Source: Laboratory of Dr. B. Jill Venton - University of Virginia
Gas chromatography (GC) is used to separate and detect small molecular weight compounds in the gas phase. The sample is either a gas or a liquid that is vaporized in the injection port. Typically, the compounds analyzed are less than 1,000 Da, because it is difficult to vaporize larger compounds. GC is popular for environmental monitoring and industrial applications because it is very reliable and can be run nearly continuously. GC is typically used in applications where small, volatile molecules are detected and with non-aqueous solutions. Liquid chromatography is more popular for measurements in aqueous samples and can be used to study larger molecules, because the molecules do not need to vaporize. GC is favored for nonpolar molecules while LC is more common for separating polar analytes.
The mobile phase for gas chromatography is a carrier gas, typically helium because of its low molecular weight and being chemically inert. Pressure is applied and the mobile phase moves the analyte through the column. The separation is accomplished using a column coated with a stationary phase. Open tubular capillary columns are the most popular columns and have the stationary phase coated on the walls of the capillary. Stationary phases a…
Two-dimensional gel electrophoresis (2DGE) is a technique that can resolve thousands of biomolecules from a mixture. This technique involves two distinct separation methods that have been coupled together: isoelectric focusing (IEF) and sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). This physically separates compounds across two axes of a gel by their isoelectric points (an electrochemical property) and their molecular weights.
The procedure in this video covers the main concepts of 2DGE and a general procedure for characterizing the composition of a complex protein solution. Three examples of this technique are shown in the applications section, including biomarker detection for disease initiation and progress, monitoring treatment in patients, and the study of proteins following posttranslational modification (PTM).
Two-dimensional, or 2D, gel electrophoresis is a technique utilizing two distinct separation methods which can separate thousands of proteins from a single mixture. One of the techniques, SDS-PAGE or sodium dodecyl sulfate polyacrylamide gel electrophoresis, cannot fully separate complex mixtures alone. 2D gel electrophoresis couples the SDS-PAGE to a second method, isoelectric focusing or IEF, which separates based on isoelectric points, allowing for the resolution of potentially a…
Source: Laboratory of Dr. Neal Abrams — SUNY College of Environmental Science and Forestry
Determining the composition of a solution is an important analytical and forensic technique. When solutions are made with water, they are referred to as being aqueous, or containing water. The primary component of a solution is referred to as the solvent, and the dissolved minor component is called the solute. The solute is dissolved in the solvent to make a solution. Water is the most common solvent in everyday life, as well as nearly all biological systems. In chemistry labs, the solvent may be another liquid, like acetone, ether, or an alcohol. The solute can be a liquid or a solid, but this experiment only addressesthe determination of solids.…
Source: Laboratory of Jeff Salacup - University of Massachusetts Amherst
The product of an organic solvent extraction, a total lipid extract (TLE), is often a complex mixture of hundreds, if not thousands, of different compounds. The researcher is often only interested in a handful of compounds. The compounds of interest may belong to one of several classes of compounds, such as alkanes, ketones, alcohols, or acids (Figure 1), and it may be useful to remove the compound classes to which it does not belong in order to get a clearer view of the compounds you are interested in. For example, a TLE may contain 1,000 compounds, but the Uk'37 sea surface temperature proxy is based on only two compounds (alkenones) and the TEX86 sea surface temperature proxy is based on only four (glycerol dialkyl glycerol tetraethers). It would behoove the researcher to remove as many of the compounds they are not interested in. This makes the instrumental analysis of the compounds of interest (alkenones or GDGTs) less likely to be complicated by other extraneous compounds.
In other cases, an upstream purification technique may have produced compounds you wish to now remove from the sample, such as the production of carboxylic acids during saponification in our …
Source: Laboratory of Dr. Paul Bower - Purdue University
The method of standard additions is a quantitative analysis method, which is often used when the sample of interest has multiple components that result in matrix effects, where the additional components may either reduce or enhance the analyte absorbance signal. That results in significant errors in the analysis results.
Standard additions are commonly used to eliminate matrix effects from a measurement, since it is assumed that the matrix affects all of the solutions equally. Additionally, it is used to correct for the chemical phase separations performed in the extraction process.
The method is performed by reading the experimental (in this case fluorescent) intensity of the unknown solution and then by measuring the intensity of the unknown with varying amounts of known standard added. The data are plotted as fluorescence intensity vs. the amount of the standard added (the unknown itself, with no standard added, is plotted ON the y-axis). The least squares line intersects the x-axis at the negative of the concentration of the unknown, as shown in Figure 1.
Source: Laboratory of Dr. B. Jill Venton - University of Virginia
Sample preparation is the way in which a sample is treated to prepare for analysis. Careful sample preparation is critical in analytical chemistry to accurately generate either a standard or unknown sample for a chemical measurement. Errors in analytical chemistry methods are categorized as random or systematic. Random errors are errors due to change and are often due to noise in instrument. Systematic errors are due to investigator or instrumental bias, which introduces an offset in the measured value. Errors in sample preparation are systematic errors, which will propagate through analysis, causing uncertainty or inaccuracies through improper calibration curves. Systematic errors can be eliminated through correct sample preparation and proper use of the instrument. Poor sample preparation can also sometimes cause harm to the instrument.…
Biosensors are devices that use a wide range of biological processes and physical properties in order to detect either a biological molecule, such as a protein or cell, or a non-biological molecule, such as a chemical component or contaminant. This interdisciplinary field utilizes electrical, optical, electrochemical, or even mechanical properties to detect the presence of the target molecule.
This video introduces the field of biosensing, and reviews common types of biosensor technologies. This video also discusses key challenges in the field, and provides insight into how biosensors are used in the field. …