Properdin, or factor P, is a single-chain plasma glycoprotein (apparent molecular mass 52-55 kDa), consisting of six thrombospondin repeat sequences between short N- and C-terminal domains. In the blood it exists as a mixture of head-to-tail dimers, trimers and tetramers, with a preponderance of trimers.
The protein is produced by a variety of leukocytes, including monocytes, T lymphocytes and neutrophils , but also by endothelial cells in which properdin synthesis and release are induced by shear stress.
Properdin participates in the alternative pathway of complement activation together with C3 and factors B, D, I and H. It stabilizes the labile C3bBb which is deposited on immune complexes or foreign surfaces. This permits amplification of C3bBb formation in competition with catabolism of C3b by factor I, which uses factor H as a cofactor. The local amplification process leads to the creation of the alternative pathway C5 convertase, C3bBb3b, and initiates the terminal pathway of complement activation.
Properdin prolongs the half-life of surface-bound C3bBb from 1½ minute to about 18 minutes. This is due to several effects: inhibition of C3b cleavage by factor I, increased affinity for factor B and inhibition of the dissociation of C3bBb into C3b and Bb. Properdin is consumed by binding to C3bBb, this binding showing an order of preference of tetramers over trimers over dimers, which is also the order of functional activity of the oligomeric forms.
Deficiency or malfunction of the molecule may lead to severe impairment of alternative pathway activation, depending on the precise nature of the defect. One parameter of functional defect in the presence of measurable levels of protein is an impaired generation of the more active tetramer and trimer forms.
The properdin gene is located on the short arm of the X chromosome, and congenital properdin deficiencies are therefore inherited as typical X-linked recessive traits.
Three types of familial deficiency have been described: type 1 (or I) is characterized by serum with very low or absent properdin activity in hemolytic assays and <0.1 μg/ml immunoreactive protein; type 2 (or II) is characterized by low but detectable levels of immunoreactive protein (»2 μg/ml) and impairment of some, but not all functional test, and type 3 (or III)- has normal levels of immunoreactive but dysfunctional protein. 10 Normal serum or plasma levels of properdin are variously quoted as about 5 or 25 µg/ml. Differences in properdin levels between healthy and diabetic persons (Chaudhry_B_2008)
The human Properdin ELISA kit is to be used for the in vitro quantitative determination of
human Properdin in serum, plasma, bronchoalveolar lavage fluid, urine and cerebrospinal
The human Properdin 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 a plate 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 Properdin.
Biotinylated tracer antibody will bind to the captured human Properdin.
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 Properdin standards (log).
The human Properdin concentration of samples, which are run concurrently with the standards, can be determined from the standard curve.
The linearity of the assay was determined by serially diluting a serum. The line obtained a correlation coefficient of 0.9966
Normal human blood samples (plasma) containing baseline levels of human Properdin, were spiked with human Properdin, in concentrations of 500 and 2000 ng/ml. Samples with and without human Properdin, were incubated for 1 hour at room temperature. Samples were measured using the ELISA. Values for human Properdin, ranged between 85.1% and 115.3%.