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July 2010 Laboratory Focus www.bioscienceworld.ca
FEATURE fragments. Limitations of LC/ MS/MS are that the instrument is expensive, operation of the instrument is complex and even when operated in a highly targeted manner, it is not as sensitive as ligand binding assays. LC/MS/MS can be set up to accomplish a number of different activi-
ties at once such as provide impurity information, identity and quantitation all within a single experiment. LC/MS/MS can also be used to identify oligonucleotides by providing both the mass and sequencing the oligonucleotide. Absorption Spectroscopy
is a very quick and inexpensive approach to monitoring total protein concentration. Prior to putting the sample on the spectrophotometer, there are a variety of sample treatment approaches (including the Bradford assay, Lowry assay, BCA assay) that when these reagents are
combined with peptides/proteins, it leads to a change in the absorption of the sample and the intensity of the absorbance at the measured wavelength provides quantitative information. Advantages of this approach include that it is a quick and cost effective measure of determining pro-
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tein quantity. Disadvantages include that the approach lacks specificity and cannot differentiate between proteins, and thus require the sample to be very pure to be able to determine the concentration of the protein of interest. The technique provides no information regarding the identity of the protein or peptide, and measures total protein concentration, not specifically the analyte of interest. Although absorption ratios of different wavelengths can provide some insight into the purity of DNA or RNA sample, this approach cannot provide close to the same level of purity information as gel electrophoresis, HPLC or CE.
Activation Laboratories Ltd is a contract analytical laboratory that specializes in analytical testing services and analytical method development services of metals, small molecules and large biomolecules. ABOUT THE AUTHORS: Michael Hoffman PhD MBA is director of biotechnology and bioanalytical services for Activation Laboratories Ltd and has extensive experience in protein based analytical techniques; Steven Jenkins PhD, is a senior R&D scientist at Activation Laboratories Ltd and has extensive experience in separations strategies and mass spectrometry; Paul Tressel PhD, is a senior R&D scientist and has extensive experience in biomolecule analysis; Mona Raita MSc is a Senior Research Scientist at Activation Laboratories Ltd and specializes in ligand binding assays and oligonucleotide assays; and Katie Moyer PhD is a Senior Research Scientist at Activation Laboratories Ltd and specializes in oligonucleotide and DNA based techniques.
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