NMR for evaluating packed columns Most attempts to assess the transport properties of analytes in a packed column have been based on bandwidth measurements. However, the total bandwidth of an analyte at the column outlet also includes contributions from outside the column, such as detector cell volume, and only total mass transport characteristics can be monitored without discrimination between the individual contributions of eddy and molecular diffusion as well as adsorption and desorption kinetics. J. Karger and colleagues at Johannes Gutenberg-Universitat (Germany) and the Universitat Leipzig (Germany) have applied pulsed field gradient NMR spectroscopy to segments of columns to study molecular diffusion in beds of spherical silica particles.
Using GC/ITMS to determine chemical markers in dust
The spectrometer was used to apply magnetic field gradient pulses with amplitudes up to 25 T/m, which allowed the measurement of molecular displacements as small as 100 nm. Using cyclohexane as the probe liquid, they found that the internal pore system of the silica particles gave rise to two modes of molecular migration: restricted motion with molecular mean square displacements on the order of 700 nm and unrestricted motion with effective diffusivities decreasing with increasing loading over nearly 2 orders of magnitude. In beds of nonporous particles cyclohexane diffusivity is found to be reduced by a factor of ~ 3 Using slow-diffusing poly(dimethylsiloxane) (PDMS) as a probe for nonporous silica they found a molecular mean square displacement of PDMS at least 2 orders of magnitude smaller than that of cvclohexane (J Phv; Chem 1996 100 7729-34")
Gram-negative bacterial lipopolysaccharide. All markers were determined as trimethylsilyl (TMS) derivatives in electron impact mode. The researchers found that compared with SIM analysis using a conventional quadrupole GC/MS/MS instrument the ion-trap system yielded similar detection sensitivity and superior specificity for TMS-derivatized 3-hydroxy fatty acids, muramic acid, and ergosterol. They suggest that ITMS should be useful for determining microbial substances in other environmental samples as well. (J. Mass Spectrom. .996,31,389-96)
Inhalation of organic dust is suspected to cause various respiratory disorders, but little is known about which chemical compounds in dust are responsible for the development of specific symptoms. Although substances of microbial and fungal origin, such as lipopolysaccharides, glucans, and allergenic proteins, are among the suspected causative agents, commonly used microbial tests lack the sensitivity for determining these substances in organic dust. Anita Saraf and Lennart Larsson of the University of Lund (Sweden) have used GC/tandem ITMS to determine ergosterol, muramic acid, and 3-hydroxy fatty acids with 10-18 carbon atoms in organic dust as markers for fungal bioion-trap tandem mass spectra of 3-hydroxy fatty acids in a housi mass, bacterial peptidoglycan, and dust sample with fragmentation of m/z 175 and monitoring of m/z 73 and 131. (Adapted with permission of John Wiley & Sons.)
Rainbow blot showing human proteins actin (green), Bc1-XL (black), Bc1-Xs (black), Bc1 -2 (red), and Bax (brown) for a cell line (C) and pairs of normal (N) and tumor (T) tissue. (Courtesy of J. C. Reed and S. Kraiewski.)
Multiple antigen detection Immunoblotting is widely used to detect and compare relative levels of proteins in biological materials. When the protein sample is limited, it is desirable to gain aa much information as possible from the protein blot This task is accomplished by stripping off the first antibody and reprobing with a second antibody. However, this stripping procedure, which usually requires high concentrations of sodium dodecyl sulfate and reducing agents, elutes some of the proteins from the blot As a result, ,here is progressively less sample with each round of stripping and reprobing. John C. Reed and colleagues from the Burnham Institute have developed a method for multiple reprobings of protein blots without antibody stripping The procedure uses horseradish peroxidase (HRP) detection with a chemiluminescent substrate. After proteins are detected by enhanced chemiluminescence, the antigen-antibody complexes are reacted with a chromogenic substrate such as 3,3'-diaminobenzidine, which renders the complexes unreactive to subsequent reprobings of the membrane with additional antibodies. No stripping is required; therefore, the authors find that signal intensity remains strong even when the blot is reprobed 12 or more rimes. The procedure also works with a "rainbow Western" in which four different HRP colorimetric substrates are used seauentiallv for detecting four different antigens on the same blot (Anal Biochem 236 1996 221-28)
Analytical Chemistry News & Features, July 1, 1996 4 0 1 A
