News releasedfromcells activated with recombinant mouse interferon-y and lipopolysaccharide. The sensor's limit of detection is 1 uM NO. The sGC-based sensor does not respond to common interferents, including nitrate, nitrite, and oxygen. However, the presence of glutathione was found to decrease its sensitivity. "The [glutathione interference] problem is unique to guanylate cyclase," says Barker. Cytochrome c'-based NO sensors are not affected by glutathione. Because glutathione is produced by macrophages, the sGC-based sensors had to be calibrated in a phosphate buffer containing the same amount of glutathione present in the cellular environment of interest. The heme domain and sGC are both relatively unstable proteins and, therefore, denature at high temperatures. "That is where the cytochrome c'-based sensors have an advantage—they are much more stable," says Barker. The sGC-based sensors can be used at room temperature for several hours, but they cannot be stored at room temperature overnight. However, sensors stored at 0-4 °C remain stable for several days. "They're like a lot of biological materials; you'd better keep them in the fridge," remarks Kopelman. Britt Erickson
that serves as an initial screen is performed with the porogen solvent, which, in this case, is methylene chloride. A MIP is most likely to bind to a template in the porogen solvent. Therefore, complete release of the template into the porogen solvent is a good indicator that the MIP is no good. "A rapid and quantitative release of template when washing the polymers in the porogenic solvent is a good indicator for absence of high affinity imprinted sites," says Sellergren. "On the other hand, retention of template is no evidence for their presence. However, a quantitative release ecn be used aa s ariterion for discarding materials, thus saving time in the further screening for selectivity." To test their assumption that release of the template into the porogen solvent means that the MIP is not selective, Sellergren and Lanza screened all of the MIPs— even the ones that had completely released the template in the initial screen— for selectivity for rebinding of terbutylazine (a triazine herbicide that had been used as the template molecule). The initial release test indicated that only polymers made with (trifluoromethyl) acrylic acid (TFM) and methacrylic acid (MAA) should be expected to show an enhanced affinity for the template molecule in methylene chloride. The results of the rebinding experiments were as they expected. "Only the polymers that significantly retained the Rapid MIPs template in the release test showed selective rebinding proDerties " savs Sellergren screening "Thus a complete release in the porogenic Selectivity is the word for molecularly imsolvent seems to be a valid criteri[on] to printed polymers (MIPs). Unfortunately, exclude materials" many factors involved in the synthesis of MIPs ultimately affect their recognition Polymers made with MAA as the funcproperties—the type and concentration of tional monomer rhowed the highest selecthe functional monomer, the cross-linking tivity for rhe template. Interestingly, , fullmonomer, the solvent (porogen), and the scale version of the MAA polymer, which template, as well as the experimental condi- was used as a chromatographic stationary tions. Synthesizing new MIPs or optimizing phase, displayed the highest capacity faca given MIP requires systematically varytor for atrazine in acetonitrile (k' = 18) that ing these factors and analyzing the resulthas been reported under similar condiing materials. tions. The retention of atrazine waa ssgnificantly higher with a polymer imprinted That's where Borje Sellergren and with terbutylazine than with a polymer Francesca Lanza of Johannes Gutenberg imprinted with atrazine. Sellergren susUniversity Mainz (Germany) come in. like pects that this is due to the difference in Toshifumi Takeuchi and his co-workers at basicity and hydrophobicity of the temHiroshima City University Qapan) (Anal plates and because the terbutylazine site Chem. 1999, 71, 285-90), Sellergren nnd can be expected to be sterically capable of Lanza have developed a method for rapidly accommodating atrazine. synthesizing and screening "mini" MIPs, which have been produced in a scaledAccording to Sellergren, the most imdown synthetic process (Anal. Chem. portant outcome of this work is the 1999, 71,2092-96)) However, ,n Selleramount oftimeit will save in synthesizing gren and Lanza's method, the washing step and optimizing MIPs. Although only six 374 A
Analytical Chemistry News & Features, June 1, 1999
MIPs were synthesized in this work, the method should be applicable to screening large numbers of MIPs. "There is no doubt that the screening methods developed by us and [Takeuchi's group] will bring significant savings in terms of time and reagents in the search for MIPs with desired performance," says Sellergren. Sellergren says they still have important steps to consider. "It is important to demonstrate the usefulness of the method in the imprinting of difficult templates where the present established protocol fails to produce MIPs with the desired recognition properties," he says. "Furthermore, because the consumption of template is typically a few milligrams, the imprinting of more precious templates can now be systematically assessed." In addition, he says it is important, in many cases, to find MIPs that recognize analytes in water or to find materials with higher sample load capacities or improved mass transfer properties. "In view of the large number of options it be of interest to find ways to further miniaturize the process" he says. Celia Henry NEWS FROM ABRF '99
The Association of Biomolecular Resource Facilities Elizabeth Zubritsky reports from Durham, NC.
N e w biological standards considered Despite the enormous growth in bioanalytical techniques, the sole biological standard is still bovine serum albumin (BSA). However, the quality and compliance committee of the Association of Biomolecuiar Resource Facilities (ABRF) hopes to change that. The committee is working on plans for additional biological standards—peptides, proteins, and possibly other molecules—according to a report given by Alan J. Smith, chairman of the committee's working group for peptide standards. At this point, the committee has spoken to the National Institute of Standards and Technology (NIST) about developing new biolocical standards, but no formal ar-
