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Glutathione S-Transferase Fluorescent Activity Kit (Invitrogen™)

The Glutathione S-Transferase Activity research-use-only kit is a fluorescent activity assay designed for the quantification and detection of glutathione S-transferase activity in serum, plasma, urine and cell lysates.

This complete, ready-to-use kit includes black 96-well plate(s), glutathione S-transferase standard (10 U/mL), glutathione S-transferase substrate, and other components to perform the assay. A 96-well microplate reader capable of reading the fluorescent emission at 460 nm, with excitation at 370-410 nm, is required for use of this kit.

Performance characteristics
• Assay type: fluorescent activity kit
• Sample types: serum, plasma, urine, and cell lysates
• Sensitivity: 2.70 mU/mL
• Standard curve range: 7.8 mU/mL–500 mU/mL
• Reactivity: species independent

Background
The Glutathione S-Transferase (GST) family of isozymes function to detoxify and neutralize a wide variety of electrophilic molecules by mediating their conjugation with reduced glutathione1. Human GSTs are encoded by 5 gene families, expressed in almost all tissues as four cytosolic and one microsomal forms. Dividing the family by isoelectric points, the basic alpha (pI 8–11), the neutral mu (pI 5–7), and acidic pi (pH<5) classes are populated by additional subclasses, each isozyme displaying differential specificity for given electrophilic molecules. This assay has been validated for human urine, serum, EDTA, heparin plasma, toadfish liver (Opsanus tau), and oyster hemolymph samples. Most cell lysates should also be compatible. GST activity varies across tissues and species, so this kit should measure GST activity from sources other than human.

Assay principle
The Glutathione S-Transferase Fluorescent Activity kit is designed to quantitatively measure the activity of GST present in a variety of samples. A GST standard is provided to generate a standard curve for the assay and all samples should be read off the standard curve. The kit utilizes a non-fluorescent molecule that is a substrate for the GST enzyme that covalently attaches to glutathione (GSH) to yield a highly fluorescent product. Mixing the sample or standard with the supplied detection reagent and GSH and incubating at room temperature for 30 minutes yields a fluorescent product which is read at 460 nm in a fluorescent plate reader with excitation at 390 nm.

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Active Rap1 Pull-Down and Detection Kit (Thermo Scientific™)

The Thermo Scientific Pierce Active Rap1 Pull-Down and Detection Kit is a complete kit for selective enrichment and detection of GTP-bound Rap1 GTPase through specific protein interaction with the RalGDS protein-binding domain.

The Active Rap1 Pull-Down and Detection Kit includes purified GST-RalGDS Rap-binding domain (RBD), glutathione agarose resin, positive and negative controls (GTPγS and GDP, respectively), lysis/binding/wash buffer, anti-Rap1 antibody, sample buffer, spin columns and collection tubes. The kit was validated using lysates from NIH 3T3 cells, a cell line that is known to have robust Rap1 activity.

Features of the Active Rap1 Pull-Down and Detection Kit:

Highly sensitive and accurate—optimized reagents, specific anti-Rap1 antibody and Western blot procedure ensure accurate controls and semi-quantitative results
Validated—functionally tested for Rap1 detection to ensure quality and performance
Compatible—effective with a variety of cell types from mouse, rat and human sources

Applications:
• Follow activation of Rap1 GTPase during cell differentiation, migration, division and cytoskeletal rearrangement
• Study active Rap1 signaling in cell junctions and adhesions
• Monitor Rap1 activity after stimulation with growth factors
• Screen small molecule inhibitors for their effects on Rap1 activity

The Active Rap1 Pull-Down and Detection Kit was validated for function and specificity of the active Rap1 enrichment method using cell lysates treated with GTPγS to activate endogenous Rap1 and compared to lysates treated with GDP to inactivate the small GTPase. GTPγS treatment traps Rap1 in the GTP-bound form (active), resulting in a strong signal when endogenous Rap1 is present. GDP treatment pushes Rap1 into the GDP-bound state (inactive), resulting in minimal or no signal, regardless of Rap1 protein levels. This kit was optimized for Western blot detection using an HRP-conjugated secondary antibody (Goat Anti-rabbit IgG, Part No. 31460) and Thermo Scientific SuperSignal West Pico Chemiluminescent Substrate (Part No. 34080). The kit contains sufficient components for 30 pull-down assays.

Rap Background:
The Rap GTPases are part of the Ras family of GTPases and are encoded by Rap1a, Rap1b and Rap2. Rap GTPases are structurally similar to Ras GTPases and have similar effector and activator proteins, although Rap GTPases have different functional activities than Ras. While Ras is involved in cell proliferation and survival, Rap1 regulates cytoskeletal rearrangements, cell adhesion and cell junction formation.

