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1.
Circulating tumor cells (CTCs), shed from primary tumors and disseminated into peripheral blood, are playing a major role in metastasis. Even after isolation of CTCs from blood, the target cells are mixed with a population of other cell types. Here, we propose a new method for analyses of cell mixture at the single-cell level using a microfluidic device that contains arrayed electroactive microwells. Dielectrophoretic (DEP) force, induced by the electrodes patterned on the bottom surface of the microwells, allows efficient trapping and stable positioning of single cells for high-throughput biochemical analyses. We demonstrated that various on-chip analyses including immunostaining, viability/apoptosis assay and fluorescent in situ hybridization (FISH) at the single-cell level could be conducted just by applying specific reagents for each assay. Our simple method should greatly help discrimination and analysis of rare cancer cells among a population of blood cells. 相似文献
2.
Purification and Characterization of Sucrose Synthase from Peach (Prunus persica) Fruit 总被引:4,自引:0,他引:4
Sucrose synthase (EC 2.4.1.13
[EC]
) was purified from peach fruit(Prunus persica) to a single band of protein on SDS-PAGE byammonium sulfate fractionation, DEAE-cellulose (DE-52) chromatography,Sepharose CL-6B gel filtration, PBA-60 affinity chromatographyand Sephadex G-200 gel filtration. The molecular weight wasestimated to be 360,000 by gel filtration. The enzyme was foundto be a tetramer of identical 87-kDa subunits. The maximum activityfor the synthesis and cleavage of sucrose was observed at pH8.5 and pH 7.0, respectively. The enzymatic reaction followedtypical Michaelis-Menten kinetics in both directions, with thefollowing parameters: Km(fructose), 4.8 mmM; Km(UDPglucose),0.033 mM; Km(sucrose), 62.5 mM; Km(UDP), 0.080 mM. Other properties,such as substrate specificity and the effects of divalent cations,were also investigated. The relationship between the enzymeand the accumulation of sucrose in peach fruit is discussed.
Present address: Laboratory of Horticulture, Faculty of Agriculture,Nagoya University, Chikusa, Nagoya 464, Japan. (Received May 2, 1988; Accepted September 14, 1988) 相似文献
3.
An alkaline solution of bismuth subnitrate reacted well with the cell membranes and cell walls of formaldehyde-glutaraldehyde potassium permanganate fixed Alternaria spores, demonstrating them with greater contrast than in sections stained with uranyl acetate and lead citrate. Optimal fine structure of fungal spores was obtained by en bloc staining with alkaline bismuth solution after aldehyde and permanganate fixation. The contrast of the cell organelles and cell walls was high enough in sections cut after the alkaline bismuth en bloc stain for direct ultrastructural observation. Our results indicate that the alkaline bismuth stain is useful either as an en bloc or section stain for aldehyde and permanganate fixed fungal spores. 相似文献
4.
5.
The uptake of sorbitol into vacuoles from immature flesh ofapple fruit (Maluspumila Mill, var domestica Schneid.) was facilitatedby 106 M ABA, while such uptake into protoplasts wasnot stimulated. By contrast, the application of 105 MIAA facilitated uptake of sorbitol into protoplasts but didnot significantly into vacuoles. (Received July 17, 1990; Accepted December 25, 1990) 相似文献
6.
7.
NADP+-Dependent Sorbitol Dehydrogenase Found in Apple Leaves 总被引:1,自引:0,他引:1
An NADP+-dependent sorbitol dehydrogenase that catalyzes sorbitoland glucose was found in apple leaves. The partially purifiedenzyme had optimum activity at pH 9.6 and a Km value of 128mM for sorbitol. Among the polyols studied, this enzyme showedthe most activity for sorbitol.
1This paper is contribution A-173 of the Fruit Tree ResearchStation. (Received June 4, 1984; Accepted July 31, 1984) 相似文献
8.
Subcellular Localization of Sorbitol-6-phosphate Dehydrogenase in Protoplast from Apple Cotyledons 总被引:1,自引:0,他引:1
The subcellular localization of sorbitol-6-phosphate (S6P) dehydrogenasein protoplasts of apple cotyledons was examined by differentialcentrifugation and linear sucrose density gradient centrifugation(3060%, w/w). The distribution of S6P dehydrogenase activitywas 55% in the 500 x g pellet of the homogenate and 35% in thesupernatant of 105,000 x g. When the x g pellet was recentrifugedin a linear sucrose density gradient, one major peak of activitywas found at a density of 1.23. This peak coincided with themajor peak of chlorophyll and NADP+-triose-P dehydrogenase activity.When the 500 x g pellet was sonicated, the major peak of S6Pdehydrogenase activity shifted to a lighter density (d=1.18).The shifted peak also coincided with the peak of chlorophyll.The enzyme detected in the major peak of chlorophyll (d=1.23)was partially solubilized by sonic or detergent treatment, butnot by hypotonic solution. The results supported the localizationof S6P dehydrogenase in chloroplasts, and presumably their associationwith thylakoid membranes. Part of the enzyme was assumed tobe naturally present in the cytosol, too. (Received November 4, 1980; Accepted January 21, 1981) 相似文献
9.
A new type of sorbitol oxidase which converts sorbitol to glucosein the absence of NAD or NADP was found in the leaf of the apple.The partially purified enzyme consumed 1/2 mole of oxygen inconverting one mole of sorbitol to glucose (sorbitol+1/2 O2 glucose+H2O). The enzyme had its optimum activity at pH 4.0,and it had a Km value of 100 mM for sorbitol and showed a weakdependency on divalent cations. This sorbitol oxidase seemsto play an important role in the utilization of accumulatedsorbitol.
1This paper is contribution A-109, Fruit Tree Research Station (Received March 15, 1980; ) 相似文献
10.
Hideo Kawaguchi Alain A. Verts Shohei Okino Masayuki Inui Hideaki Yukawa 《Applied microbiology》2006,72(5):3418-3428
The aerobic microorganism Corynebacterium glutamicum was metabolically engineered to broaden its substrate utilization range to include the pentose sugar xylose, which is commonly found in agricultural residues and other lignocellulosic biomass. We demonstrated the functionality of the corynebacterial xylB gene encoding xylulokinase and constructed two recombinant C. glutamicum strains capable of utilizing xylose by cloning the Escherichia coli gene xylA encoding xylose isomerase, either alone (strain CRX1) or in combination with the E. coli gene xylB (strain CRX2). These genes were provided on a high-copy-number plasmid and were under the control of the constitutive promoter trc derived from plasmid pTrc99A. Both recombinant strains were able to grow in mineral medium containing xylose as the sole carbon source, but strain CRX2 grew faster on xylose than strain CRX1. We previously reported the use of oxygen deprivation conditions to arrest cell replication in C. glutamicum and divert carbon source utilization towards product production rather than towards vegetative functions (M. Inui, S. Murakami, S. Okino, H. Kawaguchi, A. A. Vertès, and H. Yukawa, J. Mol. Microbiol. Biotechnol. 7:182-196, 2004). Under these conditions, strain CRX2 efficiently consumed xylose and produced predominantly lactic and succinic acids without growth. Moreover, in mineral medium containing a sugar mixture of 5% glucose and 2.5% xylose, oxygen-deprived strain CRX2 cells simultaneously consumed both sugars, demonstrating the absence of diauxic phenomena relative to the new xylA-xylB construct, albeit glucose-mediated regulation still exerted a measurable influence on xylose consumption kinetics. 相似文献