Lateral flow assays are the most robust, mature immuno sensor available today. Performance in some applications has historically been limited by difficulties in multiplexing and quantification. Novel approaches have been developed and commercialized in recent years that allow for the development and manufacturing of highly multiplexed arrays in lateral flow assays. The patented Symbolics (tm) approach is one such methodology. Symboics (tm) allows for the creation of arrays in lateral flow fields that develop evenly, allowing in turn for the creation of highly complex features such as letters and symbols and also allows for creation of multiplex assays with advanced features such as internal controls. This presentation introduces the principles of multiplexed arraying and the Symbolics technology

1. Patented technology facilitating multiplexing, quantification
and alpha-numeric result generation in rapid assays
Patented
www.Symbolicsdx.com

2. Intuitive Results, Better Results
SymbolicsTM
pixilation technology allows for new approaches to
result generation in lateral flow, including:
1. Geometric symbols
2. Alpha -numeric symbols.
3. Multiplexing (spot arrays or other formats)
4. Other advanced design features

3. Background
• Symbolics LLC was founded in 2010 as a joint venture between
DCN Inc and BioDot Inc
• Wholly owned by DCN and BioDot
• Key management and joint development team in place
• Initial patent allowed, January 2013
• Subsequent filings in progress

4. The Problem
• Lateral flow assays are limited to linear results,
created by the placement of lines of capture
reagents across the width of a flow path
• User Error: Linear results are non-intuitive
and can be difficult to interpret
• Multiplexing and quantification are hard:
– Lines are stacked behind one another
– Line bleed occurs
– Cross reactivity issues are worsened
– Flow rate decreases non-linearly with
distance from origin: quantification of
multiplexed lines is difficult and slow
The Problem with Lines

5. The Problem
Traditional manufacturing methods create
large structures in flow paths
Large, discreet linear features cause flow
pattern disturbances, preventing formation
of features behind them over relatively large
distances in the direction of flow.
High AffinityLow AffinityNo Affinity
Creating other types of features using standard production methods
results in imperfect feature geometry
Strong leading edge binding due to high affinity binding reagents is
extreme in drop sizes used in standard approaches. Reagents that are
useful for lateral flow (with decent affinity) will not form complete
features if spotted on a membrane in standard spot sizes

6. In Other Words….
• The industry has stuck with lines because they are the only way
to produce even, reproducible features in a lateral flow path.
• Alternatives currently in market, such as the “+” symbol rely on
pre-printed ink features to create part of the symbol
• Digital devices returning easily interpreted alpha-numeric results
such as “Pregnant” require expensive, patented reader
technologies in the cassettes, resulting in complexity and margin
hits for manufacturers
Pre-printed ink line

7. The Unseen Impact of Linear Results
• Limited success of lateral flow systems in multiplexed or quantitative
applications
• Limitations on user-friendliness of consumer assays
• Limitation in overall market growth
Average 7% CAGR does not account for the growing potential in
consumer markets, for multiplexed applications such as Traumatic
Brain Injury, Kidney, Cardiac or Dehydration panels, or for other novel
quantitative applications: There is a lot of untapped potential.

8. The Solution: Pixelation
• The problems of incomplete feature formation can be overcome through the
use of appropriately sized features (“pixels”) spaced at appropriate distances in
the flow path.
• This makes the formation of larger, complex features possible.
• Pixels are created using precise methods for dispensing and spacing discrete
spots of protein in the lateral flow field. Volumes range from the picoliter range
to the low nanoliter range at carefully controlled pitch (spacing).
• Dispensing is done using a variety of technologies including piezo-electric or
solenoid-based dispensers.

9. Intuitive Results, Better Results
SymbolicsTM
pixilation technology allows for new approaches to
result generation in lateral flow, including:
1. Geometric symbols
2. Alpha -numeric symbols.
3. Multiplexing (spot arrays or other formats)
4. Other advanced design features

10. The Opportunity
• Revolutionize entire rapid testing market segments by
facilitating the development and manufacturing of novel lateral
flow assays with a range of advantages:
– A new generation of user-centered consumer assays that
are intuitive, simple to interpret but do not require digital
interpretation
– Better quantification
– Higher dynamic range
– Internal controls
– Multiplexing of analytes, at virtually any density required for
diagnostics or testing in most market segments (high or low)
– Lower reagent usage

11. Where we are
• 1 issued US patent: 13/343,681: “Lateral flow assays using two
dimensional features”
• 1 published PCT: PCT/US2012/021586
• Patent portfolio expansion underway with multiple US and
international filings
• Licensing strategy in place
• Licenses and development support available

12. Offering a complete solution or a simple license
*Licenses are available without
using support services offered by
DCN or equipment offered by
BioDot

13. Getting Started
• Symbolics technology is available for license and integration into
your assay development programs
• DCN and BioDot are available if so desired to support your assay
development or re-formatting program

14. Licensing
A two level license is offered
1.Development License
1. Non exclusive
2. Low cost
3. Limited term
4. Patent grant-back
2.Manufacturing and Commercialization License
1. To be negotiated

