Availability of adequate amounts of the pure allergens is essential to further understand their molecular structures, which is a pre-requisite for the development of more efficacious allergen immunotherapy. The amino acid sequences and coding genes of Hsp70 were retrieved from Uniprot database. The gene of Hsp70 was synthesized and cloned into prokaryotic expression vector pET-28a (+). The expression of His-tagged recombinant Hsp70 in transformant E. coli BL21 was induced by IPTG (final concentration = 1 mmol/L), and the expression products were analyzed by SDS-PAGE. Then, the bacteria were collected and disrupted by sonication. Hsp70 was purified by Ni-NTA protein purification kit (Sangon, C600320-0001), and the recombinant protein was eluted with elution buffer (containing 50mM Tris, 300mM NaCl and 500mM imidazole, Ph 8.0). Then, the endotoxin was removed by Endotoxin Removal Spin Columns (Thermo Fisher, NO. 88273). Purified protein was resuspended in 1×phosphate buffered solution (PBS), and analyzed by SDS-PAGE. The protein concentration was measured by the protein concentration assay kit (Sangon, C503071-0250).
Protein concentration assay kit
Manufactured by Sangon
The Protein Concentration Assay Kit is a reagent-based solution for determining the concentration of proteins in a sample. It utilizes a colorimetric method to quantify the total protein content, providing a reliable and accurate measurement.
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Lab products found in correlation
2 protocols using protein concentration assay kit
1
Recombinant Hsp70 Allergen Purification
2
Recombinant Hsp 70 Protein Production
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Availability of adequate amounts of the pure allergens is essential to further understand their molecular structures, which is a pre-requisite for the development of more e cacious allergen immunotherapy. The amino acid sequences and coding genes of Hsp 70 were retrieved from Uniprot database. The gene of Hsp 70 was synthesized and cloned into prokaryotic expression vector pET-28a (+). The expression of His-tagged recombinant Hsp 70 in transformant E. coli BL21 was induced by IPTG ( nal concentration = 1 mmol/L), and the expression products were analyzed by SDS-PAGE. Then, the bacteria were collected and disrupted by sonication. Hsp 70 was puri ed by Ni-NTA protein puri cation kit (Sangon, C600320-0001), and protein concentration was measured by the protein concentration assay kit (Sangon, C503071-0250).
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