The specificity of Rap1 and Ras is mediated by their respective upstream regulators and downstream effectors. The GEFs for Rap contain a CDC25 homology domain that mediates the GDP/GTP exchange reaction and a REM domain (Ras exchange motif). Some Rap GEFs include C3G, Epac1 and 2, RasGRP2, PDZ-GEF1 and 2 and PLCε. The binding domain used in this kit is RalGDS, a GEF that contains an RA domain to which Rap1 has a higher binding affinity than Ras. The RapGAPs, including Rap1GAP and the Spa-1 family, insert an asparagine side chain into the nucleotide-binding pocket to catalyze the GTP hydrolysis reaction. Rap effector proteins, including RAPL, Riam, AF-6, Krit1, RacGEFs, Tiam1, Vav2, Rho GAPs and ARAP3, are involved in cell-cell junctions and adhesion and are often localized to the membrane or at cell-cell junctions. Besides Ras, there is also cross-talk between Rap and Rho GTPases.

More Product Data
Measure activation of small GTPases via their specific downstream effectors

NA-Star™ Influenza Neuraminidase Inhibitor Resistance Detection Reagent Set (Invitrogen™)

The NA-Star® Influenza Neuraminidase Inhibitor Resistance Detection Reagent Set is designed for the rapid and sensitive quantitation of influenza neuraminidase inhibitor resistance in dilutions of virus culture medium in a 96-well microplate format. The NA-Star® Influenza Neuraminidase Inhibitor Reagent kit provides all necessary assay reagents enabling improved global assay standardization and more accurate comparison of results lab-to-lab.

The NA-Star® Influenza Neuraminidase Inhibitor Resistance Detection Kit includes everything you need to quantitate neuraminidase activity and neuraminidase inhibitor resistance in avian, equine, human (types A and B), and porcine influenza viruses. The kit's fast and easy protocol and convenient 96-well plate format make it ideal for monitoring influenza virus neuraminidase inhibitor resistance, as well as high-throughput inhibitor compound screening.

• Includes pre-made solutions for all required reagents, enabling you to standardize assays and compare results between labs easily.
• Provides up to 50-fold higher sensitivity than assays using fluorescent methylumbelliferone N-acetylneuraminic acid (MUNANA) substrate.
• Dynamic range of four orders of magnitude enables you to quantitate virus isolates over a broad range of virus concentration and with varying levels of neuraminidase activity without performing numerous sample dilutions.
• Fast and easy protocol enables you to complete assays in less than 1.5 hours, providing rapid assay throughput.

Get Results in Less than 1.5 Hours
The fast and easy protocol enables you to perform assays in less than 1.5 hours. Simply incubate your virus samples with dilutions of neuraminidase inhibitor, add NA-Star chemiluminescent substrate, incubate again, and then add the accelerator solution, which triggers light emission from the reaction product. Light signal is measured with a luminometer, including multi-mode instruments that include a luminometer mode. For best results, use a luminometer with an automatic injector to add the accelerator solution. You may use a luminometer without an injector, provided you add reagents with a multichannel pipettor and read the plate immediately.

Up to 50-Fold Higher Sensitivity than MUNANA-Based Assays
The chemiluminescent-based detection technology provides a wide dynamic range -- greater than four orders of magnitude of neuraminidase concentration (two orders of magnitude greater than fluorescent MUNANA-based assays) -- enabling you to quantitate neuraminidase inhibitor resistance levels over a broad range of virus concentration and neuraminidase activity without having to test multiple virus dilutions.

Pre-Made Reagents Make it Easy to Standardize Assays
Quality-tested protocols, and pre-made reagents eliminate variability, enabling you to easily standardize assay performance and results across experiments and different laboratories.

Monitor Drug Resistance of Human, Avian, and Livestock Influenza Viruses
The kit's ease of use, standardization, and broad range of compatible species (avian, equine, human types A and B, and porcine influenza viruses) make it an important new tool for assessing and researching drug resistance of influenza in humans, birds, and livestock.

Screen for New Inhibitors and Develop New Vaccines
The kit has many other applications, including screening for new neuraminidase inhibitors and quantitating viral neuraminidase activity for vaccine development. It is also ideal for quantitating neuraminidases from bacteria, including S. typhimurium, C. perfringens, V. cholera, and likely others.

For Research Use Only. Not for use in diagnostic procedures.

