Alain Theretz

Quantification of specific immunological reactions by atomic force microscopy

In this paper we intend to demonstrate the ability of the Atomic Force Microscope (AFM) to become a tool for the sensitive quantitation of immunological reactions. Anti-ferritin antibodies were adsorbed on silane modified silicon oxyde wafers. The adsorption process was characterized and quantified by RIA, ELISA and AFM, using gold labelled antibodies. A linear correlation was found between the antibody concentration in solution and the number of gold particles, and this method was proved to be more sensitive than classical immunoassays. Finally, the binding reaction between anti-ferritin antibodies and ferritin antigens was studied by AFM. A linear relation was also found between the antigen concentration in bulk solution, and the number of ferritin molecules. The sensitivity threshold for the ferritin detection was 0.06 μg/ml.

Rapid Bacterial Identification, Resistance, Virulence and Type Profiling using Selected Reaction Monitoring Mass Spectrometry

Mass spectrometry (MS) in Selected Reaction Monitoring (SRM) mode is proposed for in-depth characterisation of microorganisms in a multiplexed analysis. Within 60–80 minutes, the SRM method performs microbial identification (I), antibiotic-resistance detection (R), virulence assessment (V) and it provides epidemiological typing information (T). This SRM application is illustrated by the analysis of the human pathogen Staphylococcus aureus, demonstrating its promise for rapid characterisation of bacteria from positive blood cultures of sepsis patients.

Atomic force microscopy as a quantitative technique: correlation between network model approach and experimental study

Abstract Atomic force microscopy (AFM) has been used for imaging and the quantification of immunological reactions. Mouse monoclonal anti-ferritin immunoglobulins (IgG) were adsorbed onto silane-modified and well-characterized silicon wafers. The IgG adsorbed amount was quantified by radio immunoassay (RIA) using 125 I-labeled anti-ferritin antibodies, and the adsorption isotherm was found to follow a Langmuir model. The immunological reaction was then detected by AFM, either using gold labeled polyclonal anti-mouse, or by direct detection of the antigen. A mathematical model based on the network model approach was proposed in order to quantify the immunological reaction. This model was based on the relationship between the roughness induced by the immunological reaction and the number of gold particles or ferritin molecules. The proposed model was found to be well correlated with experimental data for the investigated size of gold particles. The homogeneity of the immunological reaction on the sensitized surface was demonstrated by the investigation of the specifically immobilized distribution of particles on the surface.