Name
SP-D, Human, ELISA kit
Catalog nr
HK335
(lot number and expiry date are indicated on the label)
Short description
Surfactant protein D (SP-D) belongs to the collectin familiy. The lung is the major site of synthesis of SP-D, where the molecules are produced and secreted onto the epithelial surface by alveolar type II cells and unciliated bronchial epithelial cells. SP-D is also found in different epithelial cells of the gastrointestinal tract and in epithelial cells of various exocrine glands. SP-D is an...
Size
2 x 96 det.
Application
Manual
Description
Surfactant protein D (SP-D) belongs to the collectin familiy. These proteins are oligomeric proteins composed of carbohydrate-recognition domains (CRD) attached to collagenous regions. Collectins are structurally similar to the ficolins although they make use of different CRD structures: C-type lectin domain for the collectins. The lung is the major site of synthesis of SP-D, where the molecules are produced and secreted onto the epithelial surface by alveolar type II cells and unciliated bronchial epithelial cells. SP-D is also found in different epithelial cells of the gastrointestinal tract and in epithelial cells of various exocrine glands. SP-D is an important factor in the pulmonary anti-microbial defense. The anti-microbial defense mechanisms of SP-D are direct opsonization, neutralization and agglutination. This results in limiting the infection and concurrently orchestrating the subsequent adaptive immune response. SP-D synthesis and secretion increase significantly during inflammatory stress. In inflamed lungs increased amounts of SP-D in bronchial lavage fluid and tissue are found, particularly in type II pneumocytes, Clara cells and hyperplastic goblet cells are found in inflamed lungs. The localization of SP-D in endocytic vesicles and in lysosomal granules of alveolar macrophages suggests a receptor-mediated uptake. SP-D is also involved the apoptotic process. SP-D binds to apoptotic neutrophils and enhances their clearance by alveolar macrophages. Although complete deficiency of SP-D has not been described in man, basal levels are highly variable and are largely genetically determined. SP-D levels in healthy individuals vary from 200 ng/ml to 5200 ng/ml. Mean SP-D levels increase from 700 ng/ml in children to 1100 ng/ml in adults. Bronchoalveolar fluid SP-D control levels range from 7 ng/ml to 28 µg/ml. SP-D levels can rise dramatically during acute pulmonary infections and are raised in certain non-infectious lung diseases.
The ELISA is not influenced by the presence of lipopolysaccharides.
The ELISA is not influenced by the presence of lipopolysaccharides.
Cross Reactivity
Rat SP-D weak
Species
Human
Features
Working time of 3½ hours.
Minimum concentration which can be measured is 6.3 ng/ml.
Measurable concentration range of 6.3 to 400 ng/ml.
Working volume of 100 µl/well.
Minimum concentration which can be measured is 6.3 ng/ml.
Measurable concentration range of 6.3 to 400 ng/ml.
Working volume of 100 µl/well.
Typical standard curve
Principle
The human SP-D ELISA is a ready-to-use solid-phase enzyme-linked immunosorbent assay based on the sandwich principle with a working time of 3½ hours.
The efficient format of 2 plates with twelve disposable 8-well strips allows free choice of batch size for the assay.
Samples and standards are incubated in microtiter wells coated with antibodies recognizing human SP-D.
Biotinylated tracer antibody will bind to captured human SP-D.
Streptavidin-peroxidase conjugate will bind to the biotinylated tracer antibody.
Streptavidin-peroxidase conjugate will react with the substrate, tetramethylbenzidine (TMB).
The enzyme reaction is stopped by the addition of oxalic acid.
The absorbance at 450 nm is measured with a spectrophotometer. A standard curve is obtained by plotting the absorbance (linear) versus the corresponding concentrations of the human SP-D standards (log).
The human SP-D concentration of samples, which are run concurrently with the standards, can be determined from the standard curve.
The efficient format of 2 plates with twelve disposable 8-well strips allows free choice of batch size for the assay.
Samples and standards are incubated in microtiter wells coated with antibodies recognizing human SP-D.
Biotinylated tracer antibody will bind to captured human SP-D.
Streptavidin-peroxidase conjugate will bind to the biotinylated tracer antibody.
Streptavidin-peroxidase conjugate will react with the substrate, tetramethylbenzidine (TMB).
The enzyme reaction is stopped by the addition of oxalic acid.
The absorbance at 450 nm is measured with a spectrophotometer. A standard curve is obtained by plotting the absorbance (linear) versus the corresponding concentrations of the human SP-D standards (log).
The human SP-D concentration of samples, which are run concurrently with the standards, can be determined from the standard curve.
Storage and stability
Upon receipt, store individual components at 2 - 8°C. Do not freeze.
Do not use components beyond the expiration date printed on the kit label.
The standard, tracer and streptavidin-peroxidase are stable in lyophilized form until the expiration date indicated on the kit label, if stored at 2 - 8°C.
The exact concentration of the standard is indicated on the label of the vial and the certificate of quality control.
Once reconstituted the standard is stable for 6 hours. For longer stability we recommend to store aliquots at –20°C. Stored at –20°C the standard will be stable for 1 month.
Once reconstituted, tracer and streptavidin-peroxidase are stable for 1 month if stored at 2 - 8°C.
Upon receipt, foil pouch around the plate should be vacuum-sealed and unpunctured. Any irregularities to aforementioned conditions may influence plate performance in the assay.
Return unused strips immediately to the foil pouch containing the desiccant pack and reseal along the entire edge of the zip-seal. Quality guaranteed until expiration date if stored at 2 - 8°C.
Do not use components beyond the expiration date printed on the kit label.
The standard, tracer and streptavidin-peroxidase are stable in lyophilized form until the expiration date indicated on the kit label, if stored at 2 - 8°C.
The exact concentration of the standard is indicated on the label of the vial and the certificate of quality control.
Once reconstituted the standard is stable for 6 hours. For longer stability we recommend to store aliquots at –20°C. Stored at –20°C the standard will be stable for 1 month.
Once reconstituted, tracer and streptavidin-peroxidase are stable for 1 month if stored at 2 - 8°C.
Upon receipt, foil pouch around the plate should be vacuum-sealed and unpunctured. Any irregularities to aforementioned conditions may influence plate performance in the assay.
Return unused strips immediately to the foil pouch containing the desiccant pack and reseal along the entire edge of the zip-seal. Quality guaranteed until expiration date if stored at 2 - 8°C.
Recovery
Normal human blood samples (plasma), were spiked with human SP-D in concentrations of 5 and 100 ng/ml. Samples with and without human SP-D were incubated for 1 hour at room temperature. Samples were measured using the ELISA. Values for human SP-D ranged between 83% and 133% (mean 108%).
Linearity
The linearity of the assay was determined by serially diluting a sample containing 250 ng/ml human SP-D. The diluted samples were measured in the assay. The line obtained a slope of 1.08 and a correlation coefficient of 0.999.
Precautions
For research use only. Not for use in or on humans or animals or for diagnostics. It is the responsibility of the user to comply with all local/state and Federal rules in the use of this product. Hycult Biotech is not responsible for any patent infringements that might result with the use of or derivation of this product.
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MSDS
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