“For over 24 years, we’ve worked hand in hand with industry,

academic, and government researchers to help solve difficult

assay development and protein quantification problems.”

Stephen Parent, PhD

Director of Business Development

sparent@pblbio.com

(732) 777-9123 x117

PBL Assay Science History

• Founded in 1990 by Dr. Sidney Pestka

• 35 employees based in Piscataway, NJ

• World’s largest producer of interferons and related cytokine products for the research market

– ELISA and multiplex assay kits

– Recombinant proteins

– Antibodies

– Assay services

– Custom services

High Quality Assay Solutions Interrogating

Interferon and Cytokine Biology

Assay Solutions Interrogating IFN and Cytokine Biology

• Off the Shelf Assays

– More than 70 Interferon ELISAs, antibodies and cell-

based assays

• Assay Services Offerings

– Sample Testing and Screening

– More than 40 Singulex Erenna biomarker assays

– Assay Design and Development

– Custom Services

– We provide ~ 70 different services

Assay Services Capabilities

Cell-Based Assay Development

Antiviral Assays

Cytokine/Chemokine Bioactivity Assays

Immunoassays

Development

Single Analyte & Multiplex ELISAs

Ultrasensitive Cytokine Detection Assays

using Singulex Erenna® technology

More than 40 Assays Available

Human and Mouse

Reagent Development & Characterization

Assay Optimization

Technology Transfer

Consulting in Assay Development and

Therapeutics Development

IL-17

Ultrasensitive Cytokine Detection Service

using Singulex Erenna® Immunoassay System

Modified IA process

using magnetic beads

Single Molecule

Detection (SMD)

technology50- 1000 fold higher sensitivity over existing IA

products

More than 40 Assays Available

IL-17

Singulex Single Molecule Counting (SMC)

Erenna®

4.

5.

► Elution buffer

dissociates the

complex (analyte +

detection antibody

in simple buffer)

► Transfer to 384-

well plate

► Draw eluate into the

instrument for reading

► Fluorescently labeled

antibodies introduced

to laser confocal

microscope digital

counter

► Molecules are

individually detected

and digitized

Elution Molecule counting

Proprietary Singulex technology

Sample collection

2.

3.

► Transfer samples

to 96-well plate

► Incubate with

capture magnetic

antibody-coated

beads

► Wash to remove

unbound

antibodies

► Incubate with dye-

labeled detection

antibodies

1.

► Low abundance of

certain biomarkers in

complex biological

samples

Antibody incubation Washing & fluorescent labeling

Standard immunoassay protocol

More than 40 Assays Available

Examples of Available Erenna® Assays

• IL-1b

• IL-6

• IL-17A

• IL-17F

• IL-17 A/F Heterodimer

• IFNg

• TNFa

• cTnL

• GLP-1

Parameter PBL Singulex Reported

LLOQ

(pg/ml)

0.20

0.10

0.20

Avg = 0.17

0.10

LOD

(pg/ml)

0.040

0.018

0.050

Avg = 0.04

0.03

IL-1b Assay LLOQs and LLODs

Examples of Available Cell-based IFN Assays

Cytopathic Effect (CPE) Inhibition Assay

Neutralization of 1 U of IFN in

cytopathic effect (CPE) inhibition assay

A549 human lung cancer cell line with

encephalomyocarditis virus (EMCV)

Cell-based Neutralizing Antibody Assay

Examples of Cell Proliferation & Anti-Proliferation Assays

OVCAR-3 human adenocarcinoma line TF-1 human erythroleukemia line

Proliferation quantified using soluble tetrazolium reagent

Immunoassay Products

• 10 Human Interferon ELISAs

• 3 Mouse Interferon ELISAs

• 1 Cynomolgus/Rhesus Interferon ELISA

• 1 Human soluble IFNAR2 ELISA

• 2 Multiplex ELISAs quantifying Type I, II and III IFNs

and other cytokines

• Custom Multiplex ELISAs are also possible

You can have confidence in PBL ELISAs!

• Human IFN alpha ELISA study (2008)

• Split samples run at a CRO and PBL

• Slope = 0.991

• Correlation coefficient = 0.9931

• Human IFN beta ELISA study (2009)

• Split samples run at a CRO and PBL

• Slope = 1.036

• Correlation coefficient = 0.9966

• Human IFN beta ELISA study (2014)

• P. Conliffee et al. AAPS Biotechnology Conference

• Validated analytical range 2.34 - 150 pg/ml

• Dilutional linearity up to 1 : 2187

• Interassay precision (CV) ≤13.1%

• LLOQ 2.34 pg/ml

• No IFN-b detected in normal sera.

• Median quantifiable IFN-b was 17.3 pg/ml in MS sera (n=47), with 26

quantifiable & 21 below LLOQ.

