The document presents a detailed overview of the novel ADCC reporter bioassays developed by Promega Corporation, which improve upon traditional assays by enabling better quantification of antibody effector functions. These assays utilize engineered cell lines and a frozen, thaw-and-use format to reduce variability and enhance reproducibility in testing. The document outlines the advantages of these bioassays, including their stability, precision, and ability to assess the impact of Fc receptor polymorphisms on therapeutic outcomes.

1. Promega CorporationPromega Corporation
©2013 Promega Corporation.
March 2014
ADCC Reporter Bioassays -
V and F Variants:
Novel, Bioluminescent Cell-Based Assays for Quantifying
Fc Effector Function of Antibodies
Rev 01

2. ©2013 Promega Corporation.
Promega CorporationPromega Corporation 2
Topics Presented
 Introduction to ADCC
– Problem with classic ADCC assays
 Solution: Designing a Better ADCC Bioassay
– Introducing ADCC Reporter Bioassay
 “Cells as Critical Reagents”
– Cell selection, frozen, thaw-and-use format
 V Variant ADCC assay performance
– Assay optimization, qualification, potency & stability
indicating, benchmarking
 F Variant ADCC assay performance
– Assay optimization, qualification, potency & stability Indicating
 Commercial Formats
– Kits & Cell Propagation Model, custom materials
Image source:
www.iconplc.com

3. ©2013 Promega Corporation.
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 Reflective of the mechanism of action (MOA) of the
biological product
 Well controlled (precise, accurate, robust, reproducible)
 Stability-indicating
 Usable as a QC lot-release assay
Modified from Chana Fuchs (DMA/CDER)
An Ideal Bioassay Is…
On the following pages, we demonstrate how the novel ADCC Reporter Bioassay
fulfills each of these parameters

4. ©2013 Promega Corporation.
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Characteristics of a Valid Bioassay Are…
-10 -9 -8 -7 -6 -5
0
5
10
15
20
25
30
35
plate1
plate2
plate3
plate4
Log10 [B1 antibody], g/ml
FoldofInduction
Repeatability
Design:
 Two analysts
 Three days
 Four plates per day
100% vs 50%
100% vs 75%
100% vs 125%
100% vs 150% -10 -9 -8 -7 -6 -5
0
5
10
15
20
25
30
35
100%
50%
150%
Log10 [B1 antibody], g/ml
FoldofInduction
Y=1.026X-5.126
R2=0.995
Relative Potency
Linearity
Log [control antibody], g/ml
Log [control antibody], g/ml
Validation of Analytical
Procedures:
 Accuracy
 Precision:
 Repeatability (intra-assay
precision)
 Intermediate precision (day
to day, analyst to analyst)
 Reproducibility (lab to lab)
 Specificity
 Linearity
 Range
 Robustness
- ICH Guideline Q2 [R1]

5. Promega CorporationPromega Corporation
©2013 Promega Corporation.
March 2014
The Problem with Classic ADCC
Assays

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Classic ADCC Assays Are Limited by
The performance of…
Effector cells:
 PBMCs (peripheral blood
mononuclear cells)
 NK from PBMCs
 NK cell lines
Target cells:
 Load with chromium-51 or Eu
 Monitor cell lysis (LDH, Calcein AM,
GAPDH, CytoTox-Glo™ Assay)
CytoTox-Glo™ Cytotoxicity Assay
Target
cell
Y
Primary
NK effector
cellAntibody
FcgRIIIa
Classic assay principle
Cell lysis

7. Promega CorporationPromega Corporation
©2013 Promega Corporation.
March 2014
Solution: A Better ADCC Bioassay
Introducing ADCC Reporter Bioassay

8. ©2013 Promega Corporation.
Promega CorporationPromega Corporation 8
Image source: Leibson-PJ, Immunity (1997) 6(6): 655-661; doi: 10.1016/S1074-7613(00)80441-0
Luciferase reporter is
readout of pathway
activation state
Target-cell bound Ab binds to FcgRIIIa
(CD16) on effector cell – activating
pathway
Scientific Basis of ADCC Reporter Bioassay
New reporter-based
bioassay measures a
step earlier in the
pathway

