Recombinant protein activity is highly dependent on the assay performed due to the wide array of variables within a given experimental system. Reference standards provide an effective means of controlling for these variables across assays to determine the relative protein activity. By testing the material in question alongside a reference standard, the specific activity of the protein relative to the reference material can be determined.

 

The specific activity is reported in units (U) or international units (IU) when the reference material is recognized as a World Health Organization (WHO) international standard. Converting to units allows for a comparison of protein activity across multiple vendors and to ensure a consistent level of activity in an experimental system over time.

 

BioLegend often tests our recombinant proteins against commercially available reference standards, including WHO International Reference Standards, when available. Reference standards were obtained from the National Institute for Biological Standards and Control (NIBSC), a WHO collaborating center. The specific activity of these proteins can be found in the table below.

Specific Activity of Recombinant Proteins

 

Human Recombinant Proteins

 

Recombinant Protein NIBSC Reference # BioLegend Specific Activity Cell Type Assay Type
Human GM-CSF 88/646 1.59E+04 IU/μg TF-1 Cell proliferation
Human IL-1α 86/678 3.33E+04 IU/μg D10.G4.1 Cell proliferation
Human IL-1β 86/680 5.63E+04 IU/μg D10.G4.1 Cell proliferation
Human IL-2 86/500 2.36E+04 IU/μg HT-2 Cell proliferation
Human IL-3 91/510 1.64E+03 IU/μg TF-1 Cell proliferation
Human IL-4 (E. coli expressed) 88/656 1.02E+04 IU/μg TF-1 Cell proliferation
Human IL-6 89/548 5.00E+04 IU/μg 7TD1 Cell proliferation
Human IL-12 (p70) 95/544 1.32E+04 Units/μg Human PBMCs IFN-y production
Human IL-15 95/554 3.51E+04 IU/μg MO7e Cell proliferation
Human SCF 91/682 1.13E+03 IU/μg TF-1 Cell proliferation
Human TNF-α 17/232 5.35E+04 IU/μg L929 Cell proliferation
Human VEGF-165 02/286 4.59E+03 IU/μg HUVEC Cell proliferation

 

Mouse Recombinant Proteins

 

Recombinant Protein NIBSC Reference # BioLegend Specific Activity Cell Type Assay Type
Mouse IL-2 93/566 9.07E+04 Units/μg CTLL2 Cell proliferation
Mouse IL-4 91/656 1.54E+04 Units/μg CTLL2 Cell proliferation

 

Frequently Asked Questions About Specific Activity

 

How do I determine the activity in a protein based on reference units?

 

To determine the total activity in an amount of a BioLegend recombinant protein, simply multiply the amount of recombinant protein (μg) by the reported BioLegend Specific Activity (Units/μg) found in the tables above.

 

How is protein biological activity measured?

 

The biological activity of a protein is measured using a bioassay. The assay type is dependent on the protein and may include cell proliferation assays, cytotoxic assays, binding assays, or cytokine production. For each assay, the specific activity is calculated by comparing the measured ED50 (protein concentration required to elicit a 50% maximal response) to a reference standard that was tested side-by-side.

 

Is the specific activity reported in Units (U/µg) the same as International Units (IU/μg)?

 

No. International units are only assigned to international standards provided by the WHO that are manufactured according to WHO guidelines. If the specific activity is reported as U/µg, the assay was not performed using an international standard, but it may still have been tested against a reference standard. Please refer to the table above for more information about which reference standard we use for each protein.

 

Can I compare the specific activity of recombinant proteins from different vendors?

 

Yes. If the two recombinant proteins have been tested in the same assay against the same reference standard, the specific activity of the two proteins can be directly compared. However, if the proteins were not tested against the same reference standard, the activity units may vary depending on the nature of the assay. In these cases, the two proteins would need to be compared side-by-side in the same bioassay.

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