Studium imunointerakcí pomocí AFM

Bc. Patrik Gereg

Bachelor's thesis

Immunointeractions studied using AFM

Abstract:

The success of atomic force microscopy guarantees in addition to ability of observing objects in nanometer scale, the possibility to quantify the force interaction between biomolecules. These two approaches were used for evaluation of the formation of an immunocomplex between the human serum albumin and the antibody AL-01. At first, immunoreagents’ macromolecules were displayed by scanning the surface, i. e. HSA and AL-01, and then created immunocomplex. A force spectroscopy gave us the information about the force of the immunointeraction between HSA and two types of antibodies: one specific, second non-specific. The technical solution of the measurements was provided by AFM system – Dimension FastScan BIO – by Bruker company. On the scan of the mixture HSA + AL-01, there were occured pairs of two bound biomolecules, which were identified as resulting immunocomplex. Analysis of some selected forces also confirmed a stronger interaction between HSA and AL-01 than between HSA and pure mica or non-specific antibody. However, the identification of individual macromolecules by scanning in the air and in the liquid was not straightforward.

Abstract:

Úspech atómovej silovej mikroskopie zaručuje okrem schopnosti pozorovania objektov v nanometrovej mierke aj možnosť kvantitatívne určiť silové interakcie medzi biomolekulami. Práve tieto dva prístupy sa využili pre vyhodnotenie vzniku imunokomplexu medzi ľudským sérovým albumínom a protilátkou AL 01. Skenovaním povrchu sa zobrazili najskôr samotné makromolekuly imunoreaktantov, t. j. HSA a AL-01, a potom vytvorený imunokomplex. Silová spektroskopia poskytla informácie o sile imunointerakcií, a to medzi HSA a dvomi typmi protilátok: špecifickou a nešpecifickou. Technické riešenie meraní bolo zabezpečené systémom AFM – Dimension FastScan BIO – firmy Bruker. Na snímke so zmesou HSA + AL-01 sa okrem zhlukov nachádzali dvojice viazaných biomolekúl, ktoré sa označili za vzniknutý imunokomplex. Analýza vybraných síl takisto potvrdila silnejšiu interakciu medzi HSA a AL-01 ako medzi HSA a čistou sľudou alebo nešpecifickou protilátkou. Identifikácia jednotlivých makromolekúl pri skenovaní na vzduchu i v kvapaline však nebola jednoznačná.

Keywords

Atómová silová mikroskopia, AFM, Silová spektroskopia, FD-krivky, Imunointerakcia, Imunokomplex, Ľudský sérový albumín, Protilátka AL-01, Atomic Force Microscopy, Force Spectroscopy, FD-curves, Immunointeraction, Immunocomplex, Human Serum Albumin, Antibody AL-01

Language used: Slovak

Date on which the thesis was submitted / produced: 18. 5. 2015

Thesis defence

• Date of defence: 17. 6. 2015
• Supervisor: doc. RNDr. Petr Skládal, CSc.

Masaryk University

Faculty of Science

Bachelor programme / field:
Biochemistry / Biochemistry