9. ©2013 Promega Corporation.
Promega CorporationPromega Corporation 9
Target
cell
Y
= NFAT-RE-luc
FcgRIIIa
(V158 or
F158)
Antibody
Improving Upon the Classic ADCC Bioassay
Effector cell
(engineered
Jurkat)
Reporter-based ADCC bioassay
Glo
Target
cell
Y Primary
NK effector
cellAntibody
FcgRIIIa
Classic ADCC bioassay
Specific signal is from target cell:
 High variability of assay results –
mainly due to primary NK cells
 Spontaneous lysis of target & effector
cells results in high background
 Complex & tedious to perform
Signal is from effector cell:
 Reduced variability - replace NK cells
with genetically engineered stable cell line
 ADCC MOA-based bioassay
 Simple & easy to perform
 Improved bioassay performance with
robust reagents and assay design
Cell lysis
Parekh, BS et al (2012). mAbs, 4(3), 310-318; doi: 10.4161/mabs.19873

10. ©2013 Promega Corporation.
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Why an F Variant Version of the Assay is Needed
Polymorphism in the FcgRIIIa Receptor Impacts ADCC Response
FcyRIIIa
158F/V or F/F 158 V/V
>85% population ~10-15% population
Less efficient Ab
binding and ADCC
More efficient Ab
binding and ADCC
 The FcgRIIIa receptor has a
polymorphism at amino acid 158 with
human genotypes of VV, FV and FF. The
V variant receptor has higher affinity for
Abs. Patients with FV or FF genotype
respond less well to Ab drugs having
ADCC MOA.
 Many new generation mAb drugs are being designed to improve Ab
affinity for the F variant of the receptor, to improve patient response.
 Drug developers need a low variability ADCC bioassay specifically to
measure the new drug potency via the F158 variant of FcgRIIIa, and
regulatory agencies are asking for this information.
See the section beginning on page 34,
“Polymorphism in the FcgRIIIa Receptor”
for information on our newly developed
F Variant ADCC Reporter Bioassay.
V158
or
F158

11. Promega CorporationPromega Corporation
©2013 Promega Corporation.
March 2014
“Cells As Critical Reagents”

12. ©2013 Promega Corporation.
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Cells as Critical Reagents
Traditional cell-based assay using fresh cells from cell culture (1-2 weeks)
Cell culture Cell count, harvest,
prepare for assay
Thaw for
culture
Read plates
New assay format using frozen, thaw-and-use cells: convenience and improved
run-to-run reproducibility
Thaw-and-Use Resuspend in assay
buffer, plate for assay
Read plates
Time: 1-2 weeks
Time: <24hr
See pages 50-51 for
information on a
convenient, low cost
luminometer from
Promega:
GloMax® Discover

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No Cell Culture Required
ADCC Reporter Bioassay features Frozen, Thaw-and-Use Cells
1. Human cell lines (Jurkat, WIL2-S, Raji)
- Developed for immediate thaw-and-use in bioassay
- Designed for good recovery and robust response upon thawing
2. Thaw-and-Use format
- Cell propagation conditions & defined freezing protocol control assay
performance for a consistent bioassay response
- No pre-culturing prior to assay means less variability introduced
- Indefinite storage
- Identical cells in bioassay, day to day
3. Minimizes pre-assay planning, time & labor
- Ample cell banks provide long-term supply
means no
cell culture
required
Thaw-and-Use
cells
+

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Biological Performance Equivalent to Fresh Cells
Fresh cells from continuous culture Frozen, thaw-and-use cells
EC50=3.5ng/ml
FI=34-fold
EC50=2.9ng/ml
FI=41-fold
Assay specifics:
E:T ratio = 6:1; fresh WIL2-S cells as target cells;
Bio-Glo™ reagent; 20hr induction for fresh Jurkat cells
assay; 6hr induction for frozen Jurkat cells assay.

