Development of a method for detection and quantitative analysis of engeneered endolysin LysAm24-SMAP in biological samples

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Abstract

In recent years modified bacteriophage lysins are widely investigated for the purposes of antibacterial therapy development. Thus, effective and precise methods for the quantitative analysis of these enzymes are of high demand. The enzyme-linked immunosorbent assay (ELISA) method has been developed for the detection of recombinant modified endolysin LysAm24-SMAP in biological samples. The optimal parameters for protein detection were determined, particularly, the influence of salt and the composition of the buffer system for samples preparation was studied. The applicability of the immunodetection system of the genetically engineered endolysin LysAm24-SMAP in various biological samples with enzyme concentrations from 0.4 ng/ml was demonstrated. Also, the influence of matrix effects in animals’ organs and tissues homogenates samples, producer strain lysates and their individual components during the analysis was assessed and it was shown that 0.65 M NaCl addition in the ELISA buffer is crucial for achieving correct results and reduces non-specific interactions in the case of LysAm24-SMAP. The effectiveness of the developed system in the immunochemical control of the bacteriolytic enzyme was confirmed.

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About the authors

A. A. Klimova

Gamaleya National Research Centre for Epidemiology and Microbiology, Ministry of Health of the Russian Federation; Sechenov First Moscow State Medical University (Sechenov University), Ministry of Health of the Russian Federation

Email: northernnatalia@gmail.com
Russian Federation, Moscow, 123098; Moscow, 119991

I. V. Grigoriev

Gamaleya National Research Centre for Epidemiology and Microbiology, Ministry of Health of the Russian Federation

Email: northernnatalia@gmail.com
Russian Federation, Moscow, 123098

D. V. Vasina

Gamaleya National Research Centre for Epidemiology and Microbiology, Ministry of Health of the Russian Federation

Email: northernnatalia@gmail.com
Russian Federation, Moscow, 123098

M. N. Anurova

Sechenov First Moscow State Medical University (Sechenov University), Ministry of Health of the Russian Federation

Email: northernnatalia@gmail.com
Russian Federation, Moscow, 119991

V. A. Gushchin

Gamaleya National Research Centre for Epidemiology and Microbiology, Ministry of Health of the Russian Federation; Lomonosov Moscow State University

Email: northernnatalia@gmail.com
Russian Federation, Moscow, 123098; Moscow, 119991

N. P. Antonova

Gamaleya National Research Centre for Epidemiology and Microbiology, Ministry of Health of the Russian Federation

Author for correspondence.
Email: northernnatalia@gmail.com
Russian Federation, Moscow, 123098

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