micrometastasis detection methods and clinical importance
Glycoprotein Methods and Protocols - P1
Ngày tải lên :
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... UV monitor and collect fractions using a fraction col-
lector and subject to the appropriate carbohydrate and antibody analyses.
3.6. Analysis of Mucins
Methods for the detection and analysis ... antibodies and
lectins or coating methods such as the glycan detection method and enzyme-linked
immunosorbent assays (ELISA). All these techniques have advantages and disadvan-
tages. Solution methods ... serine proteases and esterases, including
acetylcholinesterase, and should therefore be handled in a fume cupboard with extreme
care! DFP is supplied in 1-g vials with a septum, and prior to dilution,...
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Glycoprotein Methods and Protocols - P2
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... nature and
limitations of the individual techniques, and (2) interpretation and reporting of mucin
staining.
1.1. Nature and Limitations of Mucin-Staining Methods
Mucin histochemistry, lectin, and ... al.
Histological Methods for Detection of Mucin 43
21. Jass, J. R. and Smith, M. (1992) Sialic acid and epithelial differentiation in colorectal
polyps and cancer—a morphological, mucin and lectin ... Rhodes, J. M. and Milton, J. D. (1986) Lectin Methods and Protocols, Humana, Totowa,
NJ.
50. Liener, I. E., Sharon, N., and Goldstein, I. J. (1998) The Lectins: Properties, Functions,
and Applications...
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Glycoprotein Methods and Protocols - P3
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... Chemical, Lectin, and Antibody Methods
Michael A. McGuckin and David J. Thornton
1. Introduction
Detection and quantitation of mucins can be important in both the research and
clinical settings. ... Mucins 45
45
From:
Methods in Molecular Biology, Vol. 125: Glycoprotein Methods and Protocols: The Mucins
Edited by: A. Corfield © Humana Press Inc., Totowa, NJ
4
Detection and Quantitation of ... 1 and 2 (2,3) of this volume for the preparation of
secreted and membrane-associated mucins, respectively, and to Chapter 7 (4) for a
discussion of methods for mucin separation.
50 McGuckin and...
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Glycoprotein Methods and Protocols - P4
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... poly-
L
-lysine for 5 min
and dried at room temperature.
3. Methods
3.1. Observation and Measurement
of Mucous Thickness on Unfixed Mucosal Sections (
see
Notes 1–4)
This is a rapid and simple method ... will be greatly
reduced and discontinuous.
7. Careful handling of the cryostat sections on the slide during fixing and staining is impor-
tant because excessive washing and so on can remove mucus ... to rat, frog and human gastric mucosa: influence of feeding
prostaglandin, N-acetyl-cysteine and other agents. Clin. Sci. 63, 187–195.
4. Bell, A. E., Allen, A., Morris, E. R., and Ross-Murphy,...
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Glycoprotein Methods and Protocols - P5
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... samples no. 1 and 2 (values a and e):
x = g/c × a/e (no dimension)
Biosynthesis and Secretion of Mucin 65
65
From:
Methods in Molecular Biology, Vol. 125: Glycoprotein Methods and Protocols: ... PhosphorImager.
4. The identification of mucin precursors and mature mucins using polypeptide-specific
antisera and SDS-PAGE is elaborated in Chapters 20 and 21, and in several references
(1,4–11). The precursor ... NJ
6
Quantitation of Biosynthesis
and Secretion of Mucin Using Metabolic Labeling
Jan Dekker, B. Jan-Willem Van Klinken,
Hans A. Büller, and Alexandra W. C. Einerhand
1. Introduction
Most epithelial...
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Glycoprotein Methods and Protocols - P6
Ngày tải lên :
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... small gel, use 1.6 g of
agarose and 160 mL of buffer, and for a large gel, use 2.8 g of agarose and 280 mL of
buffer.
2. Leave to cool before pouring (hand hot) and insert well-forming comb (see ... 1.5 mm thick and 1 cm wide. There is a compromise between
band broadness and amount of sample to be loaded. Larger amounts of sample tend to
yield poorer quality data; i.e., the bands become more ... other identified mucins.
Separation and Identification of Mucin 77
7
Separation and Identification
of Mucins and Their Glycoforms
David J. Thornton, Nagma Khan, and John K. Sheehan
1. Introduction
This...
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Glycoprotein Methods and Protocols - P7
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... 2.400
a
The values for ρ and η
rel.
are from ref. 14.
Heterogeneity and Size Distribution of Gel-Forming Mucins 87
87
From:
Methods in Molecular Biology, Vol. 125: Glycoprotein Methods and Protocols: ... variety of different sizes and should be chosen according to
sample volume and concentration.
Heterogeneity and Size Distribution of Gel-Forming Mucins 91
mucins on these grids; and the addition of ... K. E., and Chapman J. A. (1985) Mica sandwhich
technique for preparing macromolecules for rotary shadowing. J. Ultrastruct. Res. 91,
66–76.
14. Kawahara K and Tanford C. (1966) Viscosity and density...
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Glycoprotein Methods and Protocols - P8
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... glassblower. It has to withstand high tempera-
ture in the presence of strong acid vapor, and hold positive and negative gas pressure
during each hydrolysis run.
2. The detection of amino acids can ... 113
113
From:
Methods in Molecular Biology, Vol. 125: Glycoprotein Methods and Protocols: The Mucins
Edited by: A. Corfield © Humana Press Inc., Totowa, NJ
10
Amino Acid Analysis of Mucins
Jun X. Yan and ... 2 min for pump and column equilibration with the initial composition of
the mobile phase.
