IL-12 Bioassay

Measure the Ability of Biologics to Activate or Inhibit Cytokine Signals

Interleukin-12 (IL-12) is a member of the IL-12 cytokine family, which includes IL-12, IL-23, IL-27, IL-35 and IL-39. Both IL-12 and IL-23 are composite cytokines, sharing a common IL-12 p40 subunit and IL-12Rβ1 receptor. Cytokine specificity is derived from the unique IL-12 p35 subunit binding to IL-12Rβ2 and the IL-23p19 subunit binding to IL-23R. Both p35 and p40 genes need to be expressed within the same cell to produce the active heterodimer and subsequent IL-12 signaling. IL-12 is secreted following stimulation of phagocytes and dendritic cells by bacteria and other microorganisms.

The IL-12 Bioassay is a bioluminescent cell-based assay designed to measure IL-12 stimulation or inhibition. The IL-12 Bioassay Cells have been engineered to express luc2P in response to IL-12 signaling. When IL-12 binds to IL-12 Bioassay Cells, the receptor transduces intracellular signals resulting in luminescence. The bioluminescent signal is detected and quantified using Bio-Glo™ Reagent.

Assay Components

The IL-12 Bioassay Cells are provided in a thaw-and-use format as cryopreserved cells that can be thawed, plated and used in an assay without the need for cell propagation (Cat.# JA2601, JA2605).

IL-12 Bioassay Cells are also available in a Cell Propagation Model (CPM) format, as cryopreserved cells that can be thawed, propagated and banked for long-term use (Cat.# J3042).

Workflow

Standardized Reagents Improve Results

Thaw-and-use format provides significant time and labor savings over traditional methods while reducing variability.
Kit includes all of the required reagents in standardized formats.
Easily amenable to high-throughput workflows.

Assay Principle

IL-12 Bioassay schematic. — **Representation of the IL-12 Bioassay.** The IL-12 Bioassay consists of a genetically engineered cell line, IL-12 Bioassay Cells. When IL-12 binds to the IL-12 receptor (IL-12R), receptor-mediated pathway signaling induces luminescence that can be detected upon addition of Bio-Glo™ Reagent and quantified with a luminometer. Inhibition of IL-12 binding by anti-IL-12 or anti-IL-12R antibodies results in a decrease in luminescence.

Assay Performance Data

Accuracy and precision are critical to bioassays and the development of biologics. This assay was developed and prequalified according to ICH guidelines for accuracy and precision, giving it the performance characteristics required for many applications of a bioassay used for antibody drug discovery research and development.

Accuracy: Measure of how close results are to a reference or "target" value.

IntraLab Precision: Measure of repeatability from multiple assays of the same homogeneous sample under the same operating conditions over a short time (same analyst; same lab; same day).

Intermediate Precision: Measure of repeatability of multiple assays of the same homogeneous sample with different analysts or equipment or on different days.

Linearity: Ability of the assay to provide measured values that are proportional to the concentration of the sample.

The IL-12 Bioassay Shows Precision, Accuracy and Linearity.

Parameter	Results
Accuracy	% Expected Relative Potency	% Recovery
	50	101.9
	70	99.4
	140	98.7
	200	108.5
Repeatability (% CV)	100% (Reference)	3.99
Intermediate Precision (% CV)		9.6
Linearity (r²)		0.994
Linearity (y = mx + b)		y = 1.096x – 7.15
A 50–200% theoretical potency series of ustekinumab (anti-IL-12) was analyzed in triplicate in three independent experiments performed on three days by two analysts. Bio-Glo™ Reagent was added and luminescence quantified using the GloMax® Discover Detection System. Data were analyzed and relative potencies calculated after parallelism determination using JMP® software. Data were generated using thaw-and-use cells.

The IL-12 Bioassay responds to recombinant IL-12.

The IL-12 Bioassay indicates stability.

Product Kit Formats

Kits are available in 1X and 5X sizes.

Protocols

Complete Protocol

IL-12 Bioassay, Propagation Model Technical ManualPDF (1832 KB) – English

IL-12 Bioassay Technical ManualPDF (1131 KB) – English

What's in the box?

Item	Part #	Size
IL-12 Bioassay Cells (CPM)	J304A	2 × 0.65ml

SDS

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Certificate of Analysis

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Use Restrictions

Not For Medical Diagnostic Use.

Storage Conditions

Please see the Protocol for detailed storage conditions.

Patents and Disclaimers

NOT FOR MEDICAL DIAGNOSTIC USE. FOR IN VITRO USE ONLY. BY USE OF THIS PRODUCT, RECIPIENT AGREES TO BE BOUND BY THE TERMS OF THIS LIMITED USE STATEMENT. If the recipient is not willing to accept the conditions of this limited use statement, and the product is unused, Promega will accept return of the unused product and provide the recipient with a full refund.

This product may not be further sold or transferred by the recipient and may be used only by the recipient, and then only for (1) research use, (2) discovery, development and monitoring of biologic drugs and vaccines, (3) quality assurance testing of biologic drugs and vaccines, and (4) product release assays for biologic drugs and vaccines. No other commercial use is allowed. "Commercial use" means any and all uses of this product by recipient for monetary or other consideration, including providing a service, information or data to unaffiliated third parties, and resale of this product for any use. Recipient has no right to modify, derivatize, genetically engineer or otherwise create variations of the cells or genes stably transfected within the cells except that recipient may propagate and store the cells for long-term use. In addition, recipient must use Bio-Glo™ Luciferase Assay System purchased from Promega Corporation for all luminescence assays using this product or contact Promega to obtain a license for use of this product with reagents other than Promega's. PROMEGA MAKES NO REPRESENTATIONS OR WARRANTIES OF ANY KIND, EITHER EXPRESSED OR IMPLIED, INCLUDING AS TO MERCHANTABILITY OR FITNESS FOR A PARTICULAR PURPOSE WITH REGARDS TO THIS PRODUCT. The terms of this agreement shall be governed under the laws of the State of Wisconsin, USA.

Cells were obtained under license from AdVec Inc.

What's in the box?

Item	Part #	Size
IL-12 Bioassay Cells	J305A	1 × 0.65ml
Fetal Bovine Serum	J121A	1 × 4ml
RPMI 1640 Medium	G708A	1 × 36ml
Bio-Glo™ Luciferase Assay Buffer	G719A	1 × 10ml
Bio-Glo™ Luciferase Assay Substrate	G720A	1 × 1 vial

SDS

Search for SDS

Certificate of Analysis

Search by lot number

Use Restrictions

Not For Medical Diagnostic Use.

Storage Conditions

Please see the Protocol for detailed storage conditions.

Patents and Disclaimers

NOT FOR MEDICAL DIAGNOSTIC USE. FOR IN VITRO USE ONLY. BY USE OF THIS PRODUCT, RECIPIENT AGREES TO BE BOUND BY THE TERMS OF THIS LIMITED USE STATEMENT. If the recipient is not willing to accept the conditions of this limited use statement, and the product is unused, Promega will accept return of the unused product and provide the recipient with a full refund.

This product may not be further sold or transferred by the recipient and may be used only by the recipient, and then only for (1) research use, (2) discovery, development and monitoring of biologic drugs and vaccines, (3) quality assurance testing of biologic drugs and vaccines, and (4) product release assays for biologic drugs and vaccines. No other commercial use is allowed. "Commercial use” means any and all uses of this product by recipient for monetary or other consideration, including providing a service, information or data to unaffiliated third parties, and resale of this product for any use. Recipient has no right to propagate, modify, derivatize, genetically engineer or otherwise create variations of the cells or genes stably transfected within the cells. In addition, recipient must use Bio-Glo™ Luciferase Assay System purchased from Promega Corporation for all luminescence assays using this product or contact Promega to obtain a license for use of this product with reagents other than Promega’s. PROMEGA MAKES NO REPRESENTATIONS OR WARRANTIES OF ANY KIND, EITHER EXPRESSED OR IMPLIED, INCLUDING AS TO MERCHANTABILITY OR FITNESS FOR A PARTICULAR PURPOSE WITH REGARDS TO THIS PRODUCT. The terms of this agreement shall be governed under the laws of the State of Wisconsin, USA.

Cells were obtained under license from AdVec Inc.

What's in the box?

Item	Part #	Size
IL-12 Bioassay Cells	J305A	5 × 0.65ml
Fetal Bovine Serum	J121A	5 × 4ml
RPMI 1640 Medium	G708A	5 × 36ml
Bio-Glo™ Luciferase Assay Buffer	G719A	5 × 10ml
Bio-Glo™ Luciferase Assay Substrate	G720A	5 × 1 vial

SDS

Search for SDS

Certificate of Analysis

Search by lot number

Use Restrictions

Not For Medical Diagnostic Use.

Storage Conditions

Please see the Protocol for detailed storage conditions.

Patents and Disclaimers

NOT FOR MEDICAL DIAGNOSTIC USE. FOR IN VITRO USE ONLY. BY USE OF THIS PRODUCT, RECIPIENT AGREES TO BE BOUND BY THE TERMS OF THIS LIMITED USE STATEMENT. If the recipient is not willing to accept the conditions of this limited use statement, and the product is unused, Promega will accept return of the unused product and provide the recipient with a full refund.

This product may not be further sold or transferred by the recipient and may be used only by the recipient, and then only for (1) research use, (2) discovery, development and monitoring of biologic drugs and vaccines, (3) quality assurance testing of biologic drugs and vaccines, and (4) product release assays for biologic drugs and vaccines. No other commercial use is allowed. "Commercial use” means any and all uses of this product by recipient for monetary or other consideration, including providing a service, information or data to unaffiliated third parties, and resale of this product for any use. Recipient has no right to propagate, modify, derivatize, genetically engineer or otherwise create variations of the cells or genes stably transfected within the cells. In addition, recipient must use Bio-Glo™ Luciferase Assay System purchased from Promega Corporation for all luminescence assays using this product or contact Promega to obtain a license for use of this product with reagents other than Promega’s. PROMEGA MAKES NO REPRESENTATIONS OR WARRANTIES OF ANY KIND, EITHER EXPRESSED OR IMPLIED, INCLUDING AS TO MERCHANTABILITY OR FITNESS FOR A PARTICULAR PURPOSE WITH REGARDS TO THIS PRODUCT. The terms of this agreement shall be governed under the laws of the State of Wisconsin, USA.

Cells were obtained under license from AdVec Inc.

Frequently Used With

GloMax® Discover System

High-performance microplate reader for detecting luminescence, fluorescence and absorbance.

GM3000

Bio-Glo™ Luciferase Assay System

Bioluminescent detection reagent qualified for use with bioassays that utilize firefly luciferase.

G7940, G7941

IL-12 Bioassay

Part Numbers: J3042, JA2601, JA2605

A Simpler, More Consistent IL-12 Bioassay