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Frozen, Thaw-and-Use WIL2-S Target Cells Provide
Convenience
Kit Control Ab Rituximab
ADCC Reporter Bioassay response to ADCC Bioassay Control Antibody (left) or Rituximab
(right). The EC50 response using frozen, thaw-and-use WIL2-S cells was 16.8ng/ml for Control Ab,
Anti-CD20. For Rituximab, 1.94ng/ml.
Assay specifics:
E:T ratio = 6:1; 6hr induction; Bio-Glo™ reagent

16. ©2013 Promega Corporation.
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Frozen, Thaw-and-Use Raji Target Cells, Too…
ADCC Reporter Bioassay response to ADCC Bioassay Control Antibody (left) or Rituximab
(right). The EC50 response using frozen, thaw-and-use Raji cells was 59.7ng/ml for Control Ab,
Anti-CD20. For Rituximab, 17.0ng/ml.
Kit Control Ab Rituximab
Assay specifics:
E:T ratio = 6:1; 6hr induction; Bio-Glo™ Reagent

17. ©2013 Promega Corporation.
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Complete QC on Cells Ensures Consistent Results
Production cell batches are rigorously tested:
 STR analysis – cell ID profile (human)
 CO1 analysis (cytochrome oxidase) – test
for presence of species (human and other
potential contaminants)
 Cell doubling time under propagation
conditions
 Mycoplasma (Hoechst and direct culture)
 Sterility
 Cell density
 Cell viability after thaw
 Fill volume
 ADCC Reporter Bioassay (EC50 and fold induction)

18. Promega CorporationPromega Corporation
©2013 Promega Corporation.
March 2014
Performance of ADCC Reporter
Bioassay (V Variant)

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Assessing Critical Assay Parameters
Induction time E:T ratio (constant Effector cell #)
Other parameters tested:
 Assay buffer: serum concentration, use of low IgG serum
 Cell numbers per well
 Pre-plating and incubation time: target cell plating, antibody/target cells incubation
 White flat-, V- or U-bottom assay plates

20. ©2013 Promega Corporation.
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Bioassay Development Using DOE
Target cell / Ab Jurkat cell Target cell
run induction time hr incubation time(mins) plating number (K) plating number (K)
1 5.5 30 75 10
2 5.5 30 75 12.5
3 5.5 30 90 12
4 5.5 30 90 15
5 5.5 45 75 10
6 5.5 45 75 12.5
7 5.5 45 90 12
8 5.5 45 90 15
9 6 30 75 10
10 6 30 75 12.5
11 6 30 90 12
12 6 30 90 15
13 6 45 75 10
14 6 45 75 12.5
15 6 45 90 12
16 6 45 90 15
Variables:
 Induction time
 Target/Ab pre-incubation
 Effector cell number
 Target cell number
Outputs & Results:
Good response (fold induction) = 19-27
Good (low) L-term* values = 0.1-0.2
*L-term is a measure of assay precision around the EC50
determination (log width of the 95% confidence interval around
logEC50)

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 Parallelism and measurement of Potency relative to the reference antibody
 Linearity & Accuracy of observed versus expected potencies across the
desired working range of potencies
 Precision
- intra-assay
- intermediate (inter-assay) precision
 Specificity to show response is dependent on specific antibody and the
presence of target cells and FcgRIIIa on effector cells, and not other
components
 Stability-indicating to show the bioassay is capable of detecting loss of
structural integrity of an antibody
These qualification studies are critical to demonstrate a useful
and effective ADCC bioassay
Useful and Effective ADCC Bioassay Demonstrated
Qualification Studies:

22. ©2013 Promega Corporation.
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V Variant: Able to Measure Potencies of On-Market
mAb Biologic Drugs
Rituximab, anti-CD20, chimeric
IgG1. Approved to treat B-cell
non-Hodgkin lymphoma and
chronic lymphocytic leukemia.
Trastuzumab, anti-HER2,
humanized IgG1. approved to
treat HER2-positive breast
cancer and stomach cancer.
Cetuximab, anti-EGFR,
chimeric IgG1. Approved to treat
colorectal cancer and certain
types of head and neck cancer.
Panitumumab, anti-EGFR,
human IgG2 with NO ADCC.
Three best-selling, on-market mAb biologic drugs that posses ADCC as main MOA
Rituximab tests performed using ADCC Reporter Bioassay, Complete Kit; Trastuzumab and
Cetuximab using Core kit. Both V Variant.

