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Screening a large number of recombinant clones for the production of soluble target proteins can be time consuming and laborious. Sigma’s patented HIS-Select® iLAP® (Integrated Lysis and Affinity Purification) column and plate technology meets the need of researchers who screen large numbers of soluble HIS-tagged proteins by combining cell lysis and HIS-tagged protein purification in one-step.
- One-Step cell lysis and HIS-tagged protein purification
- Efficient cell lysis without harvesting cells from culture
- Non-charged, hydrophilic linkage reduces non-specific protein binding
- Ideal for rapid colony screening
Available in two convenient formats:
HIS-Select iLAP 5 mL column
HIS-Select iLAP 96-well plate
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Figure 1: Comparison of iLAP Purification with Traditional Methods. By using the HIS-Select iLAP 5 mL Column, the number of sample handling steps is greatly reduced, resulting in significant timesavings. Additionally, the procedure does not require the purchase of additional reagents or equipment. 96-well plate format also available. |
HIS-Select iLAP 5 mL column (H9913—Order Now)
The HIS-Select iLAP Columns are single-use, disposable columns designed for the one-step purification of histidine-tagged proteins directly from a 5 mL bacterial culture. Each column contains nickel chelate resin for the purification, and five lysis reagent tablets, which include all the necessary detergents and enzymes needed for efficient cell lysis and protein extraction thereby eliminating the need to harvest cells from culture.
- Patented technology enables robust and efficient cell lysis and histidine-tagged protein capture.
- In less than one hour, purify at least 1 mg of histidine-tagged protein directly from 5 mL of bacterial culture.
- Purified protein is suitable for use with many downstream applications including protein assays, SDS-PAGE, Western blotting, and MALDI.
| Efficient and Selective: High Purity and High Yield |
GST-MAT iLAP Purification |
Lane |
Sample Added |
µl |
1 |
Wide Range MW Marker (M4038) |
3.5 |
2 |
BL21 E. coli TB culture expressing GST-MAT |
10 |
3 |
iLAP Column Flow Through |
10 |
4 |
iLAP Column Wash |
10 |
5 |
iLAP Column Elution |
10 |
| Total protein in Elution = 2.52 mg |
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Figure 2: Recombinant GST-MAT (metal affinity tag) was expressed in BL21 E. coli cells. Five milliliters of bacterial culture were added directly to the HIS-Select iLAP column. The column was incubated, with mixing, for 15 minutes at room temperature. The resin was allowed to settle by gravity flow, and was washed twice with 2 ml of wash buffer (300 mM NaCl, 50 mM sodium phosphate, pH 8.0, 5 mM imidazole). Purified protein was eluted with 2 ml of elution buffer (300 mM NaCl, 50 mM sodium phosphate, pH 8.0, 200 mM imidazole). Total protein was quantitated using the Bradford assay.
HIS-Select iLAP 96-well plate (H9412—Order Now)
The iLAP 96-well clear plate is coated with a proprietary high-density nickel chelate matrix. This matrix provides high-capacity affinity binding of recombinant fusion proteins with histidine-containing tags. An additional coating contains all of the reagents and chemicals necessary for the lysis of bacterial cells for the purification of recombinant proteins along with protease inhibitors to help prevent the proteolytic breakdown of proteins.
- Add bacterial culture directly to the plate for simultaneous cell lysis and histidine-tagged protein capture
- Binds greater than 4 μm of HIS-tagged protein per well
- Efficiently confirm expression of histidine-tagged protein, detect protein-protein interactions, or rapidly screen multiple constructs
- Purified protein is suitable for use with many downstream applications including protein assays, SDS-PAGE, Western blotting, and MALDI
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