Skip to main content Accessibility help
×
Hostname: page-component-8448b6f56d-tj2md Total loading time: 0 Render date: 2024-04-17T22:25:36.385Z Has data issue: false hasContentIssue false
This chapter is part of a book that is no longer available to purchase from Cambridge Core

12 - Spectroscopic techniques: I Spectrophotometric techniques

Keith Wilson
Affiliation:
University of Hertfordshire
John Walker
Affiliation:
University of Hertfordshire
Get access

Summary

INTRODUCTION

Spectroscopic techniques employ light to interact with matter and thus probe certain features of a sample to learn about its consistency or structure. Light is electromagnetic radiation, a phenomenon exhibiting different energies, and dependent on that energy, different molecular features can be probed. The basic principles of interaction of electromagnetic radiation with matter are treated in this chapter. There is no obvious logical dividing point to split the applications of electromagnetic radiation into parts treated separately. The justification for the split presented in this text is purely pragmatic and based on ‘common practice’. The applications considered in this chapter use visible or UV light to probe consistency and conformational structure of biological molecules. Usually, these methods are the first analytical procedures used by a biochemical scientist. The applications covered in Chapter 13 present a higher level of complexity in undertaking and are employed at a later stage in biochemical or biophysical characterisation.

An understanding of the properties of electromagnetic radiation and its interaction with matter leads to an appreciation of the variety of types of spectra and, consequently, different spectroscopic techniques and their applications to the solution of biological problems.

Properties of electromagnetic radiation

The interaction of electromagnetic radiation with matter is a quantum phenomenon and dependent upon both the properties of the radiation and the appropriate structural parts of the samples involved. This is not surprising, since the origin of electromagnetic radiation is due to energy changes within matter itself.

Type
Chapter
Information
Publisher: Cambridge University Press
Print publication year: 2010

Access options

Get access to the full version of this content by using one of the access options below. (Log in options will check for institutional or personal access. Content may require purchase if you do not have access.)

