Preparation of compound-specific and group-specific antibodies to 7-methylguanine and related 7-alkylguanines and their use in immunoaffinity purification

The preparation and characteristics of compound-specific and group-specific antibodies against 7-alkylguanines (7-alkGua) are described. A compound-specific antibody against 7-methylguanine was prepared using a hapten bound to carrier protein through the N2 position. In a competitive enzyme-linked immunosorbent assay (ELISA) 7-methylguanine (7-MeGua) showed 50% inhibition (I50%) at 10 pmol/well at room temperature, but the inhibition was found to be 40 times better at 4 degrees C (I50% at 250 fmol/well). When the antibody was bound to protein A-Sepharose CL4B 7-MeGua was retained in immunoaffinity columns. A group-specific antibody to 7-alkGua was prepared using 7-(2-carboxyethyl)guanine (7-CEGua) bound to carrier protein via the carboxyl group. In a competitive ELISA, this antibody cross-reacted well with 7-CEGua, 7-ethylguanine (7-EtGua), 7-(2-hydroxyethyl)guanine (7-HOEtGua) and 7-(2',3'-dihydroxy)-propylguanine (7-DHPGua) and some inhibition was seen with 7-MeGua. Immunoaffinity columns prepared from this antibody retained a number of 7-alkGua of diverse structure. 7-EtGua in calf thymus DNA treated with diethylsulphate and ethylnitrosourea was isolated by immunoaffinity purification and quantified by HPLC-fluorescence. These results illustrate the potential of immunoaffinity purification for both individual DNA adducts and groups of adducts.     

Optical properties of amorphous carbon thin films prepared by plasma deposition in a graphite hollow cathode

Carbon films have been produced using a graphite hollow cathode and an r.f. plasma of hydrogen or hydrogen and argon. No hydrocarbon gas was used. The films were subjected to heat treatments up to 700 °C in air. The optical properties of the films were studied using Fourier transform IR, UV-visible, Raman, ellipsometry and photoluminescence techniques before and after the annealing. Films prepared without argon have a narrower band gap and a different photoluminescence response from films prepared with argon. The index of refraction was seen to have values between 1.7 and 2.2 depending on the preparation conditions. The optical band gap of the films was generally greater than 2 eV and the annealing process both increased the gap and reduced the density of states responsible for the large photoluminescence signal.     

Structure of metal site in Cd-substituted His117Gly mutant of azurin with and without addition of imidazole derivatives

The present work uses 111mCd-perturbed angular correlations of gamma-rays (PAC) to investigate the structure of the metal site of the His117Gly mutant of Pseudomonas aeruginosa azurin in aqueous solution and the effect on the structure upon addition of the following exogenous ligands: imidazole, 4-methyl imidazole, 1-methyl imidazole, 2-methyl imidazole and histidine. The nuclear quadrupole interaction of cadmium bound to the mutant without addition of exogenous ligands shows a strong pH dependence with three different nuclear quadrupole interactions consistent with two pKa values at about 7.2 and 8.6 at 2 degrees C. Addition of the imidazole derivatives resulted in a significant change in the PAC spectrum showing that they coordinate. This is in accordance with observations by EPR for the same mutant with copper at the metal site [den Blaauwen, T. & Canters, G. W. (1993) J. Am. Chem. Soc. 115, 1121-1129]. However, whereas EPR and ultraviolet/visual absorption show that the characteristics of the wild-type copper protein are regained by addition of the imidazole derivatives with the exception of the possible bidentates (histidine and histamine), the comparison of the PAC results to model calculations shows that the cadmium ion must be fourfold coordinated in most cases, probably binding an additional water or hydroxide ligand. A fourfold coordination is in contrast to cadmium-substituted wild-type azurin where PAC data inferred a threefold coordination by a Cys and two His residues [Danielsen, E. Bauer, R., Hemmingsen, L., Andersen. M., Bjerrum, M. J., Butz, T., Tr?ger, W., Canters, G. W., Hoitink, C. W. G., Karlsson, G., Hansson, O. & Messerschmidt, A. (1995) J. Biol. Chem. 270, 573-580]     

Biofiltration of toluene in the absence and the presence of ethyl acetate under continuous and intermittent loading

BACKGROUND: Two peat biofilters were used for the removal of toluene from air for one year. One biofilter was fed with pure toluene and the other received 1:1 (by weight) ethyl acetate:toluene mixture. RESULTS: The biofilters were operated under continuous loading: the toluene inlet load (IL) at which 80% removal occurred was 116 g m^?3 h^?1 at 57 s gas residence time. Maximum elimination capacity of 360 g m^?3 h^?1 was obtained at an IL of 745 g m^?3 h^?1. The elimination of toluene was inhibited by the presence of ethyl acetate. Intermittent loading, with pollutants supplied for 16 h/day, 5 days/week, did not significantly affect the removal efficiency (RE). Biomass was fully activated in 2 h after night closures, but 6 h were required to recover RE after weekend closures. Live cell density remained relatively constant over the operational period, while the dead cell fraction increased. Finally, a 15 day starvation period was applied and operation then re‐started. Performance was restored with similar re‐acclimatization period to that after weekend closures, and a reduction in dead cell fraction was observed. CONCLUSION: This study demonstrates the capacity of the system to handle intermittent loading conditions that are common in industrial practices, including long‐term starvation. Copyright © 2008 Society of Chemical Industry     