• MS samples across different sites showed excellent correlation (r2=0.9975)

VeriKine Human Interferon ELISA Kits

• Human IFNa Multisubtype Serum Kit

• Validated at numerous sites

• Kit standard calibrated to NIH human IFN alpha

standard

• Dual detection range

– 12.5 – 1000 pg/ml

• LLOD: 1.6 pg/ml; LLOQ: 5.7 pg/ml

• Intra-assay, Inter-assay, and Inter-lot CV < 8%

• Compatible with 50% normal human serum, 50%

plasma, TCM and nasal lavage

• Unique broad subtype cross reactivity

• Detects Intron-A®, Roferon®, pegylated interferon alphas

• Human Interferon Beta Serum Kit

• Developed to meet ICH guidelines on validation of

analytical procedures

• Validated at 3 independent labs to date

• Kit standard calibrated to NIH human IFN beta

standard (correlates with NIBSC standard)

• The most sensitive human IFN beta ELISA in the

market

• Detection range: 1.2 - 150 pg/ml

• LLOD: 0.3 pg/ml; LLOQ: ~2 pg/ml

• Intra-assay, Inter-assay, and Inter-lot CV < 8%

• Compatible with normal human serum, plasma,

tissue culture media

• Characterized on MS patient sera

• Detects Betaseron®, Avonex®, Cynomolgus,

Rhesus IFN beta

• Detects IFN beta bound to soluble IFNAR2

receptors

VeriKine-HS High Sensitivity

Human Interferon Beta Serum ELISA Kit

90% of Patients on IFN

Beta therapy have

quantifiable IFN Beta

2% of Healthy Donors have

1-2 pg/ml IFN-Beta

No elevation of IFN Beta

on other therapies (p=0.4)

IFN Beta in Normal and MS Patient Sera

Detected by ELISA

IFN

Bet

a Tre

ated

n=29

No

IFN

Tre

atm

ent n

-25

NH

S n=

193

0102030405060708090

100110120

IFN

-Bet

a (

pg

/m

l)

New Human IFN Alpha All Subtype ELISA Kit Released in 1/2015

41105 41110 New - 41115

DMEM Normal Hu Serum Normal Hu Serum

Subtype LLOQ LLOD LLOQ LLOD LLOQ LLOD

α1 (αD) 59.21 15.07 27.75 27.75 0.63 0.18

α2a (αA) 7.13 2.00 5.72 1.62 0.57 0.16

α4a (αM1) 123.68 39.52 137.14 51.42 0.58 0.16

α5 (αG) 40.63 11.39 58.26 22.34 0.86 0.24

α6 (αK) 10.02 3.14 6.05 1.78 0.61 0.18

α7 (αJ1) 8.57 2.96 6.15 1.97 1.34 0.39

α8 (αB2) 6.84 2.95 3.54 1.08 4.97 1.60

α10 (αC) 10.03 3.60 5.98 1.79 2.69 0.90

α14 (αH) 16.18 5.73 9.73 2.61 4.13 1.36

α16 (αWA) 162.98 49.17 208.54 81.68 2.96 0.90

α17 (αI) 6.06 2.07 4.60 1.36 1.69 0.56

α21 (αF) - - - - 14.93 5.12

500-12.5 pg/mL 1000-12.5 pg/mL 125-1.95 pg/mL

New Assay sees all 12 subtypes well; more sensitive

PBL’s New Human IFNa All Subtype ELISA (Cat. 41115)

is the most sensitive assay kit on the market

Mean LLOD &

LLOQ values on

ALL 12 human

IFN-α subtypes

LLOD & LLOQ values

from IFN-α2 alone;

subtype quantitation

information available

LLOD & LLOQ values from IFN-α2 alone.

No subtype quantitation information available.

*Competitor data obtained from public sources,

provided for comparison only.

0

2

4

6

8

10

12

PBL 41115 PBL 41110 Competitor A Competitor B Competitor C Competitor D(non-ELISA technology)

LLOD (pg/ml) LLOQ (pg/ml)

Interferon Receptor Assays

ELISA assays for soluble Type I and Type II IFN receptors

Soluble Type I and Type II Receptors are elevated in Males

sIFNAR2 ELISA Assay

sIFNAR2 Levels in Autoimmunity

Normal n

=59

SLE n=111

RA n=16

Sjogren's n=11

Scleroderma n=10

0

2500

5000

7500

100001000035000

p=0.008

sIF

NA

R2 p

g/

ml

Reagent Products

• Proteins

– Human Interferons

– Mouse Interferons

– Rat Interferons

– Other Interferons

– Human Cytokines & Growth

Factors expressed in Human Cells

• Polyclonal Antibodies

– PAbs to Human Interferons

– PAbs to Mouse Interferons

– PAbs to Rat Interferons

– Labeled PAbs

• Monoclonal Antibodies

– MAbs to Human Interferons

– MAbs to Mouse Interferons

– MAbs to Rat Interferons

– MAbs to Porcine Interferons

– Labeled MAbs

PBL Assay Science Team

PRaDAS Quality Control

Tom Lavoie Sehrish Ajmal*

Taher Fatakdawala* Karen Zipf*

Robert Niemczyk* Production Team

Xiao-Hong Lin* Daniela Rotaru

Tara Stauffer* Karen Murphy

Michael Skawinski* Rocio Acosta

Lara Izotova* Ksenia Makarenko

Eric Zhu* Natasha Kugakolova

Barbara Schwartz

Nirupama Putcha Collaborators

Sanjna Sanghvi Ray Donnelly (FDA/NIH)

Katherine Carrino-Bednarski Phyllis Conliffe (Smithers Avanza)

Jonathan Ferreira Ira DuBey (Smithers Avanza)