Antimicrobial proteins (AMP) are a fundamental element of the primary response against pathogens. AMPs are small endogenous cationic molecules expressed by phagocytic and epithelial cells. The antimicrobial activity of AMPs is directed towards a broad spectrum of pathogens, like Gram-positive & negative bacteria, viruses, yeast and fungi. AMPs aid in innate and adaptive immunity via direct inactivation and by immunomodulatory activity like leukocyte migration. RNase 7 is an protein secreted by a variety of epithelial tissues and is a member of the RNase A superfamily. This family shares sequence and structural similarities such as conserved cysteine residues as well as conserved histidines and a lysine in the active center catalysing the ribonuclease activity. RNase 7 is a 14.5 KDa protein with distinct ribonuclease activity. However, the antimicrobial activity is independent form the ribonuclease activity which might be associated with antiviral activity. On a per molar basis, RNase 7 is one of the most potent AMPs. RNase 7 contributes to the sterility in several systems. It is described to be important in sterility of the kidney and urinary tract as well as its contribution to the skin barrier protection. In the urinary system it is constitutively expressed by the intercalated cells in the renal collecting tubules and is present in the urine at such levels to kill bacteria at baseline. In the skin it contributes to control the growth of microorganisms on the skin surface, and the expression levels can be further induced under control of proinflammatory cytokines. The bactericidal activity of RNase 7 has been associated with its ability to bind and permeate the bacterial cell membrane. This requires clustering of lysine residues. RNase 7 is able to bind LPS and peptidoglycans. In ocular surface, signal transduction associated with RNase 7 expression is mediated via MAPKs but not NF-κB signalling pathways. IL1β leads to an increased expression of RNase 7.
The human RNase 7 ELISA kit is to be used for the in vitro quantitative determination of human RNase 7 in serum, plasma, bronchoalveolar lavage fluid, urine and cell culture supernatant samples.
The human RNase 7 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 RNase 7. Biotinylated tracer antibody will bind to the captured human RNase 7. 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 RNase 7 standards (log). The human RNase 7 concentration of samples, which are run concurrently with the standards, can be determined from the standard curve.