applichem © 2010 1/14 there is another top address in darmstadt! applichem cheluminate
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AppliChem © 2010 1/14
There is another top address in Darmstadt!
AppliChem CheLuminate
AppliChem © 2010 2/14
There is another top address in Darmstadt!
• Chemiluminescence detection kits• replaces radioactive assays• extremely sensitive
CheLuminate
AppliChem © 2010 3/14
There is another top address in Darmstadt!
N
N•
NH2 O-
O
NH
NH
NH2 O
O
N
N
NH2 O-
O-
OO
HRP, enhanceraminophthalate + N2 + h
luminol luminol radical anion luminol endoperoxide
catalyst
HO2 HO2
• Horseradish Peroxidase-driven
• Luminol-based
• plus enhancer
• New: plus nucleophilic acylation catalyst
CheLuminate
AppliChem © 2010 4/14
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• Chemiluminescence detection kits
o for ELISA
o for highest sensitivity
o for standard applications(with phenolic enhancer)
- CheLuminate-HRP ELISA FemtoDetect- CheLuminate-HRP ELISA FemtoDetect Plus
- CheLuminate-HRP FemtoDetect- CheLuminate-HRP FemtoDetect Plus- CheLuminate-HRP PicoDetect Extended
- CheLuminate-HRP PicoDetect
CheLuminate
AppliChem © 2010 5/14
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• AppliChem's products:
10-12
10-12
10-14
10-15
CheLuminate
AppliChem © 2010 6/14
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• Sensitivity and stability
Comparison of CheLuminate PicoDetect Extended with a competitor (B)
CheLuminate
AppliChem © 2010 7/14
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• AppliChem CheLuminate – Sales arguments
o competitive price
o highest sensitivity available – from pico (10-12) to femto (10-15) range
o extremely high stability of reagents – up to 24 h signal duration
o compatible with most detection systems / instruments
CheLuminate
AppliChem © 2010 8/14
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• Competitors
o Pierce (Thermo Scientific) Pierce ECL, SuperSignal™ West Pico,SuperSignal™ West Femto
o Millipore Immobilon™ Western HRP
o GE healthcare ECL™
CheLuminate
AppliChem © 2010 9/14
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• Literature
CheLuminate
AppliChem © 2010 10/14
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Bio-/Chemiluminescence• Luciferase System
o Luciferase is one of the most sesnitive reporter in gene expression studies; no radioactivity
o No cellular background activity of luciferase in most assay systems
o Linear activity over a wide range
o Easy to perform
APMBT
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AppliChem © 2010 11/14
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Bio-/Chemiluminescence• Luciferase System with APMBT
o Luciferase from Photinus pyralis (Pluc)o Luciferase from Renilla reniformis (Rluc)
o application of both luciferases in the same assay possible:internal control or investigation of two transcription signals
o This assay is compatible with protein assays!*
o first, measurement of Pluc, then Rluc under Rluc assay conditions, Pluc inactive, because APMBT quenches Pluc
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o APMBT = 2-(4-Aminophenyl)-6-methylbenzothiazole)o * Reference: Hampf, M. & Gossen, M. (2006) Anal. Biochem. 356, 94-99o comparable to Promega‘s DualGlow
AppliChem © 2010 12/14
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Bio-/Chemiluminescence• BCA
o Simple, sensitive photometric assay for proteins. o Faster and easier than Lowry assay.
o and simple buffer salts. More stable at alkaline pH.
o With greater tolerance to interference from non-ionic detergents,Zu bearbeiten
o The principle of the bicinchoninic acid (BCA) assay is similar to the Lowry procedure, in that both rely on the formation of a Cu2+-protein complex under alkaline condition followed by reduction of the Cu2+ to Cu1+.
o The amount of reduction is proportional to the protein present. In the second step, BCA forms a complex with Cu1+, which is purple colored and is detectable at 562 nm.
o Protein + Cu2+ => Cu1+ + BCA => Cu1+-BCA complex o market leader probably Pierce
AppliChem © 2010 13/14
There is another top address in Darmstadt!
Bio-/Chemiluminescence
e.g. Histology Alkaline Phosphatase Detection:
BCIP / NBT Diformazan
e.g. CloningBlue/White Selection:X-Gal / IPTG
e.g. Reporter GenesFirefly Luciferase Detection:
Luciferin Light
e.g. Western Blots Horseradish Peroxidase Detection:
Luminol Light
H2O2
ATP
o substrates & inducers for the detection of
• genes & proteins,• enzyme activities
• Overview
CheLuminate-HRP
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