Navigation Links
Fluorometric Protease Assays in the SpectraMax Gemini Microplate Spectrofluorometer: Example Using Caspase3 (MaxLine Application Note #35)

loss in product fluorescent intensity. Figure 3 shows the fluorescence emission (at 441 nm) of the AMC product in the presence of graded concentrations of substrate and with excitation wavelengths ranging from 345 nm to 370 nm.

The quenching effect was especially severe at excitation wavelengths 345 nm to 355 nm, and was still appreciable at 360 nm. However, it was essentially eliminated at excitation wavelengths greater than 364 nm. As the excitation wavelength increased, the fluorescence of the product predictably decreased. At an excitation wavelength of 368 nm, however, AMC fluorescence was decreased by only 40%.


Recommended instrument settings, choice of microplates, and estimated assay sensitivity
Based on the results shown in Figure 3, 368 nm was selected as the excitation wavelength. The next step was to select the emission wavelength and cutoff filter. To help in the selection process, (signal-background)/background1/2 was plotted versus wavelength. The plots were prepared from scans with no emission cutoff filters, with a 420 nm cutoff filter and a 435 nm cutoff filter (Figure 4.) The optimum emission wavelength appeared to be approximately 467 nm. The cutoff filters gave comparable results in that region and both were somewhat better than no cutoff filter. This was confirmed experimentally by running the enzyme assay and blanks at different emission wavelengths with multiple replicates and estimating limits of quantitation. Based on the results above, we recommend that Z-DEVD-AMC-based caspase-3 assays be run on the SPECTRAmax GEMINI with excitation/emission wavelengths set to 368/467 nm with a 435 nm or a 420 nm emission cutoff filter.


CHOICE OF MICROPLATES
White plates give a higher fluorescent signal (and background) than do black plates because the light is reflected, rather than absorbed. (The
'"/>

Source:


Page: All 1 2 3 4 5 6

Related biology technology :

