Introduction
Resistin is a 12.5 kDa cysteine-rich hormone secreted by adipose tissue. It is also known as XCP-1 (CEBPE regulated myeloid-specific secreted cysteine-rich protein precursor 1), FIZZ3 (found in inflammatory zone 3), or ADSF (adipocyte-specific secretory factor). The length of the resistin is 108 amino acids in humans, and 114 amino acids in mouse and rat; the molecular weight is ~12.5 kDa. Resistin is an adipokine with physiologic role regarding its involvement with obesity and type II diabetes mellitus (T2DM).
Resistin has a high sequence homology among species (43% in a mature protein). Crystal structures of resistin reveal an unusual composition of several subunits that are held together by non-covalent interactions which make up its structure. Each protein subunit comprises a carboxy-terminal disulfide-rich Beta-sandwich "head" domain and an amino-terminal alpha-helical "tail" segment. The globular domain from resistin contains five disulfide bonds.
Some studies have shown the important role of resistin linking obesity to T2DM. The underlying belief among those in support of this theory is that serum resistin levels will increase with increased adiposity. Conversely, serum resistin levels have been found to decline with decreased adiposity following medical treatment. This fact takes on significant implications considering the well understood link between central obesity and insulin resistance; marked peculiarities of T2DM. Furthermore, many studies have shown the positive correlations between resistin levels and insulin resistance, and a direct correlation between resistin levels and subjects with T2DM, indicating that such serum resistin increases are accountable for the insulin resistance apparently associated with increased adiposity. In addition to its role in insulin resistance in obese subjects, resistin also plays a role in inflammation and energy homeostasis.
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 Resistin 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.