5. Figures 1 and 2 show typical chromatograms of the separation of amino acid standards
and hydrolyzed...
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Glycoprotein Methods and Protocols - P9
Ngày tải lên :
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... Kirk, M., Barnes, S., Lowary, T. L., and Hart, G.
W. (1996) Selective detection and site-analysis of O-GlcNAc-modified glycopeptides by
β-elimination and tandem electrospray mass spectrometry. ... Curmi, P. M. G., Williams, K. L., and Gooley, A. A. (1996)
Sequencing Glycopeptides 121
121
From:
Methods in Molecular Biology, Vol. 125: Glycoprotein Methods and Protocols: The Mucins
Edited ... peptides.
Acta. Chem. Scand. 4, 283–293.
4. Edman, P and Begg, G., (1967) A protein sequencer. Eur. J. Biochem. 1, 80–91.
5. Gooley, A. A., Classon, B. J., Marschalek, R., and Williams, K. L. (1991)...
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Glycoprotein Methods and Protocols - P10
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... 129
129
From:
Methods in Molecular Biology, Vol. 125: Glycoprotein Methods and Protocols: The Mucins
Edited by: A. Corfield © Humana Press Inc., Totowa, NJ
12
Synthetic Peptides for the Analysis and Preparation
of ... monoclonal antibodies (mAbs) and fragments have been
produced by various methods. Some of these have found applications for radio-
immunoscintigraphy and targeted therapy of cancer, and others have been ... information on antibody specificity and permits identification of
significant features of an epitope that may contribute to antibody recognition and bind-
ing (see Notes 10 and 11).
1.3. Purification...
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Glycoprotein Methods and Protocols - P11
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... lysates and media. Nonspecific
bands may be present and can be eliminated from consideration using the pSVL vector
control lanes for comparison (Fig. 2A,B, lanes 1 and 3). The sizes of specific bands
reported ... polymerase, and incubate the reaction at 72˚C to allow extension of the linked
templates. Add 2 µg each of primers 5 and 6, and continue the PCR with 30 cycles at 94˚C
for 30 s, 50˚C for 1 min and 72˚C ... 195–205.
12. Bhargava, A. K., Woitach, J. T., Davidson, E. A., and Bhavanandan, V. P. (1990) Cloning
and cDNA sequence of a bovine submaxillary gland mucin-like protein containing two
distinct domains....
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Glycoprotein Methods and Protocols - P12
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... J., and Stone, B. A. (1983) Detection of neutral
and amino sugars form glycoproteins and polysaccharides as their alditol-acetates. J.
Chromatogr. 256, 419–427.
27. Takemoto, H., Hase, S., and ... Nakamura, M., and Ohkura, Y. (1987) Fluoro-
metric high performance liquid chromatography of N-acetyl- and N-glycolyl neuraminic
acids and its application to their microdetermination in human and animal ... acid (Neu5Gc), and keto-deoxy-
nonulosonic acid (Kdn), respectively. Substituents of the hydroxyl groups present on
From:
Methods in Molecular Biology, Vol. 125: Glycoprotein Methods and Protocols:...
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Glycoprotein Methods and Protocols - P13
Ngày tải lên :
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... M. J. and Hounsell, E. F. (1998) HPLC and HPAEC of oligosaccharides and glyco-
peptides, in Glycoscience Protocols (Hounsell, E. F., ed.), Humana, Totowa, NJ, pp. 79–100.
14. Kakehi, K., and Honda, ... 0.5 cm column) and elute with three
column vol of HPLC-grade water.
5. Collect the effluent and eluent and then pool (2.5-mL volume).
6. Load onto a PD10-Sephadex GM25 column and elute with HPLC-grade ... cm)
in a water jacket at 55°C and HPLC pump with refractive index and fluorescence detectors.
5. GlycoSepH™ and GlycoSepC™ HPLC column (Oxford GlycoSciences).
3. Methods
3.1. Alkaline Borohydride...
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Glycoprotein Methods and Protocols - P14
Ngày tải lên :
08/10/2012, 10:25
... or three at
the most) and the facilities that are available in the laboratory environment will define
From:
Methods in Molecular Biology, Vol. 125: Glycoprotein Methods and Protocols: The Mucins
Edited ... carbons 4 and 5 and also the exocyclic chain of
sialic acid when it is not 8 or 9 substituted. The oligosaccharide-alditol is cleaved in
two fragments, one corresponding to the 3-branch and the ... was introduced in the 1970s and rapidly
received a large application for analyzing the sequence of N-acetyllactosamine- and
oligomannosidic-type glycans.
Originally, and before the development...
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Glycoprotein Methods and Protocols - P15
Ngày tải lên :
08/10/2012, 10:25
... hygroscopic and must be thoroughly dried before weighing
and making up as standards. Standards of 1 mM each of 2-, 3-, and 6-sulfo-N-
acetylglucosamine in water may be aliquoted and stored at –20°C, and ... Harrison and Packer
2. Materials
2.1. Solvents and Reagents
1. Water, carbonate-free (see Note 1), and sparged with a constant flow of argon.
2. 0.1 M NaOH, made up with decarbonated water, and similarly ... equipment and the hydrolysis method
are similar for both methods. Although this method is not particularly sensitive by
today’s analytical standards ( >1 nmol), it provides fast and reproducible...
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