Study of the affinity characteristics of monoclonal antibodies by solid-phase immunoenzyme analysis

An approach is proposed for measuring the binding constant (Kb) for monoclonal antibodies (MA) interacting with an immobilized antigen in indirect ELISA. This approach allows the measurement of optical density (A405) in the peroxidase reaction initiated by the conjugate at different concentrations (C0) of antibodies. Using the Scatchard plots, the dependence of A405/C0 = f (A405) for the whole range of MA concentrations was examined, and the tangential of the slope (tg alpha = Kb) of the linear portion of the antigen molecule was calculated. Analysis of MA affinity parameters by using this approach may find wide use in immunodiagnostic studies aimed at measuring antigen and antibody concentrations in biological fluids as well as for estimating the efficiency of vector drugs in which the diagnostic or therapeutic component is conjugated with the vector (MA or F(ab) fragment) responsible for the drug transport to target cells. The method proposed was used for testing mouse (BALB/C) monoclonal antibodies (IgG1) to pig insulin produced by various hybridomas as well as for estimating the effect on MA of pH, temperature and hydrophobization. The minimal detectable concentration (method sensitivity) was found to depend on Kb.     

Inhibition of lactophage activity by quinolinilporphyrin and its zinc compex

The influence of free base of quinolinilporphyrin and its Zn complex on infectivity of lactophages E3, E5 and E17 has been studied. The results of our investigations show that inhibition of lactophage activity by Zn complex of quinolinilporphyrin at concentration 10 microM and 20 microM was in the range 53-62% and 65-85%, respectively. The presence of this porphyrin in nutrient medium prevents the propagation of bacteriophage infection in Lactococcus lactis, but does not affect the phage adsorption process. The free base of quinolinilporphyrin slightly inhibits the activity of lactophages.     

Induction of synthesis and activation of Penicillium commune alpha-L-rhamnosidase

For the first time the influence of a number of synthetic porfirin and fluoren derivatives, coordinative germanium substances and surfactants on the biosynthesis and activity of Penicillium commune 266 alpha-L-rhamnosidase has been studied. It is shown that some of porfirin derivatives and coordinative germanium substances may be used as inducers of biosynthesis (143-430%) and also as stimulators of alpha-L-rhamnosidase activity (15-44%). It is more expedient to use biscitratgermanium acid in a complex with asparaginic acid (430%) or threonine (370%) as inducers of biosynthesis of enzyme. At the same time the use of porfirin derivatives, in particular meso-tetra(N-methyl-6-chinoline)porfirin tosilate or its manganese complex, is more efficient as stimulators (40%) of alpha-L-rhamnosidase activity. The utilization of such surfactants as tregaloso-, peptido- and rhamnolipids is not expedient. These compounds either do not influence or inhibit biosynthesis and activity of P. commune alpha-L-rhamnosidase. Only in concentration of 0.01% rhamnolipid increased the activity of enzyme by 7.5%.     

Kinetics of immunoglobulin G transport through the multi-layer epithelial-hematic barrier of the respiratory tract]

A new class of drugs is now utilized for in vivo diagnoses and therapy of many widespread diseases. In these pharmacological and diagnostic preparations the active substance is conjugated with a vector which transports the drug to specific biological targets. Monoclonal antibodies are the most commonly used vectors: estimation of their permeability through multilayer and unilayer biomembranes is an important step in the analysis of efficiency of vector drugs. Experiments with Sprague-Dawley rats (mature females weighing 500 to 160 g) have demonstrated the ability of immunoglobulins G to penetrate through the respiratory epithelial-hematic barrier. Using solid phase ELISA, it was found that 5-25% of the total amount of mouse antiinsulin immunoglobulins G1 injected into the trachea under hexenal anesthesia can penetrate into the blood plasma. Accumulation of antibodies in the blood begins 4 hours and ceases 32 hours after the drug application in a dose of 400 micrograms. The kinetics of transmembrane transport is described by an S-like saturation function: C(t) = Cmax/(1+e-(at-b]. Penetration of monoclonal antibodies into the blood is accompanied by their distribution in the organs and tissues as well as by their clearance from the blood plasma. The clearance of monoclonal antibodies is characterized by a 24 hour half-life and is described by an exponential equation: C(t) = C0 x exp-kt. An algorithm for the interaction of these processes which should be taken into account during measurements of the transport of monoclonal antibodies and their complexes through biomembranes is proposed.(ABSTRACT TRUNCATED AT 250 WORDS)     

Study of the specificity of the estradiol-binding system in the uteri of guinea pigs]

The authors analyzed the affinity of the steroids belonging to the estran series to the estradiol-binding system of the guinea pig uteri. The presence of free hydroxyl groups in positions 3 (phenol) and 17beta and their interorientation proved to determine the interaction with the recptor system of the guinea pig uterus. The data obtained indicated that the biological activity of the steroids was determined by the peculiarities of their structural interaction with the receptor systems of the uteri.