A
A
A AR RT
T
T
T
Adaptive binding
Allosteric activation
A=Aptamer T=target R=Ribozyme
Apta-switch/beacon“OFF”
Apta-switch/beacon“ON”
“INPUT”Signal
“OUTPUT”Signal
AT
ModularDomains
“Functional”Aptamer
Your PlatformChoice implement
Negative/CounterSelection
5’
T7
PrimerExtension
5’
RNA library
Hammerhead ribozyme motif
N55
N55
5’
Mg++ dependentCleavage site
Transcription
Synthesized N55 random oligo library
Aptamer
pre
clv
PositiveSelection
RT-PCR
PAGEPartitioning
Promoter
Selection
PAGEPartitioning
(+) Target(-) Target(Buffer alone or Counter-target)
LibraryLibrary
Purify Pre-cleaved
Purify Cleaved
Random Region
Fluorophore
Optional:1. RT-PCR2. Transcription3. Refolding
Refolding
Apta-sw
itch Selection S
trategy
Monitoring Enrichment(Rounds of Selection)
(-) + (-) + (-) + (-) +
Target Example
Co2+, Ni2+, Cd2+
Zn2+, Mn2+
Caffeine
Rev Peptide
Phosphorylated ERK2,Unphosphorylted ERK2
Metal Ions
Small Organics
Peptides
Proteins
Apta-switch™ (aptamer that produces a self-cleavage output signal)
FlashGel™analysis
(5 minute run)
Apta-switch™Apta-switch™ Demonstration Kit Demonstration Kit(Theophylline/Caffeine)(Theophylline/Caffeine)
100 M
The
ophy
lline
Specificity Against Theophylline vs. Caffeine
100
90
80
70
60
Ladd
erDEPC o
nly
RXN Buf
fer o
nly
1 m
M C
affe
ine
100 M
Caf
fein
e10
M
Caf
fein
e
10
M T
heop
hyllin
e
1 M
Caf
fein
e1 M
The
ophy
lline
1 minute reaction at 23oC,then stopped with stop buffer containing excess EDTA.
1 m
M T
heop
hyllin
e
500-fold sensitivity range
Caffeine
Theophylline
“Forget Antibodies. Use Aptamers!”
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