Amplex™ Acetylcholine/Acetlycholinesterase Assay Kit (Invitrogen™)

The Amplex® Red Acetylcholine/Acetylcholinesterase Assay Kit provides an ultrasensitive method for detecting acetylcholinesterase (AChE) activity in a fluorescence microplate reader or fluorometer.

See our complete line of Fluorescence Microplate assays.

• Detect AChE activity levels as low as 0.002 U/mL in one hour
• Detect acetylcholine levels as low as 0.3 µM using excess AChE
• Achieve detection ranges of 0.3 µM to 100 µM acetylcholine
• Format allows for multiple time point measurements
• Designed for minimal autofluorescence interference

AChE activity is monitored indirectly using 10-acetyl-3,7-dihydroxyphenoxazine (Amplex® Red reagent), a sensitive fluorogenic probe for hydrogen peroxide. AChE converts the acetylcholine substrate to choline, which is then oxidized by choline oxidase to betaine and hydrogen peroxide. In the presence of horseradish peroxidase, hydrogen peroxide reacts with the Amplex® Red reagent in a 1:1 stoichiometric ratio to generate the highly fluorescent product resorufin.

Because resorufin has absorption and fluorescence emission maxima of approximately 571 nm and 585 nm, respectively, there is little interference from autofluorescence in most biological samples. Experiments with purified AChE from electric eel indicate that the Amplex® Red Acetylcholine/Acetylcholinesterase Assay Kit can detect AChE levels as low as 0.002 U/mL using a reaction time of one hour. By providing an excess of AChE in the assay, the kit can also be used to detect acetylcholine levels as low as 0.3 µM, with a range of detection from 0.3 µM to 100 µM acetylcholine.

Use Amplex® Red Assays for a Broad Range of Investigations
A wide variety of validated Amplex® Red assays are available for studying cell signaling and lipids, neurobiology, inflammation and immune function, and metabolism. We also offer Amplex® UltraRed Reagent (Cat. No. A36006), a second-generation reagent providing greater sensitivity and brighter fluorescence, and the Amplex® Red/UltraRed Stop Reagent (Cat. No. A33855). The Amplex® Red/UltraRed Stop Reagent provides convenience and control by allowing the fluorescence signal-generating reaction to be terminated at a user-determined time point. After addition of the stop reagent, the fluorescence signal remains stable for at least three hours. Custom assay design and packaging are also available.

EnzChek™ Phospholipase A1 Assay Kit (Invitrogen™)

This kits takes our stand-alone assay for phospholipase A1, PLA1 (A10070) and combines the necessary reagents to run 2 to 10 complete 96 well microplate assays that monitor activity in purified enzyme preparations and cell lysates. The importance of phospholipases in cellular signaling, lipid metabolism, inflammatory responses and pathological disorders related to these processes has stimulated demand for fluorescence-based activity monitoring methods. In particular the phospholipases resident in plasma and endothelium can perturb circulating LDL and HDL particles, creating pro-artherogenic forms. Recent evidence is drawing a link between these lipases and the progression of several severe neurodegenerative diseases, including Alzheimer's. Molecular Probes’ fluorogenic phospholipase A1 substrates is designed to provide continuous monitoring of phospholipase A1 (PLA1) in purified enzyme preparations, cell lysates and living cells. PLA-1 improves upon two existing reagents in two ways: first it is now PLA-1 specific and secondly it has improved the assay quality by decreasing initial background noise.

BACE1 (β-Secretase) FRET Assay Kit, red

BACE1 (β-secretase) is a key enzyme involved in the production of Amyloid beta-peptides (Abeta) found in extracellular amyloid plaques of Alzheimers disease (AD). In some cases, early onset familial AD can be attributed to a "Swedish" mutation in the amyloid precursor protein (APP), dramatically enhancing the cleavage of this protein by BACE1. This and other genetic and pathological evidence has led to therapeutic approaches focusing on the inhibition of BACE1 and other APP-cleaving enzymes, such as gamma-secretase. The BACE1 fluorescence resonance energy transfer (FRET) Assay Kit provides a sensitive and efficient method for screening potential BACE1 inhibitors. This kit uses purified baculovirus-expressed BACE1 and a new red FRET peptide substrate based on the "Swedish" mutant.

