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Gel Filtration in Insulin Preparation

Gel Filtration in Insulin Preparation. Presented to: CHEE 450 April 1 st , 2009 Samina Dhalla, Robert Macnamara and Terri Semler. Presentation Overview. Background Design Conclusion Questions. Background. Background.

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Gel Filtration in Insulin Preparation

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  1. Gel Filtration in Insulin Preparation Presented to: CHEE 450 April 1st, 2009 Samina Dhalla, Robert Macnamara and Terri Semler

  2. PresentationOverview • Background • Design • Conclusion • Questions

  3. Background

  4. Background • Gel filtration is used in this biological process to separate insulin from unwanted components • Gel filtration is used to separate molecules based on size • Two methods of separation: • Group separations • High-resolution fractionation http://www.siumed.edu/~bbartholomew/images/chapter6/F06-09.jpg

  5. Design

  6. Loading Sample

  7. Gel Media • Consists of porous matrices composed of spherical particles • Size • Packing density • Porosity • Chemical and physical stability • Factors to be considered: • Resolution desired • Molecular weights of the target proteins and contaminants to be separated • Rate and scale of filtration http://www5.gelifesciences.com/aptrix/upp00919.nsf/4a0f132842ea4d354a25685d0011fa04/50c849d0d5b16ba0c1256e92003e865b/$FILE/GE_Gel%20Filtration.swf

  8. Types of Gels • Bio-Gel P-4 was chosen

  9. Column Dimensions • Design Considerations: • Maximum height for column is 100cm before gel will be crushed • Sample volume injected should be 25% of the column volume • Specifications From SuperPro • Approach Velocity of 300 cm/hr • Total feed of 14,110 L in 20 cycles through filter

  10. Recommended Column Parameters are based on a total volume per batch of 14,110 L with 20 loads per batch being processed through the column

  11. Conclusion

  12. Conclusion • Decided on Bio-Gel P-4 manufactured by Bio-Rad • Using three columns stacked consecutively on one another manufactured by GE Health • Total elution time of 4.23 hours

  13. references

  14. References Amersham Biosciences. “Gel Filtration – Principles and Methods”, Sweden (2002), pp 1-120. Belter, P., Cussler, E., and Hu, W. “Elution Chromatography” in “Bioseparations – Downstream Processing for Biotechnology”, United States (1988), pp 181 – 188. Bio-Rad Laboratories. “Bio-Gel P Polyacrylamide Gel, Instruction Manual”, California, U.S. (2000), pp 1 -14. Sigma-Aldrich (2009). Retrieved on March 23, 2009 from: http://www.sigmaaldrich.com/canada-english.html

  15. Questions ?

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