15. Technical Notes
Facilitating multiplexing, quantification and non-traditional
result generation in rapid assays
Patented

16. Assay Development Strategy
• Assay development follows essentially the same process as
standard lateral flow
• Development requires higher fidelity dispensing equipment
• All of the same labels, reagents and many of the same materials
can be applied
• For higher fidelity quantitative applications, some alternative
material choices may yield better results
• An understanding of the affinity of the reagents being used is
useful given the interaction between reagent affinity and
feature spacing and size
• Assistance can be provided by DCN Inc if required

17. Manufacturing
• Manufacturing process design is similar to standard lateral flow
• Dispensing processes require higher fidelity in terms of
placement and volume
• Dispensing processes must be robust to ensure fidelity of
feature formation
• Consideration must be given to feature position during cutting,
so tracking features (fiducials) must be included in the card
design
• Virtually all other processes will stay the same as in standard
lateral flow
• Equipment can be provided by BioDot Inc if required

18. Table top equipment for R&D and batch manufacturing
Reel-to-reel equipment for high volume production

19. Cutter with Leading Edge and Fiducial Sensors
CARD TARGET
SENSOR

20. Demonstration Applications
1. Geometric symbols
2. Alpha -numeric symbols.
3. Multiplexing.
4. Other advanced design features

21. 1. Geometric Symbols

22. 1. Geometric Symbols
Sample Application: hCG
DropVol: 0.3nL
Pitch: 0.25mm
5mm Strips
Piezo electric
dispenser
Control: 0.5mg/ml
GAM
Test: 1mg/ml anti-
hCGα
Conjugate: 8ug/ml
anti-hCGβ
Negative: 100uL 0.1%
Tween-20 in 1XPBS
10mIU/mL hCG in bufferBuffer only
anti-hCGαGAM

23. 2. Alpha – Numeric Symbols

24. 2. Alpha – Numeric Symbols
Sandwich Applications
Drop Vol: 7nL
Pitch: 0.5mm
6mm Strips
BioJet Plus
CON: 0.125mg/ml protein A
H1: 4mg/ml GP041
H2: 2mg/ml GP036
Conjugate: 10 uL of 8ug/ml
protein A
Sample: 5uL human plasma positive
for HIV-1, 100uL HIV Running Buffer
Sample: 5uL human plasma positive
for HIV-2, 100uL HIV Running Buffer

25. 2. Alpha – Numeric Symbols
Sandwich Applications

26. 2. Alpha – Numeric Symbols in Competitive Assays
DropVol: 7nL. Pitch: 0.5mm. 6mm Strips
Dispenser: BioJet Plus
CON: 0.5mg/ml GAM
COC: 0.5mg/ml Benzoylecgonine-BTG Conjugate
AMP: 0.5mg/ml Amphetamine-BSA Conjugate
Conjugates: 10ug/ml anti-Benzoylecgonine & 3.5ug/ml anti-Amphetamine
Sample: 100uL PBS+
Positive Sample: 500ng/ml benzoylecgonine or amphetamine

27. Drop Vol: 0.3nL Pitch: 0.25mm 5mm Strips
Piezo electric dispenser
Control: 0.5mg/ml GAM
C: 0.5mg/ml Benzoylecgonine-BTG Conjugate
A: 0.5mg/ml Amphetamine-BSA Conjugate
Conjugates: 10ug/ml anti-Benzoylecgonine & 3.5ug/ml anti-Amphetamine
Negative Sample: 100uL PBS+ Positive Sample: 500ng/ml benzoylecgonine &
500ng/ml amphetamine in buffer
2. Alpha – Numeric Symbols in Competitive Assays

28. 3. Multiplexed Lateral Flow Arrays
• Dispenser: BioDot AD2000 Piezo
• Drop Size: 2nL
• Pitch: 1mm
• Capture reagents:
• Control: Goat anti-Mouse
• Test: Amphetamine-BSA, Morphine-
BSA
• Detectors:
• Mouse anti-Morphine Colloidal Gold
Conjugate
• Mouse anti-Amphetamine Colloidal
Gold Conjugate
• Running Buffer:
• 1XPBS, 0.1% BSA, 0.01% Tween-20
• Positive Controls:
• 100ng/ml Morpine in running buffer
• 10ug/ml Ampetamine in running buffer

29. 4. Advanced Application Concepts
• Advanced internal control features
• Flow controls for quantification
• Internal cross reactivity controls and true negatives
• Reader calibration features
• Large scale multiplexing through lateral flow arrays
• Novel quantification approaches
- “thermometer” quantification
- large dynamic range assays

30. DropVol: 0.3nL Pitch: 0.25mm 5mm Strips
Piezo electric Dispenser
Control: 0.5mg/ml GAM
Test: 1mg/ml anti-hCGα
Conjugate: 8ug/ml anti-hCGβ
Sample: 100uL 0.1% Tween-20 in 1XPBS
100mIU/mL hCG in buffer10mIU/mL hCG in buffer 1000mIU/mL hCG in buffer
4. Advanced Application Concepts
i. Analyte depletion assays