The BACE1 FRET Assay Kit offers:

• Red emission to reduce compound interference
• Excitation at 545 nm and read emission at 585 nm
• An easy-to-use, rapid, homogeneous assay that is performed in solution
• Versatile formatting for high-throughput screening
• Stable results—measurements can be made for up to 24 hours

The principle of the BACE1 FRET assay is as follows: The peptide substrate is synthesized with two fluorophores, a fluorescent donor (a rhodamine (Rh) derivative), and a proprietary quenching acceptor. The distance between these two groups has been selected so that upon light excitation, the donor (D) fluorescence energy is significantly quenched by the acceptor (A) through a quantum mechanical phenomenon known as resonance energy transfer (Figure 1). Upon cleavage by the protease, the fluorophore is separated from the quenching group, restoring the full fluorescence yield of the donor. Thus, a weakly fluorescent peptide substrate becomes highly fluorescent upon enzymatic cleavage; the increase in fluorescence is linearly related to the rate of proteolysis.

Source of BACE1 Enzyme:
A cDNA sequence encoding amino acids 1-460 of human BACE1, corresponding to the ectodomain, was expressed in recombinant, baculovirus-infected insect cells. Purified BACE1 exists as the proBACE1 form having an apparent molecular weight of 55 kDa and an N-terminal sequence of TQHGIRLPLR.

EnzChek™ Ultra Xylanase Assay Kit (Invitrogen™)

The EnzChek® Ultra Xylanase Assay Kit features a quick and convenient mix-and-read format for the detection and monitoring of xylanase activity. The increase in fluorescence resulting from xylanase activity is measured using a fluorometer or fluorescence microplate reader.

See our complete line of Fluorescence Microplate assays.

• Detect concentrations of xylanase activity as low as 1.5 mU/mL
• Suitable for a broad pH range (pH 4 to 10)
• Format allows for continuous detection of xylanase activity
• Excitation/emission maxima of ~358/455 nm, well suited for DAPI filter settings

The EnzChek® Ultra Xylanase Assay Kit provides the speed, sensitivity, and convenience required for measuring xylanase activity or for screening xylanase inhibitors in a high-throughput format. The hydrolysis of xylosidic linkages within the included Xylanase Substrate (hemicellulose polysaccharides) results in the unquenching of the attached fluorescent dyes. This kit can be used for continuous detection of xylanase activity, and offers broad dynamic and pH ranges. Each kit contains sufficient substrate for ~500 assays in a 96-well microplate format. Additionally, the kit contains a fluorescent reference standard that can be used to quantify the xylanase activity.

Histone Demethylase Fluorescent Activity Kit (Invitrogen™)

The Histone Demethylase Activity research-use-only kit is a fluorescent activity assay designed for the quantification and detection of histone demethylase activity of lysine-specific histone demethylase 1 (LSD1)-and Jumonji-type demethylases.

This kit includes black 96-well plate(s), LSD1/JMJD2A assay buffers, formaldehyde standard, and other components to perform the assay. This kit does not contain demethylase enzyme samples. A source of LSD1-type or Jumonji-type demethylase, along with any cofactors, enzyme substrates, inhibitors, and/or activators need to be supplied by the user. A fluorescence 96-well microplate reader capable of reading fluorescent emission at 510 nm, with excitation at 450 nm, is required for use of this kit.

Performance characteristics
• Assay type: fluorescent activity kit
• Sample types: lysine-specific histone demethylase 1 (LSD1) and Jumonji-type demethylases
• Standard curve range: 0.128 uM–0.64 uM for LSD1, 2.5 uM-10uM for JMJD2A
• Reactivity: human

Background
Histone Demethylase (HDM) catalyzes the site-specific demethylation of methyl-lysine residues in histones to dynamically regulate chromatin structure, gene expression, and potentially other genomic functions. Lysine-specific HDMs were first discovered in 2004 and are currently among the most actively studied formaldehyde-producing enzymes. At present, there are two known classes of HDMs: the flavin adenine dinucleotide (FAD)-dependent Lysine Specific Demethylase 1 (LSD1) family and the Fe(II)-dependent Jumonji C (JmjC) family. Although the LSD1 and JmjC HDMs employ different cofactors and catalytic mechanisms, both produce formaldehyde as a byproduct of the demethylation reaction.

Despite their biological importance, HDMs have proven difficult to quantitatively assay owing to their relatively low turnover numbers, hindering understanding of their kinetic properties, substrate specificities, and reaction mechanisms. This assay has been validated for lysine-specific histone demethylase 1 (LSD1)-and Jumonji-type demethylases. For HDM samples in cell lysates, we include a specially formulated Cell Lysis Buffer, that has been shown not to interfere with formaldehyde detection. Cell lysis buffers containing SDS and Triton X-100 inhibit the formaldehyde signal reaction and should not be used.

