Alcohol: Methods and Protocols (Methods in Molecular Biology)

Alcohol: Methods and Protocols (Methods in Molecular Biology)

Laura E. Nagy

Language: English

Pages: 420

ISBN: 1588299066

Format: PDF / Kindle (mobi) / ePub

This book examines the pleiotropic effects of ethanol in animal and cell culture models through a collection of detailed procedures written by experts in the field. Sections present clearly defined models of ethanol exposure, recent advances in the development of specific methodologies to mimic the impact of ethanol metabolism in cultured cells, and methodologies to investigate a variety of cells and tissues that are known to be disrupted by ethanol, amongst other topics.

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it flows through the feeding lines (19). If, as the pup matures and becomes more active, the feeding line becomes unhooked, the connection may be taped. While in the home cage, the dam licks the pups, stimulating their ano-genital area to induce urination and defecation. The pups in isolation will have difficulty excreting by themselves. Softly stimulating the ano-genital area with a cotton ball or a piece of soft tissue wet with warm water will stimulate excretion. This task needs to be

substrate II or Granzyme B substrate II, Z-IETD-AFC; caspase 9 substrate I, fluorogenic, AC-LEHD-AFC respectively, CalBiochem) and incubated for 12 h in a 30°C shaking water bath overnight. 5. Caspase activities are determined by measuring the fluorescence associated with released AMC (7-amino-4-methyl-coumarin, excitation at 380 nm, emission at 460 nm) or AFC (excitation at 400 nm, emission at 505 nm) in a spectrofluorimeter. Results are expressed as arbitrary units per 100 µg cell lysate

solutions are recommended for preparing 20% ethanol solutions. The determination of the appropriate volume of 20% ethanol to administer via intraperitoneal injection is based upon multiple experiments using mice of various strains, sexes, weights, and ages. For any given volume of ethanol administered, the weight of the animal was the only independent factor that predicted alcohol level 30 minutes after intraperitoneal injection. Determination of the amount of 20% ethanol to administer is based

brain corresponding to the structural precursor of the neocortex were isolated, and care was taken to exclude the structural precursors to the striatum and hippocampus. Individual cortical fragments (see Note 4) are collected in sterile 15mL conical tubes and gently triturated in trypsin/EDTA. Trypsin is inactivated with, D. M.EM containing 10% fetal bovine serum. The cell suspension is centrifuged for 5 min at 18°C, 1000 rpm (300 g). Cell pellets are resuspended in chilled, D. P. BS containing

of the intestinal epithelium. However, the effects of acetaldehyde on the TJ and AJ and its prevention by EGF and L-glutamine were confirmed in human colonic mucosa (14). Therefore, valuable information can be derived from studies that use Caco-2 cell monolayers as an experimental model of the intestinal epithelium. 13 Acetaldehyde and Intestinal Permeability 2 2.1 173 Materials Cell Culture 1. Dulbecco’s Modified Eagle’s Medium (DMEM; Gibco/BRL, Bethesda, MD) supplemented with 10% fetal

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