23. ©2013 Promega Corporation.
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Able to Measure Fc Effector Function in ADCC for
TNF Blockers
mTNF
CHO-K1
Target
cells
Y
= NFAT-RE-luc
FcgRIIIa
anti-TNF
engineered
Jurkat
effector cells
Glo
mTNF
Assay specifics:
 Effector cells: ADCC Bioassay Effector Cells,
(V Variant) frozen, thaw-and-use
 Target cells: mTNF CHO-K1 target cells
 E:T ratio: 7.5:1
An engineered CHO-K1 cell line
expressing membrane-bound TNF
was established as target cells.
Infliximab
Adalimumab

24. ©2013 Promega Corporation.
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Understanding Potency Determinations Using
Quantitative Bioassays
Relative Potency: Use a bioassay to establish potency activity of unknown
biologic relative to a reference standard
Relative potency can only be
determined when:
 The upper and lower asymptotes as
well as the slopes of the curves are
not significantly different. Hence the
curves are parallel
 Only the EC50s differ
 Relative potency calculation:
__EC50 Test Sample___
EC50 Reference Sample
y = d +
a - d
1 + (conc/c)b
a
c
b
d
Concentration
Response
Potency (% of Reference)
4-parameter logistic curve fit and potency determination
Reference
Test sample
upper asymptote
slope
lower
asymptote
EC50

25. ©2013 Promega Corporation.
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Assay Qualification Results with WIL2-S Cells
Design:
•Two analysts
• Three days
• Four plates per day
100% vs 50%
100% vs 75%
100% vs 125%
100% vs 150%
Linearity
Representative plate layout
Repeatability
Y=1.026X-5.126
R2=0.995
1 2 3 4 5 6 7 8 9 10 11 12 Plate1
A
B no Ab dilu9 dilu8 dilu7 dilu6 dilu5 dilu4 dilu3 dilu2 dilu1 100%
C no Ab dilu9 dilu8 dilu7 dilu6 dilu5 dilu4 dilu3 dilu2 dilu1 50%
D no Ab dilu9 dilu8 dilu7 dilu6 dilu5 dilu4 dilu3 dilu2 dilu1 100%
E no Ab dilu9 dilu8 dilu7 dilu6 dilu5 dilu4 dilu3 dilu2 dilu1 50%
F no Ab dilu9 dilu8 dilu7 dilu6 dilu5 dilu4 dilu3 dilu2 dilu1 100%
G no Ab dilu9 dilu8 dilu7 dilu6 dilu5 dilu4 dilu3 dilu2 dilu1 50%
H
Precision =average of RSD (%) = 7.3%
Accuracy= average of Recovery(%) = 95.8%
Antibody
Sample
Measured
Potency
(%)
Mean
Potency
(%) SD %
Recovery
(%)
RSD
(%)
day 1 48.5
day 2 50% 45.2 48.9 3.9 97.7 7.9
day 3 52.9
day 1 63.1
day 2 75% 62.9 66.4 5.9 88.5 8.9
day 3 73.2
day 1 112.1
day 2 125% 136.3 123.0 12.3 98.4 10.0
day 3 120.5
day 1 148.8
day 2 150% 150.4 147.6 3.6 98.4 2.4
day 3 143.6
Bioassay uses frozen, thaw-and-use cells for both effector & WIL2-S target cells
Good repeatability, accuracy, precision and linearity were obtained

