By Senta Reichelt (eds.)
The goal of this variation is to introduce the newbie to the fundamentals of affinity chromatography and supply sensible wisdom for the advance of affinity separation protocols. Affinity Chromatography: equipment and Protocols, 3rd Edition publications readers via new state-of-the-art protocols, molecular modelling, and the examine of ligand-target interactions. Written within the profitable Methods in Molecular Biology sequence layout, chapters comprise introductions to their respective themes, lists of the required fabrics and reagents, step by step, easily reproducible protocols, and notes on troubleshooting and keeping off recognized pitfalls.
Authoritative and simply accessible, Affinity Chromatography: tools and Protocols, 3rd Edition is designed as an invaluable source for these drawn to the quick and quantitative isolation of biomolecules with excessive purity.
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The purpose of this variation is to introduce the newbie to the fundamentals of affinity chromatography and supply functional wisdom for the improvement of affinity separation protocols. Affinity Chromatography: tools and Protocols, 3rd version courses readers via new cutting-edge protocols, molecular modelling, and the examine of ligand-target interactions.
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Extra resources for Affinity Chromatography: Methods and Protocols
1 Activation of Sepharose CL-6B 1. Place 100 mL of Sepharose CL-6B (settled gel) (see Note 5) into a sintered glass funnel and wash it sequentially with 500 mL of water/acetone (70/30 (v/v)), 500 mL of water/acetone (30/70 (v/v)) and, finally, with 1 L of acetone (see Note 6). 2. Suspend the gel in 100 mL of acetone, in a 250 mL roundbottom flask, and heat to 40 C (see Note 7), under very gentle stirring (see Note 8). 3. 0 M solution of N,N-diisopropylethylamine in acetone to the Sepharose gel (see Note 9) and, 30 min.
Taylor and Francis, Boca Raton, Chapter 8 67. Liu XC, Scouten WH (2000) Boronate affinity chromatography. In: Bailon P, Ehrlich GK, Fung WJ, Berthold W (eds) Affinity chromatography. Humana Press, Totowa, Chapter 12 68. Roming TS, Bell C, Drolet DW (1999) Aptamer affinity chromatography: combinatorial chemistry applied to protein purification. J Chromatogr B Biomed Sci Appl 731:275–284 69. Huang PY, Carbonell RG (2000) Affinity chromatographic screening of soluble combinatorial peptide libraries.
Transfer ~ 4 mL 50 % agarose resin slurry to a 20 mL plastic column. Allow storage buffer to drain until it reaches the top of the agarose bed (see Note 6). 3. Apply equilibration buffer to the agarose. Allow buffer to drain until it reaches the top of the agarose bed (see Note 7). Wash the agarose with ~ five times of the agarose bed volume (5 BV) of equilibration buffer; allow buffer to drain until it reaches the top of the agarose bed. Plug the column tip to stop buffer flow. 4. Add ~10 mL of cell lysate to plugged column and then cap column.
Affinity Chromatography: Methods and Protocols by Senta Reichelt (eds.)