Introduction

Proteomic pattern diagnostics is a promising tool for early disease detection and will help to diagnose diseases which are to date hard to detect and to reduce the number of invasive medical procedures, such as biopsies, in the future. The AquaSpec™ Technology is a unique method for the proteomic pattern measurement of aqueous fluids; in particular the acquisition of complex IR - spectra that reflect the status quo in biological fluids like serum, plasma, urine or cerebrospinal fluid. Compared to many other techniques for the measurement of proteomic pattern (e.g. MALDI-TOF), micro-biolytics generates the necessary information from the native state of the biofluid. Therefore, it has the unique and important advantage to detect the proteomic pattern “as is” (including protein interactions with the surrounding). By comparison of data sets from at least two cohorts (e.g. ill / negative controls) using our own bioinformatics package for the discovery of complex biomarker pattern models, micro-biolytics develops the new diagnosis tools. Further, micro-biolytics provides an analyzer with the integrated software ClinSpecPRO for spectra acquisition and diagnosis of the developed biomarkers.

Methods and Instrumentation

Sample preparation and data acquisition. For the development of a specialized diagnosis method, there is a need of a set of samples from each cohort. During the development it is very important to have good sample quality and the complete documentation including the best possible diagnosis of each sample (even the stage of the disease can be important). Therefore all samples have to be taken using the SOP (standard operation procedure) and the storage before measurement should be as short as possible. The samples can be measured without further sample preparation.

Serum, EDTA plasma, urine or cerebrospinal fluid samples obtained from clinicians are thawed, separated into 150µl portions and frozen at -80°C for storage and transportation. For measurement the samples are de-frozen, homogenised and measured by the AquaSpec™ – Analyzer.

Using the complete diagnostic test, the patient’s sample is transported unfrozen in the original tube (e.g. Sarstedt Monovette) and measured directly in the tube by the Analyzer.

Data analysis

There are three steps during the development of a new diagnosis. These steps are always the same for every new diagnosis development and can be used for either a first there is only a feasibility study (few samples of each cohort) or the development of the final diagnostic test (a big dataset describing the variation in total).

• finding the correct features (biomarkers) for the separation of the cohorts: using specialized bioinformatics tools, the spectral information of the cohorts is analyzed in order to find the unique spectral differences between ill and normal samples.
• training of a classificator:by using a number of highly sophisticated mathematical algorithms (chemometric techniques e.g. neural networks, support vector machines etc.) that generate models to differentiate between samples from diseased patients and from controls.
• validation of the biomarkers and the classificator: the trained classificator (optimized model) subsequently is validated by a separate dataset (not used for the development). These results will give the specifity and sensitivity of the test.