26. ©2013 Promega Corporation.
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Assay Qualification Results with Raji Cells
Day
Antibody
Sample
Measured
Potency
(%)
Mean
Potency
(%)
SD
(%)
Recovery
(%)
CV
(%)
1 49.9
2 50% 51.3 51 0.7 102 1.4
3 50.5
1 78.9
2 75% 78.8 76 5.11 101 6.7
3 70
1 118.6
2 125% 116.9 117 1.19 94 1
3 116.3
1 143.2
2 150% 142.5 145 3.91 97 2.7
3 149.6
Day
Antibody
Sample
Measured
Potency
(%)
Mean
Potency
(%)
SD
(%)
Recovery
(%)
CV
(%)
1 38.4
2 50% 47.2 41.8 7.2 83.5 17.2
3 33.2
4 48.2
1 59.6
2 75% 70.2 67.4 5.2 89.9 7.7
3 69.3
4 70.5
1 120
2 125% 132.3 129.7 7.5 103.7 5.8
3 137.8
4 128.6
1 160.2
2 150% 158.2 162.8 5.2 108.5 3.2
3 162.7
4 170
Precision: 2.95%
Accuracy (recovery avg):
98.5%
Linearity: y = 0.922x + 5.0
Precision: 8.47%
Accuracy (recovery avg):
96.4%
Linearity: y = 1.22x - 21.3
Analyst 1: Analyst 2:
Linearity:

27. ©2013 Promega Corporation.
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ADCC Reporter Bioassay (V Variant) is Specific
Assay signal is specifically
dependent on:
 Target cells expressing Ab-targeted
antigen
 Specific antibody
 Effector cells expressing FcgRIIIa
receptor

28. ©2013 Promega Corporation.
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ADCC Reporter Bioassay is Robust
-10 -9 -8 -7 -6 -5
0
10
20
30
1
6
7
14
Log10 [B1 antibody], g/ml
FoldofInduction
-10 -9 -8 -7 -6 -5
0
10
20
30
1
1
7
13
16
Log10 [B1 antibody], g/ml
FoldofInduction
Time of induction
E:T ratio and cell # per well
Run Induction time EC50
1 6.0hr 3.15x10-8 g/ml
2 5.5hr 3.83x10-8 g/ml
Run E:T ratio E cell # T cell # EC50
1 7.5:1 75k 10k 3.09x10-8 g/ml
2 6:1 90k 15k 3.83x10-8 g/ml

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Stability Indicating for Fc Effector Function
EC50 = 5.77ng/ml
EC50 = 31.0ng/ml
Rituximab:
Tositumomab:
Trastuzumab:
Activity following heat-treatment of
antibody drugs

30. ©2013 Promega Corporation.
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Miniaturizes to 384-Well Format
WIL2-S target cells Raji target cells
Assay volume per well Target cells Antibody Effector cells Bio-Glo™
96-well plate 25µl 25µl 25µl 75µl
384-well plate 5µl 5µl 5µl 15µl

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©2013 Promega Corporation.
March 2014
Antibody Variants, Glycosylation and
Benchmarking with Classic ADCC Assay

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Deglycosylated Herceptin
-12 -10 -8 -6 -4
0
5100 5
1100 6
2100 6
unt
10%unt/ 90% degly
EC50
unt
2.037e-008
10%unt/ 90% degly
7.174e-008
log [ab], (g/ml)
RLU
Deglycosylated Herceptin
-12 -10 -8 -6 -4
0
5100 5
1100 6
2100 6
unt
100% degly
EC50
unt
1.988e-008
100% degly
3.202e-008
log [ab], (g/ml)
RLU
Deglycosylated Herceptin
-12 -10 -8 -6 -4
0
5100 5
1100 6
2100 6
EC50
unt
1.720e-008
20%unt/80% degly
4.626e-008
unt
20%unt/80% degly
log [ab], (g/ml)
RLU
Deglycosylated Herceptin
-12 -10 -8 -6 -4
0
5100 5
1100 6
2100 6
unt
30%unt/ 70% degly
EC50
unt
2.002e-008
30%unt/ 70% degly
4.153e-008
log [ab], (g/ml)
RLU
Deglycosylated Herceptin
-12 -10 -8 -6 -4
0
5100 5
1100 6
2100 6
unt
40%unt/ 60% degly
EC50
unt
2.082e-008
40%unt/ 60% degly
3.486e-008
log [ab], (g/ml)RLU
Deglycosylated Herceptin
-12 -10 -8 -6 -4
0
5100 5
1100 6
2100 6
unt
50%unt/ 50% degly
EC50
unt
2.110e-008
50%unt/ 50% degly
2.990e-008
log [ab], (g/ml)
RLU
Target cells: SKBR3; Unt = 100% glycosylated
Sensitive to Detect Differences in Glycosylation

