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Cold Spring Harbor Protocols

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https://www.readbyqxmd.com/read/28679720/immunoassays
#1
Thomas O Kohl, Carl A Ascoli
The enzyme immunoassay (EIA) is one of the most powerful of all immunochemical techniques. First described in the early 1970s, these assays are now used routinely in laboratory analyses and diagnostics. In biology and biotechnology, the EIA is a valuable and versatile tool used to detect and quantitate antigens and antibodies. Application of the appropriate EIA permits rapid quantification of different antigens and antibodies (referred to here as analytes) present at very low concentrations within a mixture...
July 5, 2017: Cold Spring Harbor Protocols
https://www.readbyqxmd.com/read/28679719/ion-channels-history-diversity-and-impact
#2
Stephan Brenowitz, Ian Duguid, Paul J Kammermeier
From patch-clamp techniques to recombinant DNA technologies, three-dimensional protein modeling, and optogenetics, diverse and sophisticated methods have been used to study ion channels and how they determine the electrical properties of cells.
July 5, 2017: Cold Spring Harbor Protocols
https://www.readbyqxmd.com/read/28679718/chemostat-culture-for-yeast-physiology-and-experimental-evolution
#3
Maitreya J Dunham, Emily O Kerr, Aaron W Miller, Celia Payen
Continuous culture provides many benefits over the classical batch style of growing yeast cells. Steady-state cultures allow for precise control of growth rate and environment. Cultures can be propagated for weeks or months in these controlled environments, which is important for the study of experimental evolution. Despite these advantages, chemostats have not become a highly used system, in large part because of their historical impracticalities, including low throughput, large footprint, systematic complexity, commercial unavailability, high cost, and insufficient protocol availability...
July 5, 2017: Cold Spring Harbor Protocols
https://www.readbyqxmd.com/read/28679717/p3-buffer
#4
(no author information available yet)
No abstract text is available yet for this article.
July 5, 2017: Cold Spring Harbor Protocols
https://www.readbyqxmd.com/read/28679716/p2-buffer
#5
(no author information available yet)
No abstract text is available yet for this article.
July 5, 2017: Cold Spring Harbor Protocols
https://www.readbyqxmd.com/read/28679715/p1-buffer
#6
(no author information available yet)
No abstract text is available yet for this article.
July 5, 2017: Cold Spring Harbor Protocols
https://www.readbyqxmd.com/read/28679714/rnase-h-10-mg-ml-dnase-free
#7
(no author information available yet)
No abstract text is available yet for this article.
July 5, 2017: Cold Spring Harbor Protocols
https://www.readbyqxmd.com/read/28679713/potassium-phosphate-buffer-1-m-ph-6-5
#8
(no author information available yet)
No abstract text is available yet for this article.
July 5, 2017: Cold Spring Harbor Protocols
https://www.readbyqxmd.com/read/28679712/vitamins-1000%C3%A3
#9
(no author information available yet)
No abstract text is available yet for this article.
July 5, 2017: Cold Spring Harbor Protocols
https://www.readbyqxmd.com/read/28679711/metals-1000%C3%A3
#10
(no author information available yet)
No abstract text is available yet for this article.
July 5, 2017: Cold Spring Harbor Protocols
https://www.readbyqxmd.com/read/28679710/sulfate-limited-chemostat-medium
#11
(no author information available yet)
No abstract text is available yet for this article.
July 5, 2017: Cold Spring Harbor Protocols
https://www.readbyqxmd.com/read/28679709/phosphate-limited-chemostat-medium
#12
(no author information available yet)
No abstract text is available yet for this article.
July 5, 2017: Cold Spring Harbor Protocols
https://www.readbyqxmd.com/read/28679708/nitrogen-limited-chemostat-medium
#13
(no author information available yet)
No abstract text is available yet for this article.
July 5, 2017: Cold Spring Harbor Protocols
https://www.readbyqxmd.com/read/28679707/glucose-limited-chemostat-medium
#14
(no author information available yet)
No abstract text is available yet for this article.
