Enhancing cell culture with MACS® Premium-Grade Cytokines

  • Activity-based cytokine dosing allows for reproducible results
  • Calibrated cytokine activity values enable comparable results
  • Optimal cytokine dosing increases reliability and reduces reagent expenses
  • Known lot-specific activities save time and costs needed for batch testing
     

Variations in cell culture caused by inconsistent cytokine activity are a common phenomenon. The biological activity of a recombinant cytokine, i.e., the effect the respective cytokine has on the cells in a specific bioassay, is neither a constant nor an absolute value. 

Read through this page to learn about the pitfalls of varying cytokine activities or dosing and see how you can overcome them with the calibrated lot-specific biological activity of MACS Premium-Grade Cytokines.  

Cytokine activity

Always apply the same amount of active cytokine to get reproducible cell culture conditions.

The biological activity of recombinant cytokines is variable

Only correctly expressed and properly folded cytokines are biologically active and thus will impact the cellular response. Consequently, the total protein amount (protein weight) does not equal the amount of active cytokine (biological activity given in units). 

To receive a reproducible response in cell culture experiments, recombinant cytokines are best applied based on their biological activity instead of the protein weight.

The amount of active cytokine per microgram protein depends on the vendor and production lot

The amount of correctly expressed and folded recombinant proteins depends on the production process. Therefore, the specific biological activity of a cytokine varies between vendors and even between production lots, as shown in the figure below.

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Applying the same microgram amounts of cytokines from different vendors or production lots can result in adding different amounts of active protein.

In these specific examples, weight-based cytokine dosing may result in adding up to twice (or half) the amount of active cytokine, thereby introducing variability in cell culture experiments. 

Dosing according to the biological activity, on the other hand, minimizes those variations and allows for increased cell culture reproducibility.

We provide the lot-specific biological activity

To facilitate convenient cytokine dosing according to the biological activity, we provide the lot-specific activity with our MACS Premium-Grade Cytokines. Knowing the exact biological activity, you can always apply the same amount of active cytokine and create reproducible cell culture conditions.

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Cytokine standardization

Trust in calibrated cytokine activity to maximize comparability of your results.

Cytokine activity is a relative value

To determine a cytokine’s biological activity, the response of cultured cells to different cytokine doses is measured with bioassays. But as the results in bioassays are influenced by many factors, e.g., by applied cells or assay conditions, resulting activity values are relative. Consequently, given (minimal) biological activity values for recombinant cytokines are difficult to compare between different manufactures or even between different production batches from one supplier.

Standardization increases comparability of cytokine activity values

To be able to compare cytokine activity values with each other, bioassays need to be calibrated with a standard with known biological activity. The National Institute for Biological Standards and Control (NIBSC) provides such standards for a large number of cytokines to enable uniformity for calibration. 

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In this example, the biological activity of different cytokine batches was determined in individual experiments using the same bioassay either after calibration with a cytokine standard or without calibration. 

The experiments show that bioassay calibration reduces the variability in biological activity determination, and thus increases the comparability of the resulting unified biological activities, given in calibrated units. Hence, you can easily compare the calibrated activity of production batches of one supplier. 

However, for comparing calibrated activities between different manufactures, you need to take into consideration if the same bioassay was applied. 

Standardization increases assay comparability between different laboratories

When a common cytokine standard is used for assay calibration and the same bioassay is applied to determine the biological activity, cytokine activity becomes comparable between different experiment runs, suppliers, and production batches. This increases the comparability of results from different laboratories as shown in the bar graph below.

We calibrate the cytokine activity for you

With our MACS Premium-Grade Cytokines we provide the lot-specific activity calibrated with NIBSC standards whenever available. This maximizes comparability of your results with other laboratories and other publications. 

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Optimal cytokine dosing

Use optimal cytokine amounts to create reliable results while reducing reagent expenses.