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Linear correlation obtained between
percentage of N-glycosylated antibody in
blended antibody samples and relative
luciferase reporter activity in ADCC
Reporter Bioassay
y = 0.0127x - 0.0314
R² = 0.966
-0.100
0.000
0.100
0.200
0.300
0.400
0.500
0.600
0.700
0 10 20 30 40 50 60
RelativeactivityinreporterADCC
Percent N-glycosylated sample
y = 0.0125x - 0.0095
R² = 0.9926
0.000
0.100
0.200
0.300
0.400
0.500
0.600
0.700
0 10 20 30 40 50 60
RelativeactivityinreporterADCC
Percent N-glycosylated sample
Rituximab
Trastuzumab
_____________________________________
Small differences in Fc effector activity in
ADCC pathway activation are easily
distinguished in the ADCC Reporter Bioassay
_____________________________________
Activity Correlates with Amount of Antibody
N-Glycosylation

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Sample set: Blended mixes of fully fucosylated and afucosylated Ab samples – provides a series with a range of
ADCC potencies. Relative activity: Expressed relative to 100% fucosylated Ab (lowest potency of series).
Both assays show increase in relative potency with increased % afucosylation of mAb
WIL2-S target cells; 6hr assay. E:T 6:1LDH release assay; WIL2-S target cells; PBMCs
as effector cells. 4hr assay. E:T 25:1
Benchmarking Study: Good Correlation with Lytic
LDH Release Assay
Classic ADCC assay(1) ADCC Reporter Bioassay(2)
1. Chung, S et al (2012). mAbs, 4(3), 326-340; doi: 10.4161/mabs.19941
2. Cheng, J et al (2012) AACR, poster #4606.

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©2013 Promega Corporation.
March 2014
Polymorphism in the FcgRIIIa Receptor
Optimizing the F Variant of the ADCC Reporter Bioassay

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Effector:Target (E:T) Ratio
Suspension target cells Adherent target cells
10:1
6:1
4:1 15:1
10:1
ADCC Reporter Bioassay,
F Variant

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Induction Time
Fresh-from-culture effector cells Frozen, thaw-and-use effector cells
24hr
7hr
5hr
6hr
24hr
Jurkat cell format
ADCC Reporter Bioassay,
F Variant

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Bioassay Development Using DOE
Factors:
 Target cell density
 Target cell/antibody
incubation time
 Effector cell density
 Induction time
Outputs:
 Fold of induction
 EC50
 EC50 difference between F
and V variant Jurkat
effector cells
DOE = Design of Experiments
 Allow understanding of the
interactions between critical
assay factors
 Minimum amount of work
needed to develop robust
assays
ADCC Reporter Bioassay,
F Variant

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The F Variant Bioassay is Specific
Assay signal is specifically dependent on:
 Target cells expressing Ab-targeted antigen
 Specific antibody
 Effector cells expressing FcgRIIIa receptor
No target
cells
No Ab or
wrong Ab
FcgRIIIa
blocked
No FcgRIIIa

