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George E. Boyhan, Norman E. Schmidt, Floyd M. Woods, David G. Himelrick and William M. Randle

A spectrophotometric assay for pyruvic acid in onion has been adapted to a microplate reader. Correlations between the spectrophotometer and microplate reader ranged from 0.991 to 0.997 for sodium pyruvate standards and 0.899 to 0.934 for onion samples. Onion pungency values were slightly higher with the microplate reader for both sample and background compared to the spectrophotometer when both are used in the single wavelength mode. Comparing the spectrophotometer in the single wavelength mode to the microplate reader in the dual wavelength mode resulted in no statistically significant difference between them. Standards for both the microplate reader and spectrophotometer followed a quadratic function.

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Donald Livingstone III, Barbie Freeman, Cecile L. Tondo, Kathleen A. Cariaga, Nora H. Oleas, Alan W. Meerow, Raymond J. Schnell and David N. Kuhn

represented as error bars for differing volumes ( A ) and SG concentrations ( B ). Creation of standard curves. Samples of salmon sperm DNA ranging from 0 ng·μL −1 to 48 ng·μL −1 were read in triplicate on a Bio-tek FLx 800 microplate

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R.S. Arias, A.M. Alvarez and P.K. Murakami

The purpose of this research was to develop a rapid microplate assay for detecting and presumptively identifying pathogenic pectolytic Erwinia sp. in ornamental propagative stock and to readily distinguish them from nonpathogenic bacteria associated with Aglaonema. The assay was developed by modifying an existing crystal violet-sodium polypectate (CVP) medium to visualize the depolymerization of pectate by addition of bromcresol purple (BCP), then acidifying with a dilute hydrochloric acid (HCl) solution. The assay was sufficiently sensitive to detect latent (symptomless) infections in small sections of leaf or stem tissue. Based on inoculum titration assays, 40 to 600 colony-forming units (CFU) of Erwinia chrysanthemi (Burkholder et al.) were required to produce symptoms in Aglaonema stems and leaves, respectively. The microplate assay was able to detect the pathogen at low levels (40 to 60 CFU) in tissue segments of ≈1 cm2. In tests of bacterial strains isolated from 211 samples from Aglaonema `Silver Queen', `Emerald Beauty', and `San Luis' grown in Hawaii, only the pectolytic and pathogenic Erwinia sp. reacted positively in the microplate assay. Other bacteria associated with Aglaonema, including Pseudomonas paucimobilis (Holmes), P. vesicularis (Galarneault and Leifson), and nonpectolytic Erwinia sp., were not detected by the assay. Pectolytic strains of Ralstonia (Pseudomonas) solanacearum (Smith) were differentiated from pectolytic Erwinia sp. by the yellow color that developed in wells of the latter strains after acidification of the medium with dilute HCl. The test is visual and can be performed with minimal equipment and cost.

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Todd C. Einhorn, Cecil Stushnoff, Ann E. McSay, Phil L. Forsline, Sam Cox, Joel R.L. Ehrenkranz and Loretta Sandoval

Phlorizin is known for its role in reducing glucotoxicity and has a long history of use in diabetes research. In addition, its contribution to the pool of total phenolics adds to the overall health benefits attributed to fruit. Phlorizin is limited to Rosaceae family plants, of which apple comprises its current commercial source; however, limited information exists regarding its biodiversity among apple taxa. A subset of 22 taxa from a core collection of apple accessions representative of the global genetic diversity of apple was used to investigate the biodiversity of phlorizin present in apple shoots and in fruit relative to total phenolic content and free radical scavenging capacity. Fruit and shoots were harvested from the USDA Plant Genetic Resources Unit in Geneva, N.Y. Validation and quantification of phlorizin was conducted using a rigorous high-pressure liquid chromatography (HPLC) procedure. Total phenolics in fruit, assayed using a Folin-Ciocalteu method and expressed as gallic acid equivalents, ranged from 227 to 7181 mg·L-1

and were strongly related to 2,2' azinobis (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) antioxidant capacity for the core collection (r= 0.778). On a molar basis, phlorizin had lower antioxidant capacity than other major phenolic compounds present in apple fruit, but was more effective than ascorbic acid. Phlorizin yield in dormant apple shoots, expressed as percent weight, ranged from 0.9% to 5.5%. A rapid, 96 well micro-plate spectrophotometric assay was also developed to aid in the screening of multiple samples for selection of high phlorizin yielding apple taxa. Spectrophotometry overestimated phlorizin content as expected, but the calibration curve between HPLC and spectrophotometry was acceptable, r 2 = 0.88.

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Kevin M. Crosby, Justin Butcher, Kil Sun Yoo and Daniel I. Leskovar

taken with a microplate reader at 405 nm to determine virus infection. Leaves from ‘NM 6’ tested positive for TEV, PepMoV, and PVY ( Table 3 ). Leaves from ‘TAM Ben Villalon’ tested weakly positive for TEV and PepMoV and negative for PVY. Leaves of a

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Desire Djidonou, Amarat H. Simonne, Karen E. Koch, Jeffrey K. Brecht and Xin Zhao

mixtures were subsequently incubated in a water bath at 60 °C for 3 h followed by slow addition of 2.5 mL ice-cold 90% (v/v) H 2 SO 4 . Thereafter, the absorbance of 250 µL samples was measured at 540 nm using a microplate spectrophotometer (model Power

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Itani Tshivhandekano, Fhatuwani Nixwell Mudau and Thilivhali Emmanuel Tshikalange

yield a stock concentration of 0.04 mg·mL –1 . All samples were tested in triplicate using 96-well microplates. The startup concentration for the test samples in the first row was 500 µg·mL –1 , which was a composition of distilled water and the extract

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Amy K. Freidig and Irwin L. Goldman

DMAB [3-(dimethylamino) benzoic acid] in the presence of peroxidase to produce the detectable indamine dye. Volumes were scaled down for use in a 96-well microplate. Reagents were prepared according to the kit instructions. Thawed samples were added to

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Nobuyuki Fukuoka, Takamoto Suzuki, Keisuke Minamide and Tatsuro Hamada

ethanolic extract was dried at 60 °C and dissolved in 200 μL of distilled water, and a 5-μL aliquot of solution was brought up to 45 μL with distilled water in each well of a 96-well microplate. Then, 50 μL of Folin-Ciocalteau reagent and 50 μL of 100 mg

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Masahumi Johkan, Kazuhiro Shoji, Fumiyuki Goto, Shin-nosuke Hashida and Toshihiro Yoshihara

centrifuged at 15,000 g for 3 min (Tomy Co., Tokyo, Japan). The supernatant was applied to a 96-well plate, and the absorbance of the Chl in acetone was measured at wavelengths of 470, 645, and 663 nm with a 96-well Benchmark microplate reader (Bio