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276 Forde diseases, cancer and infectious diseases. The potential use of pDNA in vaccines has also been shown (4–8) through the expression of specific antigens on cell membranes that help to stimulate the immune system’s response and memory. As a result of these findings, there has been an increasing demand on the biotechnology industry to supply purified pDNA for gene therapy, vaccine and research applications. The contaminants that pose a particular problem in the production of purified pDNA are anionic polymers of a similar structure, charge and physical behavior to pDNA. These contaminating anionic polymers include genomic DNA (gDNA), RNA and lipopolysaccharides or endotoxins (9). Current commercial pDNA purification techniques typically require at least three chromatographic stages to remove all of these afore-mentioned contaminating species and to meet the evermore demanding purity levels required by customers (e.g.,
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Affinity Chromatography second edit
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METHODS IN MOLECULAR BIOLOGY TM Aff
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To Tina, Emmanuella, Natalie, and A
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viii Preface Affinity tags for puri
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x Contents 10. Immobilized Metal Io
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xii Contributors Tzong-Hsien Lee
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2 Roque and Lowe cell extract conta
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4 Roque and Lowe groups critical in
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6 Roque and Lowe of reinforcement e
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8 Roque and Lowe unknown factors ar
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10 Roque and Lowe receptor domains
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12 Roque and Lowe A further issue o
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14 Roque and Lowe transgenic system
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16 Roque and Lowe 6. Arsenis, C. an
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18 Roque and Lowe 39. Burton, S.J.,
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20 Roque and Lowe 71. Bhikhabhai, R
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I Various Modes of Affinity Chromat
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26 Charlton and Zachariou Since the
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28 Charlton and Zachariou 5. Equili
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30 Charlton and Zachariou 9. Elute
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32 Charlton and Zachariou 3.3. Puri
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34 Charlton and Zachariou could als
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3 Affinity Precipitation of Protein
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Affinity Precipitation of Proteins
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4 Immunoaffinity Chromatography Stu
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Immunoaffinity Chromatography 55 2.
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Immunoaffinity Chromatography 57 A
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Immunoaffinity Chromatography 59 ab
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5 Dye Ligand Chromatography Stuart
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Dye Ligand Chromatography 63 Develo
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Dye Ligand Chromatography 65 6. Mis
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Dye Ligand Chromatography 67 Fig. 1
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Dye Ligand Chromatography 69 6. Sec
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72 Tugcu chromatography, the sorpti
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74 Tugcu non-specific interactions,
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76 Tugcu the salt counter ions on t
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78 Tugcu On a plot of log K versus
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80 Tugcu Figure 2B illustrates a ty
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82 Tugcu 2.5. Running Displacement
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84 Tugcu then the operating conditi
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86 Tugcu Table 1 Trobleshooting for
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88 Tugcu 23. Torres, A. R. and Pete
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II Affinity Chromatography Using Pu
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94 Roque and Lowe market, with 14 F
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96 Roque and Lowe and RasMol V2.7.1
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98 Roque and Lowe house using, for
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100 Roque and Lowe Gly71 and Gly72)
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102 Roque and Lowe dissolved in ace
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104 Roque and Lowe equilibration bu
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106 Roque and Lowe of color. Purple
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108 Roque and Lowe 5. Fassina, G.,
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8 Phage Display of Peptides in Liga
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Phage Display of Peptides in Ligand
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9 Preparation, Analysis and Use of
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Preparation, Analysis and Use of an
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10 Immobilized Metal Ion Affinity C
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IMAC of Histidine-Tagged Fusion Pro
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152 Godat et al. tag. HQ-tagged pro
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154 Godat et al. 2. NaCl (5 M). 3.
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156 Godat et al. 4. Invert tube to
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158 Godat et al. 3. Sonicate sample
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160 Godat et al. the above protocol
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162 Godat et al. 3.7.1. Purificatio
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164 Godat et al. 3. Adding 200 mM N
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166 Godat et al. 4. The MagneHis Ni
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168 Godat et al. 22. Lin, D., Tabb,
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170 Pattenden and Thomas 1. Introdu
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172 Pattenden and Thomas functions
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174 Pattenden and Thomas of the mal
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176 Pattenden and Thomas necessary.
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178 Pattenden and Thomas 2.4. Amylo
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180 Pattenden and Thomas 9. Thoroug
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182 Pattenden and Thomas 8. Collect
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184 Pattenden and Thomas binding ef
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186 Pattenden and Thomas 15. Cells
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188 Pattenden and Thomas 23. Martin
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13 Methods for Detection of Protein
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Detection of Protein-Protein and Pr
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212 Charlton recognition sequence,
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214 Charlton when selecting Factor
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216 Charlton 2. Materials 2.1. Reag
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218 Charlton (see Notes 12 and 13).
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220 Charlton 3.3. Cleavage of Fusio
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222 Charlton 3. The catalytic mecha
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224 Charlton may dictate. However,
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226 Charlton 22. The full-length fu
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228 Charlton 22. Lien, S., Milner,
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230 Arnau et al. downstream process
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232 Arnau et al. Fig. 1. Overview o
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234 Arnau et al. Fig. 2. The pQE-1
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236 Arnau et al. 2.3.3. Step B Buff
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238 Arnau et al. Fig. 3. Immobilize
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240 Arnau et al. 6. Collect the flo
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242 Arnau et al. 4. Desalting is ne
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III Various Applications of Affinit
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248 Galaev and Mattiasson in biotec
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Page 510: 17 Monolithic Bioreactors for Macro
Page 514: Monolithic Bioreactors for Macromol
Page 546: 18 Plasmid DNA Purification Via the
Page 552: 278 Forde 12. Sample loading buffer
Page 556: 280 Forde 2. Mix 100 μl of sample,
Page 560: 282 Forde 12. Safety Warning: Picog
Page 564: 19 Affinity Chromatography of Phosp
Page 568: Affinity Chromatography of Phosphor
Page 584: 20 Protein Separation Using Immobil
Page 588: Immobilized Phospholipid Chromatogr
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21 Analysis of Proteins in Solution
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Affinity Capillary Electrophoresis
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Index Adsorption isotherm, 75, 84 A
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Index 341 Genenase I, 170, 214, 215
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Index 343 Saccharomyces cerevisae,
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