Assay principle
The Histone Demethylase Activity kit is to quantitatively measure the enzymatic activity of formaldehyde-producing enzymes such as histone demethylases. The kit is unique in that the product of these enzymatic demethylation reactions, formaldehyde, is quantitated directly by a fluorescent product. No separation or washing is required. The kit has been validated for both LSD1 and JMJD2A histone demethylases (HDMs).

The kit provides optimized buffers for the HDMs LSD1 and JMJD2A, a stable formaldehyde standard, the Formaldehyde Detection Reagent (FDR), and two 96-well plates for detecting the generated fluorescent signal. The kit allows any enzymatic reaction generating formaldehyde to be measured. The end user will have to provide the demethylase system and any cofactors necessary for activity, along with any test inhibitors or activators. The kit allows end users to produce HDM activity in many in vivo and in vitro systems and then determine the activity by measuring formaldehyde generation. For in vitro studies, the HDM reaction should be carried out in our supplied buffers using optimized reaction conditions for the demethylation.

Following the formaldehyde generating reaction, the reaction can be stopped by addition of a suitable inhibitor. The FDR is then added to all the wells. If calibration to formaldehyde is needed (for cross lab comparisons) then a formaldehyde standard curve generated from the supplied standard should be run. After a short incubation at 37°C for 30 minutes, the fluorescent product is read at 510 nm in a fluorescent plate reader with excitation at 450 nm.

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Superoxide Dismutase (SOD) Colorimetric Activity Kit (Invitrogen™)

The Superoxide Dismutase Activity research-use-only kit is a colorimetric activity assay designed for the quantification and detection of superoxide dismutase activity in serum, plasma, cells, tissues and erythrocyte lysates.

This complete, ready-to-use kit includes clear 96-well plate(s), superoxide dismutase standard (1 Unit/vial), superoxide dismutase substrate, and other components to perform the assay. A 96-well microplate reader capable of reading optical density at 450 nm is required for use of this kit.

Performance characteristics
• Assay type: colorimetric activity kit
• Sample types: serum, plasma, cells, tissues, and erythrocyte lysates
• Sensitivity: 0.044 U/mL
• Standard curve range: 0.06 U/mL–4 U/mL
• Reactivity: species independent

Background
Short-lived and highly reactive oxygen species (ROS) such as superoxide, hydroxyl radical, and hydrogen peroxide are continuously generated in vivo. In the resting state, the balance between antioxidants and oxidants is sufficient to prevent the disruption of normal physiologic functions; however, either increases in oxidants or decreases in antioxidants can disrupt this balance giving rise to elevated levels of reactive oxygen species (ROS). The cellular levels of ROS are controlled by antioxidant enzymes and small molecule antioxidants. The major antioxidant enzymes, superoxide dismutases (SODs), including copper-zinc superoxide dismutase (Cu/ZnSOD, SOD1), manganese superoxide dismutase (MnSOD, SOD2) and extracellular superoxide dismutase (EC-SOD, SOD3), all play critical roles in scavenging superoxide .

Decreased SOD activity results in elevated level of superoxide which in turn leads to decreased NO but increased peroxynitrite concentrations. The major intracellular SOD is a 32-kD copper and zinc containing homodimer (Cu/Zn SOD). The mitochondrial SOD (MnSOD) is a manganese-containing 93-kD homotetramer that is synthesized in the cytoplasm and translocated to the inner matrix of mitochondria. This assay has been validated for serum and urine samples. Superoxide dismutases are ancient enzymes that should behave in a similar manner to the colorimetric substrate. This assay should to measure SOD activity from a wide range of sources.

Assay principle
The Superoxide Dismutase Activity kit is designed to quantitatively measure SOD activity in a variety of samples. The assay measures all types of SOD activity, including Cu/Zn, Mn, and FeSOD types. A bovine erythrocyte SOD standard is provided to generate a standard curve for the assay and all samples should be read off of the standard curve. Samples are diluted in our specially colored sample diluent and added to the wells. The substrate is added followed by Xanthine Oxidase Reagent and incubated at room temperature for 20 minutes. The xanthine oxidase generates superoxide in the presence of oxygen, which converts a colorless substrate in the detection reagent into a yellow colored product. The colored product is read at 450 nm. Increasing levels of SOD in the samples causes a decrease in superoxide concentration and a reduction in yellow product.

SOD unit definition
One unit of SOD is defined as the amount of enzyme causing half the maximum inhibition of the reduction of 1.5 mM Nitro blue tetrazolium in the presence of riboflavin at 25°C and pH 7.8.

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Glutathione Reductase Fluorescent Activity Kit (Invitrogen™)

The Glutathione Reductase Activity research-use-only kit is a fluorescent activity assay designed for the quantification and detection of glutathione reductase activity in serum, plasma, RBCs and cell lysates.