References

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.)CrossRefGoogle Scholar
http://lectureonline.cl.msu.edu/%7Emmp/applist/Spectrum/s.htm
http://www.colorado.edu/physics/2000/lasers/index.html
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 InterscienceCrossRef
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
Brown, M. P. and Royer, C. (1997). Fluorescence spectroscopy as a tool to investigate protein interactions. Current Opinion in Biotechnology, 8, 45–49.CrossRefGoogle ScholarPubMed
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.CrossRefGoogle ScholarPubMed
Lakowicz, J. R. (1999). Principles of Fluorescence Spectroscopy 2nd edn. New York: Kluwer/ Plenum. (An authorative textbook on fluorescence spectroscopy.)CrossRefGoogle Scholar
Langowski, J. (2008). Protein–protein interactions determined by fluorescence correlation spectroscopy. Methods in Cell Biology, 85, 471–484.CrossRefGoogle ScholarPubMed
Prinz, A., Reither, G., Diskar, M. and Schultz, C. (2008). Fluorescence and bioluminescence procedures for functional proteomics. Proteomics, 8, 1179–1196.CrossRefGoogle ScholarPubMed
Roy, R., Hohng, S. and Ha, T. (2008). A practical guide to single-molecule FRET. Nature Methods, 5, 507–516.CrossRefGoogle ScholarPubMed
VanEngelenburg, S. B. and Palmer, A. E. (2008). Fluorescent biosensors of protein function. Current Opinion in Chemical Biology, 12, 60–65.CrossRefGoogle ScholarPubMed
http://www.invitrogen.com/site/us/en/home/support/Tutorials.html
http://www.microscopyu.com/tutorials/java/fluorescence/excitationbalancer/index.html
Bacart, J., Corbel, C., Jockers, R., Bach, S. and Couturier, C. (2008). The BRET technology and its application to screening assays. Biotechnology Journal, 3, 311–324.CrossRefGoogle ScholarPubMed
Deshpande, S. S. (2001). Principles and applications of luminescence spectroscopy. Critical Reviews in Food Science Nutrition, 41, 155–224.CrossRefGoogle ScholarPubMed
Jia, Y., Quinn, C. M., Kwak, S. and Talanian, R. V. (2008). Current in vitro kinase assay technologies: the quest for a universal format. Current Drug Discovery Technology, 5, 59–69.CrossRefGoogle ScholarPubMed
Meaney, M. S. and McGuffin, V. L. (2008). Luminescence-based methods for sensing and detection of explosives. Analytical and Bioanalytical Chemistry, 391, 2557–2576.CrossRefGoogle Scholar
http://www.turnerbiosystems.com/doc/appnotes/998_2620.html
Fasman, G. D. (1996). Circular Dichroism and the Conformational Analysis of Biomolecules. New York: Plenum Press. (An authorative textbook on circular dichroism in biochemistry.)CrossRefGoogle Scholar
Gottarelli, G., Lena, S., Masiero, S., Pieraccini, S. and Spada, G. P. (2008). The use of circular dichroism spectroscopy for studying the chiral molecular self-assembly: an overview. Chirality, 20, 471–485.CrossRefGoogle ScholarPubMed
Kelly, S. M. and Price, N. C. (2006). Circular dichroism to study protein interactions. Current Protocols in Protein Science, Chapter 20, Unit 20.10. New York: Wiley Interscience.
Martin, S. R. and Schilstra, M. J. (2008). Circular dichroism and its application to the study of biomolecules. Methods in Cell Biology, 84, 263–293.CrossRefGoogle Scholar
http://www.cryst.bbk.ac.uk/cdweb/html/
http://www.ap-lab.com/circular_dichroism.htm
Lindner, P. and Zemb, T. (2002). Neutron, X-rays and Light. Scattering Methods Applied to Soft Condensed Matter, rev. edn. Amsterdam: North-Holland. (In-depth coverage of theory and applications of light scattering at expert level.)Google Scholar
Villari, V. and Micali, N. (2008). Light scattering as spectroscopic tool for the study of disperse systems useful in pharmaceutical sciences. Journal of Pharmaceutical Science, 97, 1703–1730.CrossRefGoogle Scholar
http://www.ap-lab.com/light_scattering.htm
http://www.people.vcu.edu/~ecarpenter2/Tutorial.html
L'vov, B. V. (2005). Fifty years of atomic absorption spectrometry. Journal of Analytical Chemistry, 60, 382–392.CrossRefGoogle Scholar
Zybin, A., Koch, J., Wizemann, H. D., Franzke, J. and Niemax, K. (2005). Diode laser atomic absorption spectrometry. Spectrochimica Acta, B60, 1–11.Google Scholar
http://www.colorado.edu/physics/2000/quantumzone/index.html
http://zebu.uoregon.edu/nsf/emit.html

Save book to Kindle

To save this book to your Kindle, first ensure coreplatform@cambridge.org is added to your Approved Personal Document E-mail List under your Personal Document Settings on the Manage Your Content and Devices page of your Amazon account. Then enter the ‘name’ part of your Kindle email address below. Find out more about saving to your Kindle.

Note you can select to save to either the @free.kindle.com or @kindle.com variations. ‘@free.kindle.com’ emails are free but can only be saved to your device when it is connected to wi-fi. ‘@kindle.com’ emails can be delivered even when you are not connected to wi-fi, but note that service fees apply.

Find out more about the Kindle Personal Document Service.

Available formats
×

Save book to Dropbox

To save content items to your account, please confirm that you agree to abide by our usage policies. If this is the first time you use this feature, you will be asked to authorise Cambridge Core to connect with your account. Find out more about saving content to Dropbox.

Available formats
×

Save book to Google Drive

To save content items to your account, please confirm that you agree to abide by our usage policies. If this is the first time you use this feature, you will be asked to authorise Cambridge Core to connect with your account. Find out more about saving content to Google Drive.

Available formats
×