G25脱盐柱使用指南讲解学习.pptVIP

Desalting column protocol Mr. Yu Wang Matrix Sephadex? G-25 Fine, cross-linked dextran Bed volume 53 ml Bed height 100 mm i.d. 26 mm separation of high (Mr 5 000) from low molecular weight substances (Mr 1 000). Storage +4 to +30 °20% ethanol 长期不用后再使用，应该用5CV平衡柱子 （5*60=300ml buffer） Try these conditions first Buffer(35ms/cm): 50 mM phosphate buffer, 0.15 M NaCl, pH 7.0 Flow rate: 20 ml/min at room temperature Sample volume: 15 ml Separations: Elute the column with 106 ml (2 CV) of buffer Equilibration is not necessary between runs with the same buffer. (去除气泡）Reverse the flow direction and pump 200 ml well de-gassed buffer at a flow rate of 15 ml/min through the column. The quality of the packed bed will not normally be affected. Sephadex G-10 is well suited for the separation of biomolecules such as peptides (Mr 700) from smaller molecules (Mr 100). Sephadex G-50 is suitable for the separation of molecules Mr 30 000 from molecules Mr 1 500 such as labeled protein or DNA from unconjugated dyes. The medium is often used to remove small nucleotides from longer chain nucleic acids. Sephadex G-25 is recommended for the majority of group separations involving globular proteins. These media are excellent for removing salt and other small contaminants away from molecules that are greater than Mr 5000.