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Assay Format

Detection Range:

0.1-1,000 ng/ml

Sensitivity:

4.4 ng/ml

Specificity:

This EIA kit is designed to only detect Cyclic AMP.

Number of Targets Detected:

Species Detected:

Human

Compatible Sample Types:

Plasma
Serum

Design Principle:

Competition-based

Method of Detection:

Colorimetric

Quantitative/Semi-Quantitative:

Quantitative

Solid Support:

96-well Microplate

Product Specifications

Size:

1, 2, or 5 x 96-Well Microplate Kit

Introduction

Cyclic adenosine monophosphate (cAMP, cyclic AMP, or 3',5'-cyclic adenosine monophosphate) is a second messenger important in many biological processes. cAMP is derived from adenosine triphosphate (ATP) and used for intracellular signal transduction in many different organisms, such as transferring into cells the effects of hormones like glucagon and adrenaline, which cannot pass through the plasma membrane. cAMP functions through three main effectors: PKA, the guanine-nucleotide-exchange factor (GEF) EPAC and cyclic-nucleotide-gated ion channels. The intracellular levels of cAMP are regulated by the balance between the activities of two enzymes adenylyl cyclase (AC) and cyclic nucleotide phosphodiesterase (PDE).

Cyclic AMP has been shown to be involved in cell growth, differentiation and general metabolism, and it is important for many biological function, especially in the cardiovascular, nervous and immune systems. The measurement of intracellular Cyclic AMP in tissues and cell cultures may help to provide a clearer understanding of the physiology and pathology of many disease states.

Product Features

Strip plates and additional reagents allow for use in multiple experiments
Quantitative protein detection
Establishes normal range
The best products for confirmation of antibody array data

Application Notes

Kit Components

Pre-Coated 96-well Strip Microplate
Wash Buffer
Standard Peptide
Assay Diluent(s)
Biotinylated Peptide
HRP-Streptavidin
TMB One-Step Substrate
Stop Solution
Assay Diagram
Positive Control Sample
Capture Antibody
User Manual

Other Materials Required

Distilled or deionized water
Precision pipettes to deliver 2 µl to 1 ml volumes
Adjustable 1-25 ml pipettes for reagent preparation
100 ml and 1 liter graduated cylinders
Tubes to prepare standard and sample dilutions
Orbital shaker
Aluminum foil
Saran Wrap
Absorbent paper
Microplate reader capable of measuring absorbance at 450nm
SigmaPlot software (or other software that can perform four-parameter logistic regression models)

Protocol Outline

Prepare all reagents, samples and standards as instructed.
Add 100 µl detection antibody to each well.
Incubate 1.5 h at RT or O/N at 4°C.
Add 100 µl standard or sample to each well.
Incubate 2.5 h at RT.
Add 100 µl prepared streptavidin solution.
Incubate 45 min at RT.
Add 100 µl TMB One-Step Substrate Reagent to each well.
Incubate 30 min at RT.
Add 50 µl Stop Solution to each well.
Read plate at 450 nm immediately.

Storage/Stability

Standard, biotinylated peptide, and positive control should be stored at -20°C after arrival. Avoid multiple freeze-thaws. The remaining kit components may be stored at 4°C. Opened Microplate Wells and antibody (Item N) may be stored for up to 1 month at 2° to 8°C. Return unused wells to the pouch containing desiccant pack and reseal along entire edge.