–2 parts per 10^10.
AFS uses the same basic setup as AES and AAS. The atoms are required to be vaporised
by one of three methods (flame, electric, ICP). The atoms are excited using electromagnetic
radiation by directing a light beam into thevaporised sample. This beam must be intense,
but not spectrally pure, since only the resonantwavelengths will be absorbed, leading to
fluorescence (see Section 12.3.1).
12.8 Suggestions for further reading
General biophysics
Hoppe, W., Lohmann, W., Markl, H. and Ziegler, H. (1982).Biophysik, 2nd edn. Berlin:
Springer-Verlag. (A rich and authorative compendium of the physical basics of the life sciences.)
WEBSITES
http://lectureonline.cl.msu.edu/%7Emmp/applist/Spectrum/s.htm
http://www.colorado.edu/physics/2000/lasers/index.html
Ultraviolet and visible light spectroscopy
Simonian, M. H. and Smith, J. A. (2006). Spectrophotometric and colorimetric determination of
protein concentration.Current Protocols in Molecular Biology, Chapter 10, Unit 10.1A.
New York: Wiley Interscience
WEBSITES
http://teaching.shu.ac.uk/hwb/chemistry/tutorials/molspec/uvvisab1.htm
http://www.cem.msu.edu/~reusch/VirtualText/Spectrpy/UV-Vis/spectrum.htm#uv1
http://www.srs.dl.ac.uk/VUV/
http://phys.educ.ksu.edu/vqm/html/absorption.html
Fluorescence spectroscopy
Brown, M. P. and Royer, C. (1997). Fluorescence spectroscopy as a tool to investigate protein
interactions.Current Opinion in Biotechnology, 8 , 45–49.
Groemping, Y. and Hellmann, N. (2005). Spectroscopic methods for the determination of protein
interactions.Current Protocols in Protein Science, Chapter 20, Unit 20.8. New York: Wiley
Interscience.
Hwang, L. C. and Wohland, T. (2007). Recent advances in fluorescence cross-correlation
spectroscopy.Cell Biochemistry and Biophysics, 49 , 1–13.
519 12.8 Suggestions for further reading