40. ©2013 Promega Corporation.
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F Variant: Able to Measure Potencies of On-Market
mAb Biologic Drugs
Rituximab, anti-CD20,
chimeric IgG1. Approved to
treat B-cell non-Hodgkin
lymphoma and chronic
lymphocytic leukemia.
Trastuzumab, anti-HER2,
humanized IgG1. Approved to
treat HER2-positive breast
cancer and stomach cancer.
Cetuximab, anti-EGFR,
chimeric IgG1. Approved to
treat colorectal cancer and
certain types of head and
neck cancer.
Target cell: SK-BR-3 Target cell: A431
Three best-selling, on-market mAb biologic drugs that posses ADCC as main MOA
Tests performed using ADCC Reporter Bioassay, Core Kit (F Variant)

41. ©2013 Promega Corporation.
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Antibody IgG-Isotype Specificity
Order of response: hu IgG1, IgG3 > mouse
IgG2a >> hu IgG2, IgG4, mouse IgG1
Expanding assay applications:
1. Confirm desired ADCC activity for IgG1
and IgG3-based mAbs
2. Identify non-designed ADCC activity for
mAbs with non-ADCC MOA
2 1
Data generated using the ADCC Reporter
Bioassay, F Variant

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Detects Loss of Activity Due to Heat-Stress
EC50 increases (right shift)
 Reporter bioassay exhibits stability-indicating capability
 Can potentially be used in stability studies for therapeutic antibody
Fold induction
decreases
ADCC Reporter Bioassay,
F Variant

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Polymorphism in the FcgRIIIa Receptor Impacts
ADCC Response
Because many new generation mAb drugs are being designed to improve Ab affinity for
the F variant of the receptor, drug developers need a low variability ADCC bioassay
specifically to measure the new drug potency via the F158 variant of FcgRIIIa.
Rituximab Trastuzumab
The dual receptor bioassay approach of the Promega ADCC Reporter Bioassay, with both V
and F variants of the FcgRIIIa allow researchers to better characterize their Ab drugs.

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©2013 Promega Corporation.
March 2014
Product Formats

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Product Configurations: ADCC Reporter Bioassay,
V Variant
Engineered
Jurkat cells
Bio-Glo™
Reagent
Control
Antibody
Target WIL2-S
or Raji Cells
Core Kit
Complete
Kits
+Medium Serum
1. Core Kits:
1X kit – Cat.# G7010
5X kit – Cat.# G7018
2. Complete Kits:
WIL2-S Target Cells – Cat.# G7014
Raji Target Cells – Cat.# G7015
3. Target Kits:
WIL2-S Target Cells – Cat.# G7013
Raji Target Cells – Cat.# G7016
4. Cell Propagation Model
ADCC Bioassay Effector Cells,
Propagation Model – Cat.# G7102
2 vials of Jurkat Effector cells:
allows propagation and banking for
use in ADCC via unique LULL
Target Kits
Multiple formats flexible to your
research needs:
Engineered
Jurkat cells,
V variant
Propagation
Model

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Custom Product Configurations for ADCC
Reporter Bioassay, F Variant
Engineered Jurkat
cells, F variant
Bio-Glo™
Reagent
Control
Antibody
Target WIL2-S or
Raji Cells
Core Kit
+
Medium Serum
Target Kits
Catalog
products
Jurkat effector
cells, F variant
Propagation
Model
1. Core Kit (Part# CS1324F01)
Components:
 ADCC Bioassay Effector Cells,
F Variant: in frozen, thaw-and-use
format
 RPMI Medium
 Low IgG Serum
 Bio-Glo™ Luciferase Assay System
2. Cell Propagation Model
(Part# CS1324F08)
Components:
 ADCC Bioassay Effector Cells, F Variant,
Propagation Model: allows propagation
and banking for use in ADCC via unique
LULL with Bio-Glo™ Luciferase Assay
System