This complete, ready-to-use kit includes black 96-well plate(s), glutathione reductase standard (200 mU/mL), glutathione reductase substrate, and other components to perform the assay. A 96-well microplate reader capable of reading the fluorescent emission at 510 nm, with excitation at 390 nm, is required for use of this kit.

Performance characteristics
• Assay type: fluorescent activity kit
• Sample types: serum, plasma, RBCs, and cell lysates
• Sensitivity: 0.009 mU/mL
• Standard curve range: 0.15 mU/mL–5 mU/mL
• Reactivity: human

Background
Glutathione Reductase (GR) plays an indirect but essential role in the prevention of oxidative damage within the cell by helping to maintain appropriate levels of intracellular glutathione (GSH). GSH, in conjuction with the enzyme glutathione peroxidase (GP), is the acting reductant responsible for minimizing harmful hydrogen peroxide cellular levels. The regeneration of GSH is catalyzed by GR. GR is a ubiquitous 100-120 kDa dimeric flavoprotein that catalyzes the reduction of oxidized glutathione (GSSG) to reduced glutathione, using beta-nicotinamide dinucleotide phosphate (NADPH) as the hydrogen donor.

Molecules such as NADPH act as hydride donors in a variety of enzymatic processes. NADPH has been suggested to also act as an indirectly operating antioxidant, given its role in the re-reduction of GSSG to GSH and thus maintaining the anti-oxidative power of glutathione. This assay has been validated for human serum, EDTA and heparin plasma, and isolated erythrocytes. Most cell lysates should also be compatible. GR activity varies across tissues and species, however this kit may measure GR activity from sources other than human.

Assay principle
The Glutathione Reductase Activity kit is designed to quantitatively measure Glutathione Reductase (GR) activity in a variety of samples. A GR standard is provided to generate a standard curve for the assay and all samples should be read off the standard curve. The kit utilizes a proprietary non-fluorescent detection reagent that will covalently bind to the free thiol group on GSH generated in the reduction of oxidized glutathione (GSSG) to yield a highly fluorescent product. After mixing the sample or standard with detection reagent and incubating at room temperature, the fluorescent product is read at 510 nm in a fluorescent plate reader with excitation at 390 nm

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Learn more about other immunoassays

EnzChek™ Direct Phospholipase C Assay Kit (Invitrogen™)

This kit provides continuous monitoring of phosphotidyl choline specific phospholipase C (PLC) activity in microplate based biochemical assays. The assay uses a PCL selective fluorogenic substrate that emits a bright green emission upon cleavage.

The EnzChek® Direct Phospholipase C Assay Kit provides a simple and robust method for monitoring PC-PLC activity. Each kit provides enough reagents for 2 microplates, using 200 µl volumes in 96 well format. PC-PLC plays a crucial role in many cell signaling pathways involved in apoptosis and cell survival, as well as diseases ranging from cancer to HIV1-7. The assay uses a glycerophospho-ethanolamine with a dye-labeled sn-2 acyl chain as a substrate for PC-PLC. Substrate cleavage by PC-PLC before the phosphate releases the dye-labeled diacylglycerol, which produces a positive fluorescence signal that may be measured continuously. The reaction product has absorption and fluorescence emission maxima of 509 nm and 516 nm, respectively. Using purified enzyme from Bacillus cereus, the assay can detect as little as 10 mU⁄mL PC-PLC after one hour at room temperature. The kit has been proven useful for characterizing PC-PLC inhibition, and since it offers a direct measurement, the potential for false positives in a compound screen is eliminated.

NA-Star™ Influenza Neuraminidase Inhibitor Resistance Detection Kit (Invitrogen™)

The NA-Star® Influenza Neuraminidase Inhibitor Resistance Detection Kit is designed for the rapid and sensitive quantitation of influenza neuraminidase inhibitor resistance in dilutions of virus culture medium in a 96-well microplate format. The NA-Star® Influenza Neuraminidase Inhibitor Reagent kit provides all necessary assay reagents enabling improved global assay standardization and more accurate comparison of results lab-to-lab.

The NA-Star® Influenza Neuraminidase Inhibitor Resistance Detection Kit includes everything you need to quantitate neuraminidase activity and neuraminidase inhibitor resistance in avian, equine, human (types A and B), and porcine influenza viruses. The kit's fast and easy protocol and convenient 96-well plate format make it ideal for monitoring influenza virus neuraminidase inhibitor resistance, as well as high-throughput inhibitor compound screening.

