The DHFR2-Fn3 fusion proteins were produced in Escherichia coli (E. coli) and purified from the soluble fraction of the cell lysates by IMAC according to the methods previously reported for the parent Fn3 clones.31 The mSA-DHFR2 fusion proteins produced in E. coli and purified from the insoluble inclusion bodies of the cell lysates according to our previously reported refolding methods.6 , 43 Purified proteins were analyzed by SEC on a Superdex 200 Increase 10/300 gel filtration column (GE Healthcare Life Sciences, Cat: 28990944) in phosphate buffered saline (PBS, pH 7.4) running buffer (Figure S4 ). Fusion protein retention times were compared to those of commercial molecular weight standards (Sigma Aldrich, Cat: MWGF1000-1KT).
Superdex 200 increase 10 300 gel
Manufactured by GE Healthcare
Superdex 200 Increase 10/300 gel is a size-exclusion chromatography medium designed for the separation and purification of proteins, peptides, and other biomolecules. It has a separation range of 10,000 to 600,000 daltons and is suitable for use in a variety of applications, including protein analysis, enzyme purification, and antibody isolation.
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Lab products found in correlation
2 protocols using superdex 200 increase 10 300 gel
1
Purification and Characterization of DHFR-Fusion Proteins
2
Purification and Size-Exclusion Chromatography of TTR
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Purified TTR was
dissolved in PBS buffer, pH 7.4. The dissolved protein was treated
with thrombin protease to cleave the His-tag with overnight incubation
at 4°. The His-tag-cleaved protein was then centrifuged for 30
min at 14,000g using a benchtop microcentrifuge (Eppendorf
Centrifuge 5424g solution was clear and free of any larger aggregates).
Then, 500 μL of concentrated TTR protein solutions was loaded
on a Superdex 200 Increase 10/300 gel filtration column attached to
an AKTA pure (GE Healthcare) and eluted isocratically at 4 °C
in the same buffer with a flow rate of 0.5 mL/min, and 1 ml fractions
were collected, as shown inFigure S7 .
dissolved in PBS buffer, pH 7.4. The dissolved protein was treated
with thrombin protease to cleave the His-tag with overnight incubation
at 4°. The His-tag-cleaved protein was then centrifuged for 30
min at 14,000g using a benchtop microcentrifuge (Eppendorf
Centrifuge 5424g solution was clear and free of any larger aggregates).
Then, 500 μL of concentrated TTR protein solutions was loaded
on a Superdex 200 Increase 10/300 gel filtration column attached to
an AKTA pure (GE Healthcare) and eluted isocratically at 4 °C
in the same buffer with a flow rate of 0.5 mL/min, and 1 ml fractions
were collected, as shown in
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