47. ©2013 Promega Corporation.
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Ordering Information
ADCC Reporter Bioassays (V Variant)
Product Size Cat.
ADCC Reporter Bioassay, Core Kit 1ea G7010
ADCC Reporter Bioassay, Core Kit 5X 1ea G7018
ADCC Reporter Bioassay, Complete Kit (WIL2-S) 1ea G7014
ADCC Reporter Bioassay, Complete Kit (Raji) 1ea G7015
ADCC Reporter Bioassay, Target Kit (WIL2-S) 1ea G7013
ADCC Reporter Bioassay, Target Kit (Raji) 1ea G7016
ADCC Bioassay Effector Cells, Propagation Model 1ea Pls enquire
Contact: COD@promega.com
Bio-Glo™ Luciferase Assay System 100ml
10ml
G7940
G7941
Product Cat.
ADCC Reporter Bioassay, F Variant, Core Kit Pls enquire
ADCC Bioassay Effector Cells, F Variant, Propagation Model Pls enquire
Contact CAS@promega.com
ADCC Reporter Bioassays (F Variant)
Pre-commercial materials are available now through our
Custom Assay Services group. Simply e-mail CAS at the
address below.

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Adopted by the Contract Services Industry
 Approved manufacturing cell line switch by a pharmaceutical company
 Submitted in an IND filing by a pharmaceutical company
 In development for lot-release testing by a pharmaceutical company
 Adopted by multiple pharmaceutical companies globally
 BioAgilytix, Catalent, Covance & Charles River Laboratories are providing
ADCC Reporter Bioassay services

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Highlights of the ADCC Reporter Bioassay…
Features
 Low variability
 Engineered effector cells (Jurkat FcgRIIIa/NFAT-RE luc2)
replace primary NK cells
 “Cells as reagents” - frozen, thaw-and-use format
 Simple & robust protocol
 Broad applicability in use with suspension or adherent
target cells
 Good correlation with classic ADCC bioassay (cytolytic)
Benefits
 Demonstrates precision, accuracy, linearity, robustness,
specificity
 Can quantify potency and stability of therapeutic Ab drugs
 Can differentiate biological activity of Fc effector function
in ADCC MOA resulting from small changes in Ab
glycosylation

50. Promega CorporationPromega Corporation
©2013 Promega Corporation.
March 2014
Simple, Convenient Luminometer
Tested with ADCC Reporter Bioassay

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Easy Reporter Assay Detection
GloMax® Discover System…
Integrated for Promega’s ADCC Reporter Bioassay
And ready to run any of the following reporter, cell
health and protein interaction assays
Product Size Cat.
GloMax® Discover Multimode Detection System 1ea GM3000
Cell Health:
 CellTiter-Glo® Assay
 CellTox™ Green Assay
 Caspase-Glo® Assay
 BacTiter-Glo® Assay
Luciferase Reporters:
 Nano-Glo® Assay
 ONE-Glo™ Assay
 Dual-Glo® & DLR Assays
 Bright-Glo™ Assay
Protein Interaction Assays:
 ELISAs
 BRET
 FRET See www.promega.com/glomax for more
information

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GloMax® Discover System
Easy-To-Use
 Choose from preloaded Promega
protocols or customize your own.
 Build custom protocols using the intuitive
drag-and-drop interface.
 Export data to a network, cloud or any
drive desired.
Superior Sensitivity
 Plate mask (aperture control) for
switching between 96-well and 384-well
plates.
 Low cross-talk between wells gives you
better, more usable results.
 Broad dynamic range of the instrument
extends the linear range of your assay.
Minimal Manual Intervention
 Filter paddles and automatic filter
switching allow easy two-color
multiplexing assays or kinetic studies.
 Quick read-speeds for high efficiencies in
your laboratory.
 Compatible for 3rd party automation
control.
Quality
 IQ/OQ Service available.
 Provides required technical
elements of a 21CFR 11 compliant
system to be used with the
appropriate laboratory workflow.

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For More Information
Neal Cosby, PhD
Strategic Marketing Manager
Neal.cosby@promega.com
Custom Order Department
COD@promega.com
Or see:
www.promega.com/adcc