• Includes pre-made solutions for all required reagents, enabling you to standardize assays and compare results between labs easily.
• Provides up to 50-fold higher sensitivity than assays using fluorescent methylumbelliferone N-acetylneuraminic acid (MUNANA) substrate.
• Dynamic range of four orders of magnitude enables you to quantitate virus isolates over a broad range of virus concentration and with varying levels of neuraminidase activity without performing numerous sample dilutions.
• Fast and easy protocol enables you to complete assays in less than 1.5 hours, providing rapid assay throughput.

Complete Kit for Measuring the Neuraminidase Inhibitor Resistance of Influenza Viruses
The NA-Star® Influenza Neuraminidase Inhibitor Resistance Detection Kit includes NA-Star chemiluminescent substrate for neuraminidase, all necessary assay reagents, and microplates -- everything you need for the fast, accurate quantitation of neuraminidase inhibitor resistance in influenza virus isolates.

Get Results in Less than 1.5 Hours
The kit's fast and easy protocol enables you to perform assays in less than 1.5 hours. Simply incubate your virus samples with dilutions of neuraminidase inhibitor, add NA-Star chemiluminescent substrate, incubate again, and then add the accelerator solution, which triggers light emission from the reaction product. Light signal is measured with a luminometer, including multi-mode instruments that include a luminometer mode. For best results, use a luminometer with an automatic injector to add the accelerator solution. You may use a luminometer without an injector, provided you add reagents with a multichannel pipettor and read the plate immediately.

Up to 50-Fold Higher Sensitivity than MUNANA-Based Assays
The kit's chemiluminescent-based detection technology provides a wide dynamic range -- greater than four orders of magnitude of neuraminidase concentration (two orders of magnitude greater than fluorescent MUNANA-based assays) -- enabling you to quantitate neuraminidase inhibitor resistance levels over a broad range of virus concentration and neuraminidase activity without having to test multiple virus dilutions.

Pre-Made Reagents Make it Easy to Standardize Assays
Quality-tested protocols, microplates, and pre-made reagents eliminate variability, enabling you to easily standardize assay performance and results across experiments and different laboratories.

Monitor Drug Resistance of Human, Avian, and Livestock Influenza Viruses
The kit's ease of use, standardization, and broad range of compatible species (avian, equine, human types A and B, and porcine influenza viruses) make it an important new tool for assessing and researching drug resistance of influenza in humans, birds, and livestock.

Screen for New Inhibitors and Develop New Vaccines
The kit has many other applications, including screening for new neuraminidase inhibitors and quantitating viral neuraminidase activity for vaccine development. It is also ideal for quantitating neuraminidases from bacteria, including S. typhimurium, C. perfringens, V. cholera, and likely others.

For Research Use Only. Not for use in diagnostic procedures.

Amplex™ Red Phospholipase D Assay Kit (Invitrogen™)

The Amplex® Red Phospholipase D Assay Kit provides a sensitive and simplemethod to detect Phospholipase D (PLD) activity using a fluorescence microplate reader or fluorometer.

See our complete line of Fluorescence Microplate assays.

• Detect phospholipase D activity levels as low as 10 U/mL
• Format allows for multiple time point measurements
• Designed for minimal autofluorescence interference

PLD activity is monitored indirectly using 10-acetyl-3,7-dihydroxyphenoxazine (Amplex® Red reagent), a sensitive fluorogenic probe for hydrogen peroxide. PLD converts the phosphatidylcholine (lecithin) to choline, which is then oxidized by choline oxidase to betaine and hydrogen peroxide. In the presence of horseradish peroxidase, huydrogen peroxide reacts with the Amplex® Red reagent in a 1:1 stoichiometric ratio to generate the highly fluorescent product resorufin.

Because resorufin has absorption and fluorescence emission maxima of approximately 571 nm and 585 nm, respectively, there is little interference from autofluorescence in most biological samples.

Use Amplex® Red Assays for a Broad Range of Investigations
A wide variety of validated Amplex® Red assays are available for studying cell signaling and lipids, neurobiology, inflammation and immune function, and metabolism. We also offer Amplex® UltraRed Reagent (Cat. No. A36006), a second-generation reagent providing greater sensitivity and brighter fluorescence, and the Amplex® Red/UltraRed Stop Reagent (Cat. No. A33855). The Amplex® Red/UltraRed Stop Reagent provides convenience and control by allowing the fluorescence signal-generating reaction to be terminated at a user-determined time point. After addition of the stop reagent, the fluorescence signal remains stable for at least three hours. Custom assay design and packaging are also available.

