Amplite® Reagents and Assay Kits

The Amplite^® product series is composed of enzyme-based substrates and assays optimized for measuring horseradish peroxidase (HRP), luciferase, phosphodiesterases, proteases, histone deacetylase (HDAC) and cell signaling molecules such as NAD/NADH, NADP/NADPH, etc. The series includes assay kits spanning essentially all aspects of cellular and physiological activity, and is compatible with other AAT Bioquest products as well as classic materials.

Fluorogenic Substrates

---

AAT Bioquest's substrates are a component of many of our kits, and are also available as standalone items. Spanning a wide spectral range, from the bright Amplite^® Blue to the long-wavelength Amplite^® IR, these substrates fluoresce in the presence of the peroxidase enzyme and H2O2. This series has a high signal-to-noise ratio and is very stable. Each substrate demonstrates useful features; Amplite^® Blue does not photobleach, for example, and Amplite^® IR has both excellent water solubility and is pH-independent from pH 4-10.

Quantitation Kits

---

Our Amplite^® quantitation kits provide a rapid, dependable method of measuring a variety of biomaterials, ranging from alpha-ketoglutarate to zinc ions. Simple and efficient to use, these kits reliably measure the target materials, either in total amounts or by concentration, depending on the kit selected. The kits are designed to be all-inclusive, although some reagents are available separately as well, for more unusual experimental requirements. For example, the Amplite® Total Sulfide Quantification Kit has Sulfide Green™ AM as a central component.







Each of the Amplite^® biomarker quantitation kits employ our proprietary Amplite^® Red substrate, compatible with with common instrument platforms such as microplate or fluorescent microscope. A colorimetric format is also available for the detection of glucose.

Endotoxin Detection Kits

---

Lipopolysaccharide, aka endotoxin, is a potentially-lethal biomarker of bacterial invasion. Its detection and prevention are of outmost importance in the pharmaceutical and biomedical industries. Employing Limulus Amebocyte Lysate (LAL), these endotoxin kits generate signal proportional to the presence of endotoxin in the sample. The kits are available in either colorimetric or fluorimetric format, and employ either Endotoxin Yellow™ or Endotoxin Green™ proprietary substrates, respectively. Both kits detect a wide range of endotoxin concentration, with the fluorimetric kit having approximately twice the sensitivity (detection of concentrations as low as 0.002 EU/ml) as the colorimetric kit.

Assay Kits

---

Enzyme Detection

Enzymes act as a regulatory catalyst for the majority of biochemical reactions that facilitate cell function, including all aspects of cellular metabolism. They do so by lowering the energy barrier necessary for a chemical reaction to occur. Enzymes are present throughout the cell, some sequestered within organelles such as the mitochondria, or free in the cytoplasm. Upon reaction with a suitable substrate, enzymes change shape to form a new complex. This change, when used during experiments, can provide a signal to researchers that a reaction has occurred, as well as potentially improving the specificity of the reaction itself. Amplite^® enzyme-based assays provide a sensitive and simple method for the quantitation of a diverse array of analytes, including dehydrogenases, kinases, and other enzyme groups.

Learn More

Enzyme Activity

Enzyme activity assays are the primary tool for studying enzyme kinetics and inhibition. AAT Bioquest offers a full line of enzymatic activity kits for targets including hydrolytic enzymes, proteases, and other enzyme families.




In addition to the kits in the table above, AAT Bioquest offers a range of kits specifically for investigating the activity of caspases. Click the button below to learn more:

Learn More

Luciferase Reporter Gene

The luciferase reporter gene, originally isolated from the common eastern firefly Photinus pyralis, is the most popular choice for detection assays and other uses due to its low toxicity and ease of use. Our Amplite^® kits employ different variations of this useful gene, isolated from the firefly, sea pansy (Renilla reniformis) and aquatic copepod Gaussia prince for sensitive, efficient detection and steady signal. AAT Bioquest also offers its bioluminescent substrate Luciferin in several forms.

Horseradish Peroxidase (HRP)

The peroxidase enzyme family plays a prominent role in multiple cellular processes. They catalyze hydrogen peroxide (H2O2) oxidation of a variety of organic compounds, many of which are commonly present in living cells. For use in the laboratory, the most commonly used one is isolated from the common horseradish. Horseradish peroxidase (HRP) is a small, robust molecule of only 44kD, and can be easily conjugated to antibodies in a 4:1 ratio with little to no steric hindrance. Since H202 is a common product of many cellular pathways, the ability to quantify it makes HRP incredibly valuable for multiple biochemical applications.