July 5, 2017: Cold Spring Harbor Protocols
https://www.readbyqxmd.com/read/28679706/indirect-competitive-enzyme-linked-immunosorbent-assay-elisa
#15
Thomas O Kohl, Carl A Ascoli
The indirect competitive ELISA (indirect cELISA) pits plate-immobilized antigen against antigens in solution for binding to antigen-specific antibody. The antigens in solution are in the test sample and are first incubated with antigen-specific antibody. These antibody-antigen complexes are then added to microtiter plates whose wells have been coated with purified antigen. The wells are washed to remove unbound antigen-antibody complexes and free antigen. A reporter-labeled secondary antibody is then added followed by the addition of substrate...
July 5, 2017: Cold Spring Harbor Protocols
https://www.readbyqxmd.com/read/28679705/direct-competitive-enzyme-linked-immunosorbent-assay-elisa
#16
Thomas O Kohl, Carl A Ascoli
The competitive enzyme-linked immunosorbent assay (ELISA) (cELISA; also called an inhibition ELISA) is designed so that purified antigen competes with antigen in the test sample for binding to an antibody that has been immobilized in microtiter plate wells. The same concept works if the immobilized molecule is antigen and the competing molecules are purified labeled antibody versus antibody in a test sample. Direct cELISAs incorporate labeled antigen or antibody, whereas indirect assay configurations use reporter-labeled secondary antibodies...
July 5, 2017: Cold Spring Harbor Protocols
https://www.readbyqxmd.com/read/28679704/isolation-of-high-molecular-weight-dna-from-monolayer-cultures-of-mammalian-cells-using-proteinase-k-and-phenol
#17
Michael R Green, Joseph Sambrook
This procedure is the method of choice for purification of mammalian genomic DNA from monolayer cultures when large amounts of DNA are required, for example, for Southern blotting. Approximately 200 µg of mammalian DNA, 100-150 kb in length, is obtained from 5 × 10(7) cultured aneuploid cells (e.g., HeLa cells).
July 5, 2017: Cold Spring Harbor Protocols
https://www.readbyqxmd.com/read/28679703/spore-analysis-and-tetrad-dissection-of-schizosaccharomyces-pombe
#18
Karl Ekwall, Geneviève Thon
Here we describe the processing of Schizosaccharomyces pombe spores in batches (random spore analysis) or through tetrad dissections. Spores are usually prepared from matings between haploid strains (producing zygotic asci) or from sporulating diploids (producing azygotic asci). In random spore analysis, a snail enzyme preparation is used to digest the walls of asci to release free spores that are diluted and plated to form colonies. In tetrad dissection, a needle attached to a micromanipulator is used to pick asci and separate spores...
July 5, 2017: Cold Spring Harbor Protocols
https://www.readbyqxmd.com/read/28679702/selecting-schizosaccharomyces-pombe-diploids
#19
Karl Ekwall, Geneviève Thon
Here we describe procedures for the selection of diploid Schizosaccharomyces pombeade6-M210/ade6-M216 heteroallelic complementation is widely used to select for Ade(+) diploids. Such diploids will readily sporulate when starved of nitrogen. For some investigations, stable diploids are preferable (e.g., for genetic complementation tests), and in these cases mating an h(-) strain with an h(90) mat2-Pi-102 strain can be used to prevent sporulation. When ade6-M210/ade6-M216 mutations impact on, or show synthetic interactions with, the gene of interest, two different auxotrophic markers can be used to select complementation...
July 5, 2017: Cold Spring Harbor Protocols
https://www.readbyqxmd.com/read/28679701/setting-up-schizosaccharomyces-pombe-crosses-matings
#20
Karl Ekwall, Geneviève Thon
Here we provide methods for setting up standard crosses with Schizosaccharomyces pombe strains. All strain genotypes and pedigrees should be recorded in a laboratory strain book. Matings between two haploid strains of interest are induced on solid medium poor in nitrogen. Usually, sporulation agar (SPA) plates are preferred, but for difficult matings it is advisable to try several mating media in parallel because one medium might allow for more efficient mating. Protoplast fusion can be used to produce zygotes from sterile mutants that fail to mate...
July 5, 2017: Cold Spring Harbor Protocols
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