An improved system for the control of dissolved oxygen in freshwater aquaria

A system for the removal and control of dissolved oxygen (DO) from freshwater was designed and constructed with aquarium-type fish studies in mind. Degassed water was obtained using a partial vacuum of -14 psi, and DO regulated at an aquarium scale using electronically controlled aeration with timed partial water renewal. The degassing system was capable of producing water with approximately 1.7 mg L(-1) DO within 10 min of operation, and 0.55 mg L(-1) after 2h. The control system was capable of maintaining DO levels of ca 0.8 mg L(-1) over 48 h in the absence of aeration and further capable of precisely controlling DO levels as low as 1.16+/-0.002 mg L(-1) (mean+/-SEM) with aeration over a 48 h period.     

Lumbar lordosis. Effects of sitting and standing

STUDY DESIGN: The effect of sitting versus standing posture on lumbar lordosis was studied retrospectively by radiographic analysis of 109 patients with low back pain. OBJECTIVE: To document changes in segmental and total lumbar lordosis between sitting and standing radiographs. SUMMARY OF BACKGROUND DATA: Preservation of physiologic lumbar lordosis is an important consideration when performing fusion of the lumbar spine. The appropriate degree of lumbar lordosis has not been defined. METHODS: Total and segmental lumbar lordosis from L1 to S1 was assessed by an independent observer using the Cobb angle measurements of the lateral radiographs of the lumbar spine obtained with the patient in the sitting and standing positions. RESULTS: Lumbar lordosis averaged 49 degrees standing and 34 degrees sitting from L1 to S1, 47 degrees standing and 33 degrees sitting from L2 to S1, 31 degrees standing and 22 degrees sitting from L4 to S1, and 18 degrees standing and 15 degrees sitting from L5 to S1. CONCLUSION: Lumbar lordosis while standing was nearly 50% greater on average than sitting lumbar lordosis. The clinical significance of this data may pertain to: 1) the known correlation of increased intradiscal pressure with sitting, which may be caused by this decrease in lordosis; 2) the benefit of a sitting lumbar support that increases lordosis; and 3) the consideration of an appropriate degree of lordosis in fusion of the lumbar spine.     

Factors affecting contrasting results between self-reported and performance-based levels of physical limitation

Molecular electrostatic potential (MEP) maps of certain 3-methoxy flavone derivatives having different anti-picornavirus activities have been studied. Geometries of the molecules were optimised and charge distributions computed using the AM1 molecular orbital method. Hybridization displacement charges (HDC) were combined with the L?wdin charge distributions to compute the MEP maps. Reliability of the method of computing MEP maps was tested by studying certain other molecules for which ab initio MEP results are available. The anti-picornavirus activities of the flavones have been shown to be related with negative MEP values in two regions, one near the 3-methoxy group and another in a diagonally opposite region near the substituent attached to the C7 atom of the molecules.     

Congenital adrenal hyperplasia presenting as massive adrenal incidentalomas in the sixth decade of life: report of two patients with 21-hydroxylase deficiency

Divergent recommendations exist regarding the evaluation of adrenal incidentalomas. Recent data have indicated a prevalence of adrenal tumors of 71% in nonclassical congenital adrenal hyperplasia (CAH) and unmasked heterozygotes. These data expand the differential diagnosis of such incidental tumors and substantially modify the approach to their evaluation. We present two patients, female pseudohermaphrodites with the simple virilizing form of CAH and 21-hydroxylase deficiency, who functioned successfully as married phenotypic males. Both came to medical attention in the sixth decade by virtue of massive adrenal incidentalomas encountered in the evaluation of recurrent urinary tract infections. Each had a 46, XX karyotype, no palpable testes, and markedly elevated baseline levels of 17-hydroxyprogesterone (17-OH Prog) of 6086 ng/dL and 6750 ng/dL. Both responded appropriately to dexamethasone suppression with reduction of 17-OH Prog, androgens and, in the second patient, ACTH to normal or near normal levels. Histologic and autopsy examination of the first patient's tumor and computed tomographic characteristics of the second revealed a benign adenoma and myelolipoma respectively. We extend and confirm previous recommendations that CAH be included in the differential diagnosis of adrenal incidentaloma and that baseline 17-OH Prog. levels be obtained, with ACTH stimulation if necessary, to diagnose the presence of nonclassical CAH.