The cellular response depends on the amount of active cytokine applied

Dose-response experiments show how the cellular response in a certain bioassay depends on the amount of active cytokine applied. If cytokine levels are not saturated, small variations in cytokine concentrations can result in notable variations of the measured cellular response. Once cytokine levels are saturated, the dose-response curve reaches a plateau and the maximum cellular response in the respective cell type and culture setting is reached. 

To avoid variable results or artificial effects due to inconsistent or too high cytokine concentrations, optimal cytokine doses are usually titrated for each cell culture assay.

Varying cytokine concentrations influence the cell fate

In cell culture assays, cytokine levels are usually saturated in order to achieve the maximum cellular response. Without knowing the exact cytokine activity, one option to reach saturation is to deliberately add more cytokine than actually needed, creating a state of oversaturation.

However, evidence has increased over the years that artificially high cytokine levels in oversaturated experiments can be harmful for cells. And even if oversaturation is not directly detrimental, it can influence the experimental outcome. The following examples illustrate that the cytokine concentration may determine the cell fate:

  • IL-2 in CAR T cell expansion:
    Kaartinen et al. showed that low IL-2 concentrations favor differentiation of early memory over effector T cells. As memory T cells multiply faster than effector T cells in vivo, excessive IL-2 concentrations can significantly reduce the efficacy of CAR T cell production.
  • GM-CSF in myeloid differentiation:  
    Rautela et al. demonstrated that GM-CSF quantities orchestrate cell fate during myeloid differentiation. Low GM-CSF levels promote the granulocytic lineage, moderate quantities drive moDC differentiation, and high levels stimulate the monocytic lineage. Moreover, monocytes/macrophages generated with different doses of GM-CSF differ in their function.
  • FGF-2 (FGFb) in ESC differentiation:
    Ameri et al. found that differentiation of hESC-derived definitive endoderm into foregut/midgut cell lineage is dependent on the FGF-2 concentration: High FGF-2 levels inhibit hepatocyte differentiation and promote pulmonary cell fate, whereas moderate levels promote pancreatic differentiation.

Efficient cytokine dosing reduces reagent expenses

Moreover, inefficient reagent usage by deliberate overdosing causes a significant cost increase. Knowing the exact cytokine activity, you can refrain from oversaturation, use the optimal amount of active cytokine for your experiment, and save up to 10-fold reagent expenses.

We pave the way for optimal cytokine dosing

With the given lot-specific activity of MACS Premium Grade Cytokines, you can precisely apply cytokines according to their biological activity. In this way, you can generate reproducible cell culture conditions and aim for natural cytokine concentrations. You can avoid possibly harmful oversaturation, artificial experimental results, and at the same time reduce costs.

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Lot-to-lot testing for activity determination

Save your time and money and get directly started with your experiment. We already did the testing.

The biological activity needs to be determined for each production lot of recombinant cytokines

As only correctly expressed and properly folded proteins are biologically active, the biological activity of recombinant cytokines varies between production batches. However, manufactures often only provide the minimal biological activity specifications rather than the exact biological activity of a cytokine batch. 

Consequently, the biological activity of each cytokine batch has to be determined by the end-user with a suitable bioassay and standard. Only then the cytokine activity can be compared to previous lots and the cytokine dosing can be adjusted accordingly.

Lot-to-lot testing is time and cost intensive

The exact expenses of lot-to-lot testing will depend on the respective assay; they include material costs such as cell culture dishes, filter tips and pipettes, cultivation and coating reagents, media and supplements, as well as time to run the assay and manpower conducting and evaluating the assay. 

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This generalized overview illustrates different steps of lot-to-lot testing and indicates estimated expenses in terms of invested time.

We test each production lot for you

With MACS Premium-Grade Cytokines you can skip lot-to-lot testing. We assess the biological activity for each finally filled cytokine batch, calibrate the biological activity whenever possible to NIBSC standards, and provide this calibrated lot-specific activity value to you. Thus, you will save time and reagent expenses and can directly get started with your experiments.

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