Learn More

Alkaline Phosphatase (ALP)

Alkaline phosphatase (ALP) is an 86 kDa homodimeric protein enzyme present in multiple tissue types within the body, with the highest concentrations in the bone and liver. It is used as a biomarker for several pathologies. ALP is also a common reporter enzyme used in multiple immunoassays and other applications. It is typically conjugated to an antibody, and can be detected in multiple formats. The substrates for the Amplite^® ALP kits range from the popular para-Nitrophenylphosphate (pnPP) for colorimetric detection and 4-Methylumbelliferyl phosphate (MUP) for fluorimetric, to our proprietary substrate for luminometric platforms.

Learn More

Phosphatase Assays and Reaction Control

For researchers requiring control over the phosphatase enzyme reaction, we suggest using the paired Signal Guard™ Phosphatase Reaction Stopping Solution:

• The Signal Guard™ phosphatase reaction stopping solution is a ready to use reagent that provides a convenient tool for terminating fluorescence and colorimetric signal-generating phosphatase reactions at a user-determined time point, and also keep the fluorescence or colorimetric signal stable for up to 18 hours.
• The stopping solution is optimized and compatible with alkaline, acid and protein phosphatase.
• Compatible with nearly all colorimetric and fluorimetric phosphatase substrates, the solution requires only mild pH and only a 2-step protocol. Full details and pdf protocol available here.

NAD(P)(H) Assays

Nicotinamide adenine dinucleotides are prolific cofactors that facilitate the reversible oxidation and reduction steps in all major metabolic pathways. They are present in varying concentrations in phosphorylated (NADP and NADPH) and unphosphorylated forms (NAD+ and NADH+). Since the NAD(P)(H) cofactors link cell metabolism to other systems in the body (including transcriptional control and cell signaling) they are a target for drug discovery and oncology research. By monitoring the changes in intracellular concentration of these biomolecules, researchers gain understanding of cell behavior and pathologies.

Learn More

Cellular Energy and Metabolism

All living cells must be able to fuel themselves with some form of energy. The process of transforming outside sources of energy (e.g. sunlight, food) into adenosine triphosphate (ATP) is the beginning of all life processes. Major metabolic pathways such as glycolysis are heavily regulated and include multiple enzymes and molecular intermediates. Assays to quantify changes in the concentrations of these biomolecules give valuable information about pathological mechanisms as well as normal cell activities.

Antioxidants and ROS

Reactive oxygen species (ROS) are a major component of cell signaling, gene expression, and other activities, with most occurring in the mitochondria. The amount of ROS in the cell is usually at low concentrations, as they are used for multiple cell signaling activities and other purposes. The levels are also kept low by the presence of antioxidants, which are synthesized or imported by the cell. In high concentrations however, these chemically-reactive molecules can be very damaging and even lethal.

Being able to accurately measure ROS levels is a useful way to detect the early stages of multiple pathologies, as well as the activity of metabolites or normal/atypical cellular function.


In addition to the kits in the table above, AAT Bioquest offers a range of kits specifically for investigating the activity of Glutathione (GSH and GSSG). Click the button below to learn more:

Learn More

Amino Acids

As the building blocks of all proteins, amino acids play a role in every aspect of cellular activity. Sensitive, accurate measurements of these small compounds, both essential and non-essential, are relevant in multiple fields of study.

Nerve Signaling

Sphingomyelin and acetylcholine, along with other roles, are important to signal transduction in the central nervous system. Using our proprietary Amplite^® fluorescent substrates, these kits provide optimized protocols to give accurate, sensitive biomolecule quantification.

AAT Bioquest also offers Amplite^® assay kits for the detection of the enzymes associated with these biomolecules, namely acetylcholinesterase (AChE) and sphingomyelinase (SMase), in a several formats and colors. Acetylcholinesterase (AChE) and sphingomyelinase (SMase) assays are one of the most popular methods for investigating nerve response and function. For more in-depth information on these critical tests see below:

• Acetylcholinesterase (AChE) Assays
• Sphingomyelinase (SMase) Assays

Diagnostic Biomarkers

Specific biomolecules have distinctive changes in concentrations as a symptom of some pathologies. By measuring the changing levels of these materials against known standards, researchers gain insight into physiological conditions. Our Amplite^® kits provide sensitive quantification with a minimal sample size of only 100 µL reaction volume.

Product Ordering Information

---