EnzChek™ Ultra Amylase Assay Kit (Invitrogen™)

The EnzChek® Ultra Amylase Assay Kit provides a solution-based assay featuring the speed, high sensitivity, and convenience required for measuring amylase activity or for screening amylase inhibitors in a high-throughput format. This EnzChek® kit contains a starch derivative—the DQ™ starch substrate—that is labeled with BODIPY® FL dye to such a degree that the fluorescence is quenched. This substrate is efficiently degraded by amylase; digestion relieves the quenching and yields highly fluorescent fragments. The accompanying increase in fluorescence is proportional to amylase activity and can be monitored with a fluorescence microplate reader or fluorometer, using standard fluorescein filters.

EnzChek® Ultra Amylase Assay Kit Specifications:
• Label (Ex/Em): BODIPY® FL conjugate (~502/512 nm)
• Kit contains lyophilized substrate, 10X reaction buffer, substrate solvent, a fluorescent standard, and a detailed protocol
• Sufficient reagents are supplied for 500 assays (using a 100 µL assay volume in a 96-well microplate assay format)


Find Fluorescent Substrates for Other Glycosidases
In addition to the EnzChek® Ultra Amylase Assay Kit, we offer kits and substrates to measure xylanase, lysozyme, β-galactosidase, and more. Review Detecting Glycosidases—Section 10.2 in the Molecular Probes® Handbook for more information on these products.

For Research Use Only. Not for human or animal therapeutic or diagnostic use.

ActivX™ Azido-FP Serine Hydrolase Probe (Thermo Scientific™)

Thermo Scientific Serine Hydrolase Probes are ActivX™ Fluorophosphonate (FP) and other tagged phosphonate probes to purify or assay serine hydrolase active sites using fluorescence, biotin-affinity, or mass spectrometry.

Features of ActivX Azido-FP Serine Hydrolase Probe:

Specific—labels the reactive site of active serine hydrolases
Compatible—tags available for capture, detection and Staudinger conjugation
Flexible—use for in vitro or intracellular enzyme labeling

These ActivX™ FP Probes feature a reactive fluorophosphonate group that specifically and covalently labels the active-site serine of enzymatically active serine hydrolases. These probes are available with a desthiobiotin (biotin analog) tag for selective enrichment, TAMRA fluorophore for detection or a reactive- azido group (Staudinger reagents) that facilitates multiplex labeling when used with phosphine- or alkyne-derivatized tags. These probes can be used to assess activity or screen small molecule inhibitors against enzymes derived from cell lysates, subcellular fractions, tissues and recombinant proteins.

Applications:
• Determine serine hydrolase enzyme activity in cells and lysates
• Mapping the active-site serine of functionally diverse serine hydrolase family members (e.g. proteases, lipases, esterases)
• Screen for small molecule binding affinities and active-site inhibition
• Profile serine hydrolases using fluorescent, Western blot or mass spectrometry workflows

ActivX active-site probes are especially advantageous for determining active enzyme levels compared to other protein expression profiling techniques that only measure abundance. Because many of the proteolytic enzymes in the serine hydrolase family are expressed as inactive proenzymes (zymogens), the ActivX FP probes selective enrichment only those enzymes that are functionally active and biologically relevant at the time of labeling. This feature also makes it possible to perform selective screening of inhibitors or other conditions that alter enzyme activity.

Active serine hydrolases labeled with ActivX FP Probes can be detected and quantified by Western blot, fluorescent gel imaging or mass spectrometry by using a compatible tag. The TAMRA-FP probe can be used to label and detect active serine hydrolases in samples by fluorescent gel imaging, capillary electrophoresis or mass spectrometry. An anti-TAMRA antibody is also available for immuno-enrichment of TAMRA-FP probe-labeled proteins. The Azido-FP probe is used in combination with a phosphine- or alkyne-derivatized tag for either detection or enrichment. Desthiobiotin-FP probes can be used for streptavidin-based enrichment and detection of active serine hydrolase proteins in Western blotting or mass spectrometry.

The serine hydrolase superfamily is one of the largest, most diverse enzyme families in eukaryotic proteomes. Serine hydrolases are generally grouped into two large families: serine proteases (e.g., trypsin, elastase and thrombin) and metabolic serine hydrolases. Metabolic serine hydrolases are divided into multiple enzyme subclasses (e.g., esterases, lipases, amidases and peptidases) based on structure, catalytic mechanism and substrate preference.

Related Products
ActivX™ TAMRA-FP Serine Hydrolase Probe
ActivX™ Desthiobiotin-FP Serine Hydrolase Probe