1. Fluorescence-Based Single-Tube Assays to Rapidly Detect Human Gene Mutations
2. Matched siRNAs and Assays: Ambion + Applied Biosystems = RNAi Success
3. Using Validated siRNAs in Functional Genomic Assays
4. Nuclease Protection Assays: The Basics
5. Selection of Standards for Bio-Plex Cytokine Assays, Rev A
6. Feasibility of Multiplexing Bio-Plex Total Target and Phosphoprotein Assays
7. The Gradient Feature: Use in Optimization of Allelic Discrimination Assays, Rev B
8. The iCycler iQ Detection System for TaqMan Assays, Rev B
9. Multiplexed Quantitative Peptide Assays for Protein Biomarkers of Cardiovascular Disease in Human Plasma
10. Caspase-3 Activation - An Indicator of Apoptosis in Image-Based Assays
11. Amplification Efficiency of TaqMan Gene Expression Assays
Post Your Comments:
(Date:9/17/2014)... , Sept. 17, 2014 BioClinica®, Inc., ... eClinical Solutions, today announced three of its experts are ... for Clinical Data Management to be held September 28 ... . The Company also announced the innovative ... overall quality in clinical trials acquired earlier this year, ...
(Date:9/17/2014)... MA (PRWEB) September 17, 2014 Terracon ... version of its TerraPure tank . This addition ... is molded from FDA-compliant linear polyethylene and available in ... pre-installed fittings and ships in two weeks or less. ... features and benefits found in Terracon’s existing TerraPure tank ...
(Date:9/17/2014)... , Sept. 17, 2014 Hussey Copper ... MD-Cu 29 antimicrobial copper, announces Gilmour Academy Ice ... the first EPA-registered antimicrobial solid touch surface to help ... a reasonably priced surface that kills greater than 99.9% ... of exposure. 1 , ...
(Date:9/17/2014)... SQI Diagnostics Inc. (TSX-V: SQD) (OTCQX: SQIDF), a life ... products for advanced microarray diagnostics, today announced that its ... the United States on the OTCQX ... 17, 2014 on the OTCQX under the symbol SQIDF.  ... Venture Exchange under its existing symbol SQD. ...
Breaking Biology Technology:BioClinica to Present at 20th Annual SCDM Conference 2BioClinica to Present at 20th Annual SCDM Conference 3Terracon Introduces Pre-Engineered TerraPure® Tank for Ultrapure Water Applications 2Innovative Gilmour Academy Ice Arena changes more than 200 fixtures to Hussey Copper's MD-Cu29 antimicrobial copper to help protect against bacteria 2Innovative Gilmour Academy Ice Arena changes more than 200 fixtures to Hussey Copper's MD-Cu29 antimicrobial copper to help protect against bacteria 3SQI Diagnostics to begin Trading on the OTCQX Marketplace in the United States 2SQI Diagnostics to begin Trading on the OTCQX Marketplace in the United States 3
... Stock Exchange Symbol: MSEDMONTON, March 2 /PRNewswire-FirstCall/ - BioMS ... treatment of multiple sclerosis (MS), announced that effective on ... purchase warrants, each of which entitles the holder to ... of $5.00 per share, will be extended from March ...
... B.C. Technology Promises to Revolutionize Food Processing, ... Corporation (TSX-V: ENW) ("EnWave" or "the Company") today ... of its continuous nutraREV(TM) food dehydration equipment to ... of B.C.,s largest blueberry producers. With this sale, ...
... of "In the Zone for UC" Campaign, Centocor Ortho ... Colitis Foundation of America for Each Point Pisani Scores ... Ortho Biotech Inc. announced today a partnership with professional ... colitis (UC) and funding to support continued education and ...
Cached Biology Technology:BioMS Medical warrant extension 2EnWave Successfully Commercializes nutraREV(TM) Food Dehydration Technology 2EnWave Successfully Commercializes nutraREV(TM) Food Dehydration Technology 3EnWave Successfully Commercializes nutraREV(TM) Food Dehydration Technology 4Pro Hockey Player Fernando Pisani Looks to Shoot and Score Against Ulcerative Colitis 2Pro Hockey Player Fernando Pisani Looks to Shoot and Score Against Ulcerative Colitis 3Pro Hockey Player Fernando Pisani Looks to Shoot and Score Against Ulcerative Colitis 4
(Date:9/16/2014)... us that we inherit certain traits from our parents ... how these genes play out by taking certain drugs ... exactly what a team of researchers at the Boston ... research of epigenetics research. , Epigenetics regulates gene ... and histone proteins, which prevent permanent mutations or alterations ...
(Date:9/16/2014)... spelled doom for the dinosaurs 66 million years ago ... much greater extent than their deciduous peers, according to ... published in the journal PLOS Biology . ... thousands of fossilized leaves of angiosperms flowering ... reconstruct the ecology of a diverse plant community thriving ...
(Date:9/16/2014)... interaction between viruses, bacteria, and their environment is ... collaboration between Matthew Sullivan, associate professor in the ... Biology and Steven Hallam from the University of ... are drivers of nutrient and energy cycles that ... is changing, so are the environments these bacteria ...
Breaking Biology News(10 mins):Epigenetic drugs: A hope to treat cancer resistance and reduce cancer relapse? 2Epigenetic drugs: A hope to treat cancer resistance and reduce cancer relapse? 3Meteorite that doomed dinosaurs remade forests 2Meteorite that doomed dinosaurs remade forests 3Meteorite that doomed dinosaurs remade forests 4New research decodes virus-host interactions in ocean dead zones 2New research decodes virus-host interactions in ocean dead zones 3
... has found all forms of tobacco exposure, whether that be ... of heart attack. , The study by professors Salim Yusuf ... Medicine at McMaster University and Hamilton Health Sciences in Hamilton, ... In collaboration with colleagues from 52 countries, they calculated the ...
... Researchers have found that abnormal stimulation of a ... cells in adults causes invasive tumor-like growths in ... to regress--a finding the researchers said suggests a ... called malignant gliomas. , Arturo Alvarez-Buylla and Erica ...
... savannas contain 70% of the world's plants and are critical ... project involving researchers from the Leeds Earth and Biosphere Institute ... sensitive areas. , The researchers think we may be at ... the rainforests to shrink, so increasing the amount of carbon ...
Cached Biology News:Global study shows all tobacco bad for the heart 2Global study shows all tobacco bad for the heart 3Possible birthplace of malignant brain tumors identified 2The vicious cycle of rainforest destruction 2
100 adhesive strips....
Request Info...
Molecular Biological Resources has extensive abilities cloning and expression of prokaryotic and eukaryotic proteins. Please contact lloconne@molbiores.com to discuss your project. Contact lloconne@...
Lentigen's manufacturing platform is sufficiently versatile for use in the manufacture of any HIV-based Lentiviral vector. Lentigen will manufacture your LV of choice to the highest possible titer a...
Biology Products: