ABSTRACT
Complement convertases are enzymatic complexes that play a central role in sustaining and amplification of the complement cascade. Impairment of complement function leads directly or indirectly to pathological conditions, including higher infection rate, kidney diseases, autoimmune- or neurodegenerative diseases and ischaemia-reperfusion injury. An assay for direct measurement of activity of the convertases in patient sera is not available. Existing assays testing convertase function are based on purified complement components and, thus, convertase formation occurs under non-physiological conditions. We designed a new assay, in which C5 blocking compounds enabled separation of the complement cascade into two phases: the first ending at the stage of C5 convertases and the second ending with membrane attack complex formation. The use of rabbit erythrocytes or antibody-sensitized sheep erythrocytes as the platforms for convertase formation enabled easy readout based on measurement of haemolysis. Thus, properties of patient sera could be studied directly regarding convertase activity and membrane attack complex formation. Another advantage of this assay was the possibility to screen for host factors such as C3 nephritic factor and other anti-complement autoantibodies, or gain-of-function mutations, which prolong the half-life of complement convertases. Herein, we present proof of concept, detailed description and validation of this novel assay.
Subject(s)
Complement C3-C5 Convertases/analysis , Erythrocytes/enzymology , Immunoassay/methods , Animals , Autoantibodies/immunology , Complement C3 Nephritic Factor/immunology , Complement C3-C5 Convertases/immunology , Complement Pathway, Alternative/immunology , Complement System Proteins/immunology , Erythrocytes/immunology , Guinea Pigs , Half-Life , Humans , Rabbits , SheepABSTRACT
A film intended for mammography (Kodak SO 155 MRH-1) was tested and compared to one (Kodak SO 177 Ortho M) used earlier. Both films including the cassette could resolve 20 lp/mm. For a processing time of 90 s the new film gave the same overall image quality and irradiation dose to the breast as the old system. If, instead, the processing time was increased to 150 s, a 43% reduction in kerma could be attained with the new film. With a developing temperature of 36 degrees C and 150 s processing time, the noise is clinically acceptable. A nonparametric test showed no significant difference between the 2 films on the 0.01 level. At an X-ray tube potential difference of 25 kV, the mean absorbed dose to a 4.5-cm-thick breast was reduced from 1.7 mGy with the old combination to 1.0 mGy with the new one. The measurements were made with a moving grid.
